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Sample records for administration increases muscarinic

  1. Scopolamine administration modulates muscarinic, nicotinic and NMDA receptor systems.

    PubMed

    Falsafi, Soheil Keihan; Deli, Alev; Höger, Harald; Pollak, Arnold; Lubec, Gert

    2012-01-01

    Studies on the effect of scopolamine on memory are abundant but so far only regulation of the muscarinic receptor (M1) has been reported. We hypothesized that levels of other cholinergic brain receptors as the nicotinic receptors and the N-methyl-D-aspartate (NMDA) receptor, known to be involved in memory formation, would be modified by scopolamine administration.C57BL/6J mice were used for the experiments and divided into four groups. Two groups were given scopolamine 1 mg/kg i.p. (the first group was trained and the second group untrained) in the multiple T-maze (MTM), a paradigm for evaluation of spatial memory. Likewise, vehicle-treated mice were trained or untrained thus serving as controls. Hippocampal levels of M1, nicotinic receptor alpha 4 (Nic4) and 7 (Nic7) and subunit NR1containing complexes were determined by immunoblotting on blue native gel electrophoresis.Vehicle-treated trained mice learned the task and showed memory retrieval on day 8, while scopolamine-treatment led to significant impairment of performance in the MTM. At the day of retrieval, hippocampal levels for M1, Nic7 and NR1 were higher in the scopolamine treated groups than in vehicle-treated groups.The concerted action, i.e. the pattern of four brain receptor complexes regulated by the anticholinergic compound scopolamine, is shown. Insight into probable action mechanisms of scopolamine at the brain receptor complex level in the hippocampus is provided. Scopolamine treatment is a standard approach to test cognitive enhancers and other psychoactive compounds in pharmacological studies and therefore knowledge on mechanisms is of pivotal interest.

  2. Selective increase of alpha 1-adrenoceptors and muscarinic cholinergic receptors in rat cerebral cortex after chronic haloperidol.

    PubMed

    Pazo, J H; Levi de Stein, M; Jerusalinsky, D; Novas, M L; Raskovsky, S; Tumilasci, O R; Medina, J H; De Robertis, E

    1987-06-30

    The effect of chronic administration of haloperidol on alpha 1-, alpha 2-, and beta-adrenoceptors, cholinergic muscarinic, GABAA and benzodiazepine receptors in the cerebral cortex of the rat was investigated. Doses of 0.3 and 2 mg/kg of haloperidol during 7 days increased markedly the density of alpha 1-adrenoceptors without changes in affinity. The alpha 2- and beta-adrenoceptors were not modified after neuroleptic administration. The number of muscarinic receptors were also increased after haloperidol treatment (2 mg/kg/day). However, the GABAA and benzodiazepine binding sites remained unchanged. In the brainstem an increment in the alpha 1-, but not the beta-adrenoceptors was observed. The well known increase in the dopamine receptors in the striatum was confirmed. These observations demonstrate a multireceptor effect of haloperidol in the cerebral cortex.

  3. Development of tolerance after repeated administration of a selective muscarinic M1 antagonist biperiden in healthy human volunteers.

    PubMed

    Salin-Pascual, R J; Granados-Fuentes, D; Galicia-Polo, L; Nieves, E; Gillin, J C

    1993-02-01

    The muscarinic antagonist biperiden produces a dose-dependent inhibition of (REM) sleep on acute administration. The present study addressed the possibility of pharmacological tolerance after repeated biperiden administration. Six healthy volunteers were studied under sleep laboratory conditions in the following situations: one acclimatization, night, two baseline (that were averaged), 4 nights of biperiden administration, and 4 nights of placebo recovery administration. Six milligrams of biperiden and placebo were administered in identical capsules. Volunteers and technicians were blind to the order of the administration of the capsules. REM sleep time was reduced during the first and the second night, but was not significantly different in comparison with baseline by the third night. During placebo recovery nights, REM sleep time was not different from baseline. REM sleep latency was increased during the first and second nights of biperiden administration, but tolerance to this effect was observed by the third night. On placebo nights a dramatic shortening of REM latency was observed. The present findings support the hypothesis that anticholinergic drugs, even a selective M1 antagonist such as biperiden, induce tolerance soon after administration. A similar effect has been reported with scopolamine, a nonselective muscarinic antagonist, but the main difference is that biperiden withdrawal was not followed by an REM sleep rebound. The observed effect on REM sleep latency during placebo administration may be related to a supersensitivity to muscarinic M-1 receptors that trigger the first REM sleep period. Because short REM latency has been the main finding in the sleep of depressed patients, some implications of the present findings are discussed.

  4. Hypoxia increases exercise heart rate despite combined inhibition of β-adrenergic and muscarinic receptors.

    PubMed

    Siebenmann, C; Rasmussen, P; Sørensen, H; Bonne, T C; Zaar, M; Aachmann-Andersen, N J; Nordsborg, N B; Secher, N H; Lundby, C

    2015-06-15

    Hypoxia increases the heart rate response to exercise, but the mechanism(s) remains unclear. We tested the hypothesis that the tachycardic effect of hypoxia persists during separate, but not combined, inhibition of β-adrenergic and muscarinic receptors. Nine subjects performed incremental exercise to exhaustion in normoxia and hypoxia (fraction of inspired O2 = 12%) after intravenous administration of 1) no drugs (Cont), 2) propranolol (Prop), 3) glycopyrrolate (Glyc), or 4) Prop + Glyc. HR increased with exercise in all drug conditions (P < 0.001) but was always higher at a given workload in hypoxia than normoxia (P < 0.001). Averaged over all workloads, the difference between hypoxia and normoxia was 19.8 ± 13.8 beats/min during Cont and similar (17.2 ± 7.7 beats/min, P = 0.95) during Prop but smaller (P < 0.001) during Glyc and Prop + Glyc (9.8 ± 9.6 and 8.1 ± 7.6 beats/min, respectively). Cardiac output was enhanced by hypoxia (P < 0.002) to an extent that was similar between Cont, Glyc, and Prop + Glyc (2.3 ± 1.9, 1.7 ± 1.8, and 2.3 ± 1.2 l/min, respectively, P > 0.4) but larger during Prop (3.4 ± 1.6 l/min, P = 0.004). Our results demonstrate that the tachycardic effect of hypoxia during exercise partially relies on vagal withdrawal. Conversely, sympathoexcitation either does not contribute or increases heart rate through mechanisms other than β-adrenergic transmission. A potential candidate is α-adrenergic transmission, which could also explain why a tachycardic effect of hypoxia persists during combined β-adrenergic and muscarinic receptor inhibition.

  5. Deletion of M1 muscarinic acetylcholine receptors increases amyloid pathology in vitro and in vivo

    PubMed Central

    Davis, Albert A.; Fritz, Jason J.; Wess, Jürgen; Lah, James J.; Levey, Allan I.

    2010-01-01

    Alzheimer's disease (AD) is a progressive neurological disorder that causes dementia and poses a major public health crisis as the population ages. Aberrant processing of the amyloid precursor protein (APP) is strongly implicated as a proximal event in AD pathophysiology, but the neurochemical signals that regulate APP processing in the brain are not completely understood. Activation of muscarinic acetylcholine receptors (mAChRs) has been shown to affect APP processing and AD pathology, but less is known about the roles of specific mAChR subtypes. In this study, we used M1 mAChR knockout mice (M1KO) to isolate the effects of the M1 mAChR on APP processing in primary neurons and on the development of amyloid pathology in a transgenic mouse model of AD. We demonstrate that the loss of M1 mAChRs increases amyloidogenic APP processing in neurons, as evidenced by decreased agonist-regulated shedding of the neuroprotective APP ectodomain APPsα and increased production of toxic Aβ peptides. Expression of M1 mAChRs on the M1KO background rescued this phenotype, indicating that M1 mAChRs are sufficient to modulate non-amyloidogenic APP processing. In APPSwe/Ind transgenic mice, the loss of M1 mAChRs resulted in increased levels of brain Aβ1-40 and greater accumulation of amyloid plaque pathology. Analysis of APP metabolites in APPSwe/Ind brain tissue indicates that the loss of M1 mAChRs increases amyloidogenic APP processing. These results indicate that the M1 mAChR is an important regulator of amyloidogenesis in the brain and provide strong support for targeting the M1 mAChR as a therapeutic candidate in AD. PMID:20335454

  6. Muscarinic receptor binding increases in anterior thalamus and cingulate cortex during discriminative avoidance learning

    SciTech Connect

    Vogt, B.A.; Gabriel, M.; Vogt, L.J.; Poremba, A.; Jensen, E.L.; Kubota, Y.; Kang, E. )

    1991-06-01

    Training-induced neuronal activity develops in the mammalian limbic system during discriminative avoidance conditioning. This study explores behaviorally relevant changes in muscarinic ACh receptor binding in 52 rabbits that were trained to one of five stages of conditioned response acquisition. Sixteen naive and 10 animals yoked to criterion performance served as control cases. Upon reaching a particular stage of training, the brains were removed and autoradiographically assayed for 3H-oxotremorine-M binding with 50 nM pirenzepine (OxO-M/PZ) or for 3H-pirenzepine binding in nine limbic thalamic nuclei and cingulate cortex. Specific OxO-M/PZ binding increased in the parvocellular division of the anterodorsal nucleus early in training when the animals were first exposed to pairing of the conditional and unconditional stimuli. Elevated binding in this nucleus was maintained throughout subsequent training. In the parvocellular division of the anteroventral nucleus (AVp), OxO-M/PZ binding progressively increased throughout training, reached a peak at the criterion stage of performance, and returned to control values during extinction sessions. Peak OxO-M/PZ binding in AVp was significantly elevated over that for cases yoked to criterion performance. In the magnocellular division of the anteroventral nucleus (AVm), OxO-M/PZ binding was elevated only during criterion performance of the task, and it was unaltered in any other limbic thalamic nuclei. Specific OxO-M/PZ binding was also elevated in most layers in rostral area 29c when subjects first performed a significant behavioral discrimination. Training-induced alterations in OxO-M/PZ binding in AVp and layer Ia of area 29c were similar and highly correlated.

  7. Effects of acute administration of nicotinic and muscarinic cholinergic agonists and antagonists on performance in different cost–benefit decision making tasks in rats

    PubMed Central

    Mendez, Ian A.; Gilbert, Ryan J.; Bizon, Jennifer L.

    2012-01-01

    Rationale Alterations in cost–benefit decision making accompany numerous neuropsychiatric conditions, including schizophrenia, attention deficit hyperactivity disorder, and addiction. Central cholinergic systems have been linked to the etiology and/or treatment of many of these conditions, but little is known about the role of cholinergic signaling in cost–benefit decision making. Objectives The goal of these experiments was to determine how cholinergic signaling is involved in cost–benefit decision making, using a behavioral pharmacological approach. Methods Male Long-Evans rats were trained in either “probability discounting” or “delay discounting” tasks, in which rats made discrete-trial choices between a small food reward and a large food reward associated with either varying probabilities of omission or varying delays to delivery, respectively. The effects of acute administration of different doses of nicotinic and muscarinic acetylcholine receptor agonists and antagonists were assessed in each task. Results In the probability discounting task, acute nicotine administration (1.0 mg/kg) significantly increased choice of the large risky reward, and control experiments suggested that this was due to robust nicotine-induced impairments in behavioral flexibility. In the delay discounting task, the muscarinic antagonists scopolamine (0.03, 0.1, and 0.3 mg/kg) and atropine (0.3 mg/kg) both significantly increased choice of the small immediate reward. Neither mecamylamine nor oxotremorine produced reliable effects on either of the decision making tasks. Conclusions These data suggest that cholinergic receptors play multiple roles in decision making contexts which include consideration of reward delay or probability. These roles should be considered when targeting these receptors for therapeutic purposes. PMID:22760484

  8. Activation of muscarinic receptors in porcine airway smooth muscle elicits a transient increase in phospholipase D activity.

    PubMed

    Mamoon, A M; Smith, J; Baker, R C; Farley, J M

    1999-01-01

    Phospholipase D (PLD) is a phosphodiesterase that catalyses hydrolysis of phosphatidylcholine to produce phosphatidic acid and choline. In the presence of ethanol, PLD also catalyses the formation of phosphatidylethanol, which is a unique characteristic of this enzyme. Muscarinic receptor-induced changes in the activity of PLD were investigated in porcine tracheal smooth muscle by measuring the formation of [3H]phosphatidic acid ([3H]PA) and [3H]phosphatidylethanol ([3H]PEth) after labeling the muscle strips with [3H]palmitic acid. The cholinergic receptor agonist acetylcholine (Ach) significantly but transiently increased formation of both [3H]PA and [3H]PEth in a concentration-dependent manner (>105-400% vs. controls in the presence of 10(-6) to 10(-4) M Ach) when pretreated with 100 mM ethanol. The Ach receptor-mediated increase in PLD activity was inhibited by atropine (10(-6) M), indicating that activation of PLD occurred via muscarinic receptors. Activation of protein kinase C (PKC) by phorbol-12-myristate-13-acetate (PMA) increased PLD activity that was effectively blocked by the PKC inhibitors calphostin C (10(-8) to 10(-6) M) and GFX (10(-8) to 10(-6) M). Ach-induced increases in PLD activity were also significantly, but incompletely, inhibited by both GFX and calphostin C. From the present data, we conclude that in tracheal smooth muscle, muscarinic acetylcholine receptor-induced PLD activation is transient in nature and coupled to these receptors via PKC. However, PKC activation is not solely responsible for Ach-induced activation of PLD in porcine tracheal smooth muscle.

  9. Increasing Faculty and Administrative Effectiveness.

    ERIC Educational Resources Information Center

    Heller, Jack F.

    A unified approach to identifying and solving the fundamental problems that prevent colleges and universities from responding effectively to new demands is presented. It demonstrates the underlying causes of institutional problems using case study examples of administrators and faculty members in their institutions. Chapter 1, "Current Approaches…

  10. Brain regional acetylcholinesterase activity and muscarinic acetylcholine receptors in rats after repeated administration of cholinesterase inhibitors and its withdrawal

    SciTech Connect

    Kobayashi, Haruo . E-mail: hk1664@iwate-u.ac.jp; Suzuki, Tadahiko; Sakamoto, Maki; Hashimoto, Wataru; Kashiwada, Keiko; Sato, Itaru; Akahori, Fumiaki; Satoh, Tetsuo

    2007-03-15

    Activity of acetylcholinesterase (AChE) and specific binding of [{sup 3}H]quinuclidinyl benzilate (QNB), [{sup 3}H]pirenzepine (PZP) and [{sup 3}H]AF-DX 384 to muscarinic acetylcholine receptor (mAChR) preparations in the striatum, hippocampus and cortex of rats were determined 1, 6 and 11 days after the last treatment with an organophosphate DDVP, a carbamate propoxur or a muscarinic agonist oxotremorine as a reference for 7 and 14 days. AChE activity was markedly decreased in the three regions 1 day after the treatment with DDVP for 7 and 14 days with a gradual recovery 6 to 11 days, and much less decreased 1, 6 and 11 days after the treatment with propoxur for 7 days but not for 14 days in the hippocampus and cortex. The binding of [{sup 3}H]-QNB, PZP and AF-DX 384 in the three regions was generally decreased by the treatment with DDVP for 7 and 14 days. Such down-regulations were generally restored 6 or 11 days after the treatment for 7 but not for 14 days. The down-regulation or up-regulation as measured by [{sup 3}H]-QNB, PZP and AF-DX 384 was observed 1, 6 or 11 days after treatment with propoxur for 7 days and/or 14 days. Repeated treatment with oxotremorine produced similar effects except AChE activity to DDVP. These results suggest that repeated inhibition of AChE activity may usually cause down-regulation of mAChRs with some exception in the hippocampus when a reversible antiChE propoxur is injected.

  11. Increase in muscarinic stimulation-induced Ca(2+) response by adenovirus-mediated Stim1-mKO1 gene transfer to rat submandibular acinar cells in vivo.

    PubMed

    Morita, Takao; Nezu, Akihiro; Tojyo, Yosuke; Tanimura, Akihiko

    2013-10-01

    Adenoviruses have been used for gene transfer to salivary gland cells in vivo. Their use to study the function of salivary acinar cells was limited by a severe inflammatory response and by the destruction of fluid-secreting acinar cells. In the present study, low doses of adenovirus were administered to express Stim1-mKO1 by retrograde ductal injection to submandibular glands. The approach succeeded in increasing muscarinic stimulation-induced Ca(2+) responses in acinar cells without inflammation or decreased salivary secretions. This increased Ca(2+) response was notable upon weak muscarinic stimulation and was attributed to increased Ca(2+) release from internal stores and increased Ca(2+) entry. The basal Ca(2+) level was higher in Stim1-mKO1-expressing cells than in mKO1-expressing and non-expressing cells. Exposure of permeabilized submandibular acinar cells, where Ca(2+) concentration was fixed at 50 nM, to inositol 1,4,5-trisphosphate (IP3) produced similar effects on the release of Ca(2+) from stores in Stim1-mKO1-expressing and non-expressing cells. The low toxicity and relative specificity to acinar cells of the mild gene transfer method described herein are particularly useful for studying the molecular functions of salivary acinar cells in vivo, and may be applied to increase salivary secretions in experimental animals and human in future.

  12. Why Block Grants Should Increase Administrative Costs.

    ERIC Educational Resources Information Center

    Baker, Keith

    1983-01-01

    Federal education programs increase costs because they attach fewer strings to funds than state or local grants, and this is likely to lead to administrative empire-building. Bureaucracy tends pathologically as it grows to generate more work for itself independent of true administrative needs. Some policy implications are drawn. (MJL)

  13. Contractions of the mouse prostate elicited by acetylcholine are mediated by M(3) muscarinic receptors.

    PubMed

    White, Carl W; Short, Jennifer L; Haynes, John M; Matsui, Minoru; Ventura, Sabatino

    2011-12-01

    Increased smooth muscle tone in the human prostate contributes to the symptoms associated with benign prostatic hyperplasia. In the mouse prostate gland, cholinergic innervation is responsible for a component of the nerve-mediated contractile response. This study investigates the muscarinic receptor subtype responsible for the cholinergic contractile response in the mouse prostate gland. To characterize the muscarinic receptor subtype, mouse prostates taken from wild-type or M(3) muscarinic receptor knockout mice were mounted in organ baths. The isometric force that tissues developed in response to electrical-field stimulation or exogenously applied cholinergic agonists in the presence or absence of a range of muscarinic receptor antagonists was evaluated. Carbachol elicited reproducible and concentration-dependent contractions of the isolated mouse prostate, which were antagonized by the presence of muscarinic receptor antagonists. Calculation of antagonist affinities (pA(2) values) indicated a rank order of antagonist potencies in the mouse prostate of: darifenacin (9.08) = atropine (9.07) = 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (9.02) > cyclohexyl-hydroxy-phenyl-(3-piperidin-1-ylpropyl)silane (7.85) > cyclohexyl-(4-fluorophenyl)-hydroxy-(3-piperidin-1-ylpropyl)silane (7.39) > himbacine (7.19) > pirenzipine (6.88) > methoctramine (6.20). Furthermore, genetic deletion of the M(3) muscarinic receptor inhibited prostatic contractions to electrical-field stimulation or exogenous administration of acetylcholine. In this study we identified that the cholinergic component of contraction in the mouse prostate is mediated by the M(3) muscarinic receptor subtype. Pharmacological antagonism of the M(3) muscarinic receptor may be a beneficial additional target for the treatment of benign prostatic hyperplasia in the human prostate gland.

  14. Impaired recovery of brain muscarinic receptor sites following an adaptive down-regulation induced by repeated administration of diisopropyl fluorophosphate in aged rats

    SciTech Connect

    Pintor, A.; Fortuna, S.; De Angelis, S.; Michalek, H. )

    1990-01-01

    Potential age-related differences in the recovery rate of brain cholinesterase activity (ChE) and muscarinic acetylcholine receptor binding sites (mAChRs) following reduction induced by repeated treatment with diisopropyl fluorophosphate (DFP) were evaluated in Sprague-Dawley rats. Male 3- and 24-month old rats were s.c. injected with DFP on alternate days for 2 weeks and killed 48 hr and 7, 14, 21, 28 and 35 days after the last treatment. In the hippocampus and striatum, but not in the cerebral cortex, of control rats there as a significant age-related decline of ChE activity and maximal density of 3H-QNB binding sites (Bmax). The repeated administration of DFP during the first week caused a syndrome of cholinergic stimulation both in aged and young rats. The syndrome was more pronounced, in terms of intensity and duration in aged than in young animals resulting in 40 and 12% mortality, respectively; during the second week the syndrome attenuated in the two age-groups. The percentage inhibition of brain ChE at the end of DFP treatment did not differ between young and surviving aged rats. The down-regulation of mACRs was present in the three brain regions of both young and age rats (from 20 to 40%). Factorial analysis of variance showed significant differences for age, recovery rate, and significant interaction between age and recovery rate, both for ChE and mAChRs in young rats the three brain areas.

  15. Intrahippocampal administration of anandamide increases REM sleep.

    PubMed

    Rueda-Orozco, Pavel Ernesto; Soria-Gómez, Edgar; Montes-Rodríguez, Corinne Jennifer; Pérez-Morales, Marcel; Prospéro-García, Oscar

    2010-04-01

    A nascent literature has postulated endocannabinoids (eCBs) as strong sleep-inducing lipids, particularly rapid-eye-movement sleep (REMs), nevertheless the exact mechanisms behind this effect remain to be determined. Anandamide and 2-arachidonyl glycerol, two of the most important eCBS, are synthesized in the hippocampus. This structure also expresses a high concentration of cannabinoid receptor 1 (CB1). Recent extensive literature supports eCBs as important regulators of hippocampal activity. It has also been shown that these molecules vary their expression on the hippocampus depending on the light-dark cycle. In this context we decided to analyze the effect of intrahippocampal administration of the eCB anandamide (ANA) on the sleep-waking cycle at two points of the light-dark cycle. Our data indicate that the administration of ANA directly into the hippocampus increases REMs in a dose dependent manner during the dark but not during the light phase of the cycle. The increase of REMs was blocked by the CB1 antagonist AM251. This effect was specific for the hippocampus since ANA administrations in the surrounding cortex did not elicit any change in REMs. These results support the idea of a direct relationship between hippocampal activity and sleep mechanisms by means of eCBs. The data presented here show, for the first time that eCBs administered into the hippocampus trigger REMs and support previous studies where chemical stimulation of limbic areas triggered sleep.

  16. Subcutaneous administration of muscarinic antagonists and triple-immunostaining of the levator auris longus muscle in mice.

    PubMed

    Wright, Megan; Kim, Amy; Son, Young-Jin

    2011-09-08

    Hind limb muscles of rodents, such as gastrocnemius and tibialis anterior, are frequently used for in vivo pharmacological studies of the signals essential for the formation and maintenance of mammalian NMJs. However, drug penetration into these muscles after subcutaneous or intramuscular administration is often incomplete or uneven and many NMJs can remain unaffected. Although systemic administration with devices such as mini-pumps can improve the spatiotemporal effects, the invasive nature of this approach can cause confounding inflammatory responses and/or direct muscle damage. Moreover, complete analysis of the NMJs in a hind limb muscle is challenging because it requires time-consuming serial sectioning and extensive immunostaining. The mouse LAL is a thin, flat sheet of muscle located superficially on the dorsum of the neck. It is a fast-twitch muscle that functions to move the pinna. It contains rostral and caudal portions that originate from the midline of the cranium and extend laterally to the cartilaginous portion of each pinna. The muscle is supplied by a branch of the facial nerve that projects caudally as it exits the stylomastoid foramen. We and others have found LAL to be a convenient preparation that offers advantages for the investigation of both short and long-term in vivo effects of drugs on NMJs and muscles. First, its superficial location facilitates multiple local applications of drugs under light anesthesia. Second, its thinness (2-3 layers of muscle fibers) permits visualization and analysis of almost all the NMJs within the muscle. Third, the ease of dissecting it with its nerve intact together with the pattern of its innervation permits supplementary electrophysiological analysis in vitro. Last, and perhaps most importantly, a small applied volume (-50 μl) easily covers the entire muscle surface, provides a uniform and prolonged exposure of all its NMJs to the drug and eliminates the need for a systemic approach.

  17. Attenuation of Cocaine's Reinforcing and Discriminative Stimulus Effects via Muscarinic M1 Acetylcholine Receptor StimulationS⃞

    PubMed Central

    Conn, P. Jeffrey; Lindsley, Craig; Wess, Jürgen; Boon, Joon Y.; Fulton, Brian S.; Fink-Jensen, Anders; Caine, S. Barak

    2010-01-01

    Muscarinic cholinergic receptors modulate dopaminergic function in brain pathways thought to mediate cocaine's abuse-related effects. Here, we sought to confirm and extend in the mouse species findings that nonselective muscarinic receptor antagonists can enhance cocaine's discriminative stimulus. More importantly, we tested the hypothesis that muscarinic receptor agonists with varied receptor subtype selectivity can blunt cocaine's discriminative stimulus and reinforcing effects; we hypothesized a critical role for the M1 and/or M4 receptor subtypes in this modulation. Mice were trained to discriminate cocaine from saline, or to self-administer intravenous cocaine chronically. The nonselective muscarinic antagonists scopolamine and methylscopolamine, the nonselective muscarinic agonists oxotremorine and pilocarpine, the M1/M4-preferring agonist xanomeline, the putative M1-selective agonist (4-hydroxy-2-butynyl)-1-trimethylammonium-3-chlorocarbanilate chloride (McN-A-343), and the novel M1-selective agonist 1-(1-2-methylbenzyl)-1,4-bipiperidin-4-yl)-1H benzo[d]imidazol-2(3H)-one (TBPB) were tested as substitution and/or pretreatment to cocaine. Both muscarinic antagonists partially substituted for cocaine and enhanced its discriminative stimulus. Conversely, muscarinic agonists blunted cocaine discrimination and abolished cocaine self-administration with varying effects on food-maintained behavior. Specifically, increasing selectivity for the M1 subtype (oxotremorine < xanomeline < TBPB) conferred lesser nonspecific rate-suppressing effects, with no rate suppression for TBPB. In mutant mice lacking M1 and M4 receptors, xanomeline failed to diminish cocaine discrimination while rate-decreasing effects were intact. Our data suggest that central M1 receptor activation attenuates cocaine's abuse-related effects, whereas non-M1/M4 receptors probably contribute to undesirable effects of muscarinic stimulation. These data provide the first demonstration of anticocaine

  18. A long-acting β2-adrenergic agonist increases the expression of muscarine cholinergic subtype‑3 receptors by activating the β2-adrenoceptor cyclic adenosine monophosphate signaling pathway in airway smooth muscle cells.

    PubMed

    Liu, Yuan-Hua; Wu, Song-Ze; Wang, Gang; Huang, Ni-Wen; Liu, Chun-Tao

    2015-06-01

    The persistent administration of β2‑adrenergic (β2AR) agonists has been demonstrated to increase the risk of severe asthma, partly due to the induction of tolerance to bronchoprotection via undefined mechanisms. The present study investigated the potential effect of the long‑acting β2‑adrenergic agonist, formoterol, on the expression of muscarinic M3 receptor (M3R) in rat airway smooth muscle cells (ASMCs). Primary rat ASMCs were isolated and characterized following immunostaining with anti‑α‑smooth muscle actin antibodies. The protein expression levels of M3R and phospholipase C‑β1 (PLCβ1) were characterized by western blot analysis and the production of inositol 1,4,5‑trisphosphate (IP3) was determined using an enzyme‑linked immunosorbent assay. Formoterol increased the protein expression of M3R in rat ASMCs in a time‑ and dose‑dependent manner, which was significantly inhibited by the β2AR antagonist, ICI118,551 and the cyclic adenosine monophosphate (cAMP) inhibitor, SQ22,536. The increased protein expression of M3R was positively correlated with increased production of PLCβ1 and IP3. Furthermore, treatment with the glucocorticoid, budesonide, and the PLC inhibitor, U73,122, significantly suppressed the formoterol‑induced upregulated protein expression levels of M3R and PLCβ1 and production of IP3. The present study demonstrated that formoterol mediated the upregulation of M3R in the rat ASMCs by activating the β2AR‑cAMP signaling pathway, resulting in increased expression levels of PLCβ1 and IP3, which are key to inducing bronchoprotection tolerance. Administration of glucocorticoids or a PLC antagonist prevented formoterol‑induced bronchoprotection tolerance by suppressing the protein expression of M3R. PMID:25672589

  19. A long-acting β2-adrenergic agonist increases the expression of muscarine cholinergic subtype-3 receptors by activating the β2-adrenoceptor cyclic adenosine monophosphate signaling pathway in airway smooth muscle cells

    PubMed Central

    LIU, YUAN-HUA; WU, SONG-ZE; WANG, GANG; HUANG, NI-WEN; LIU, CHUN-TAO

    2015-01-01

    The persistent administration of β2-adrenergic (β2AR) agonists has been demonstrated to increase the risk of severe asthma, partly due to the induction of tolerance to bronchoprotection via undefined mechanisms. The present study investigated the potential effect of the long-acting β2-adrenergic agonist, formoterol, on the expression of muscarinic M3 receptor (M3R) in rat airway smooth muscle cells (ASMCs). Primary rat ASMCs were isolated and characterized following immunostaining with anti-α-smooth muscle actin antibodies. The protein expression levels of M3R and phospholipase C-β1 (PLCβ1) were characterized by western blot analysis and the production of inositol 1,4,5-trisphosphate (IP3) was determined using an enzyme-linked immunosorbent assay. Formoterol increased the protein expression of M3R in rat ASMCs in a time- and dose-dependent manner, which was significantly inhibited by the β2AR antagonist, ICI118,551 and the cyclic adenosine monophosphate (cAMP) inhibitor, SQ22,536. The increased protein expression of M3R was positively correlated with increased production of PLCβ1 and IP3. Furthermore, treatment with the glucocorticoid, budesonide, and the PLC inhibitor, U73,122, significantly suppressed the formoterol-induced upregulated protein expression levels of M3R and PLCβ1 and production of IP3. The present study demonstrated that formoterol mediated the upregulation of M3R in the rat ASMCs by activating the β2AR-cAMP signaling pathway, resulting in increased expression levels of PLCβ1 and IP3, which are key to inducing bronchoprotection tolerance. Administration of glucocorticoids or a PLC antagonist prevented formoterol-induced bronchoprotection tolerance by suppressing the protein expression of M3R. PMID:25672589

  20. Modulation of mood and cognitive performance following acute administration of single doses of Melissa officinalis (Lemon balm) with human CNS nicotinic and muscarinic receptor-binding properties.

    PubMed

    Kennedy, D O; Wake, G; Savelev, S; Tildesley, N T J; Perry, E K; Wesnes, K A; Scholey, A B

    2003-10-01

    Melissa officinalis (Lemon balm) is a herbal medicine that has traditionally been attributed with memory-enhancing properties, but which is currently more widely used as a mild sedative and sleep aid. In a previous study it was demonstrated that a commercial Melissa extract led to dose-specific increases in calmness, and dose-dependent decrements in timed memory task performance. However, the extract utilized in that study did not exhibit in vitro cholinergic receptor-binding properties. The current study involved an initial screening of samples of M. officinalis for human acetylcholinesterase inhibition and cholinergic receptor-binding properties. The cognitive and mood effects of single doses of the most cholinergically active dried leaf were then assessed in a randomized, placebo-controlled, double-blind, balanced crossover study. Following the in vitro analysis, 20 healthy, young participants received single doses of 600, 1000, and 1600 mg of encapsulated dried leaf, or a matching placebo, at 7-day intervals. Cognitive performance and mood were assessed predose and at 1, 3, and 6 h postdose using the Cognitive Drug Research computerized assessment battery and Bond-Lader visual analog scales, respectively. In vitro analysis of the chosen extract established IC(50) concentrations of 0.18 and 3.47 mg ml(-1), respectively, for the displacement of [(3)H]-(N)-nicotine and [(3)H]-(N)-scopolamine from nicotinic and muscarinic receptors in the human cerebral cortex tissue. However, no cholinesterase inhibitory properties were detected. The most notable cognitive and mood effects were improved memory performance and increased 'calmness' at all postdose time points for the highest (1600 mg) dose. However, while the profile of results was overwhelmingly favorable for the highest dose, decrements in the speed of timed memory task performance and on a rapid visual information-processing task increased with decreasing dose. These results suggest that doses of Melissa

  1. Immunochemical studies of the muscarinic acetylcholine receptor.

    PubMed

    André, C; Marullo, S; Guillet, J G; Convents, A; Lauwereys, M; Kaveri, S; Hoebeke, J; Strosberg, A D

    1987-01-01

    Muscarinic receptors have been purified from calf forebrain plasma cell membranes by affinity chromatography on a dexetimide-agarose gel. SDS-PAGE analysis showed a single 70 kDa band. Monoclonal antibodies have been prepared against these affinity purified 70 kDa protein(s). One antibody, M-35, immunoprecipitated up to 80% of digitonin-solubilized muscarinic receptors. M-35 had agonist-like effects on guinea-pig myometrium: it increased the intracellular cyclic GMP content, decreased prostaglandin-induced cyclic AMP accumulation and caused muscle contractions. The two first effects were inhibited by atropine. M-35 was used to visualize muscarinic receptors at the surface of human fibroblastic cells. In the particular cell line used, the receptors have a low affinity for pirenzepine, were negatively coupled to adenylate cyclase and mediated increase in the phosphatidyl-inositol breakdown. PMID:3040987

  2. Activation of muscarinic acetylcholine receptors via their allosteric binding sites.

    PubMed Central

    Jakubík, J; Bacáková, L; Lisá, V; el-Fakahany, E E; Tucek, S

    1996-01-01

    Ligands that bind to the allosteric-binding sites on muscarinic acetylcholine receptors alter the conformation of the classical-binding sites of these receptors and either diminish or increase their affinity for muscarinic agonists and classical antagonists. It is not known whether the resulting conformational change also affects the interaction between the receptors and the G proteins. We have now found that the muscarinic receptor allosteric modulators alcuronium, gallamine, and strychnine (acting in the absence of an agonist) alter the synthesis of cAMP in Chinese hamster ovary (CHO) cells expressing the M2 or the M4 subtype of muscarinic receptors in the same direction as the agonist carbachol. In addition, most of their effects on the production of inositol phosphates in CHO cells expressing the M1 or the M3 muscarinic receptor subtypes are also similar to (although much weaker than) those of carbachol. The agonist-like effects of the allosteric modulators are not observed in CHO cells that have not been transfected with the gene for any of the subtypes of muscarinic receptors. The effects of alcuronium on the formation of cAMP and inositol phosphates are not prevented by the classical muscarinic antagonist quinuclidinyl benzilate. These observations demonstrate for the first time that the G protein-mediated functional responses of muscarinic receptors can be evoked not only from their classical, but also from their allosteric, binding sites. This represents a new mechanism of receptor activation. PMID:8710935

  3. Muscarinic presynaptic modulation in GABAergic pallidal synapses of the rat.

    PubMed

    Hernández-Martínez, Ricardo; Aceves, José J; Rueda-Orozco, Pavel E; Hernández-Flores, Teresa; Hernández-González, Omar; Tapia, Dagoberto; Galarraga, Elvira; Bargas, José

    2015-02-01

    The external globus pallidus (GPe) is central for basal ganglia processing. It expresses muscarinic cholinergic receptors and receives cholinergic afferents from the pedunculopontine nuclei (PPN) and other regions. The role of these receptors and afferents is unknown. Muscarinic M1-type receptors are expressed by synapses from striatal projection neurons (SPNs). Because axons from SPNs project to the GPe, one hypothesis is that striatopallidal GABAergic terminals may be modulated by M1 receptors. Alternatively, some M1 receptors may be postsynaptic in some pallidal neurons. Evidence of muscarinic modulation in any of these elements would suggest that cholinergic afferents from the PPN, or other sources, could modulate the function of the GPe. In this study, we show this evidence using striatopallidal slice preparations: after field stimulation in the striatum, the cholinergic muscarinic receptor agonist muscarine significantly reduced the amplitude of inhibitory postsynaptic currents (IPSCs) from synapses that exhibited short-term synaptic facilitation. This inhibition was associated with significant increases in paired-pulse facilitation, and quantal content was proportional to IPSC amplitude. These actions were blocked by atropine, pirenzepine, and mamba toxin-7, suggesting that receptors involved were M1. In addition, we found that some pallidal neurons have functional postsynaptic M1 receptors. Moreover, some evoked IPSCs exhibited short-term depression and a different kind of modulation: they were indirectly modulated by muscarine via the activation of presynaptic cannabinoid CB1 receptors. Thus pallidal synapses presenting distinct forms of short-term plasticity were modulated differently. PMID:25392165

  4. Muscarinic contribution to the acute cortical effects of vagus nerve stimulation

    NASA Astrophysics Data System (ADS)

    Nichols, Justin A.

    2011-12-01

    Electrical stimulation of the vagus nerve (VNS) has been used to treat more than 60,000 patients with drug-resistant epilepsy and is under investigation as a treatment for several other neurological disorders and conditions. Among these, VNS increases memory performance and enhances recovery of motor and cognitive function in animal models of traumatic brain injury. Recent research indicates that pairing brief VNS with tones multiple-times a day for several weeks induces long-term, input specific cortical plasticity, which can be used to re-normalize the pathological cortical reorganization and eliminate a behavioral correlate of chronic tinnitus in noise exposed rats. Despite the therapeutic potential, the mechanisms of action of VNS remain speculative. In chapter 2 of this dissertation, the acute effects of VNS on cortical synchrony, excitability, and temporal processing are examined. In anesthetized rats implanted with multi-electrode arrays, VNS increased and decorrelated spontaneous multi-unit activity, and suppressed entrainment to repetitive noise burst stimulation at 6 to 8 Hz, but not after systemic administration of the muscarinic antagonist scopolamine. Chapter 3 focuses on VNS-tone pairing induced cortical plasticity. Pairing VNS with a tone one hundred times in anesthetized rats resulted in frequency specific plasticity in 31% of the auditory cortex sites. Half of these sites exhibited a frequency specific increase in firing rate and half exhibited a frequency specific decrease. Muscarinic receptor blockade with scopolamine almost entirely prevented the frequency specific increases, but not decreases. Collectively, these experiments demonstrate the capacity for VNS to not only acutely influence cortical synchrony, and excitability, but to also influence temporal and spectral tuning via muscarinic receptor activation. These results strengthen the hypothesis that acetylcholine and muscarinic receptors are involved in the mechanisms of action of VNS and

  5. Evidence for the pharmacological similarity between the central presynaptic muscarinic autoreceptor and postsynaptic muscarinic receptors.

    PubMed Central

    Bowen, D. M.; Marek, K. L.

    1982-01-01

    Twenty antagonist substances with varying potencies for central and peripheral postsynaptic muscarinic receptors have been examined for effects on the central presynaptic muscarinic autoreceptor. This has been monitored by measuring the stimulating effects of the substances on acetylcholine synthesis by rat neocortical tissue prisms. Dose-response curves for selected agents showed that maximal stimulation of synthesis was to 136-140% of the value without an antagonist. At a concentration of 1 microM, 17 of the substances caused a significant increase in synthesis, whilst at 0.01 microM significant stimulation occurred with only atropine, dexetimide, N-methyl-piperdin-4-yl (R)-2-cyclohexyl-2-hydroxyl-2-phenylacetate, quinuclidinyl benzilate (QNB) and scopolamine. Linear regression analysis between synthesis values obtained with the substances and published data for the effects on either cholinoceptor-agonist induced contraction of guinea-pig ileum or the binding of [3H]-QNB to rat forebrain membranes gave correlation coefficients of r = 0.84 (P less than 0.01), and r = 0.75 (P less than 0.02) respectively. The results provide no indication of a pharmacological difference between the central presynaptic muscarinic autoreceptor and central and peripheral postsynaptic muscarinic receptors. PMID:7186824

  6. Modulation of neuritogenesis by astrocyte muscarinic receptors.

    PubMed

    Guizzetti, Marina; Moore, Nadia H; Giordano, Gennaro; Costa, Lucio G

    2008-11-14

    Astrocytes have been shown to release factors that have promoting or inhibiting effects on neuronal development. However, mechanisms controlling the release of such factors from astrocytes are not well established. Astrocytes express muscarinic receptors whose activation stimulates a robust intracellular signaling, although the role of these receptors in glial cells is not well understood. Acetylcholine and acetylcholine receptors are present in the brain before synaptogenesis occurs and are believed to be involved in neuronal maturation. The present study was undertaken to investigate whether stimulation of muscarinic receptors in astrocytes would modulate neurite outgrowth in hippocampal neurons. Rat hippocampal neurons, co-cultured with rat cortical astrocytes previously exposed to the cholinergic agonist carbachol, displayed longer neurites. The effect of carbachol in astrocytes was due to the activation of M3 muscarinic receptors. Exposure of astrocytes to carbachol increased the expression of the extracellular matrix proteins fibronectin and laminin-1 in these cells. This effect was mediated in part by an increase in laminin-1 and fibronectin mRNA levels and in part by the up-regulation of the production and release of plasminogen activator inhibitor-1, an inhibitor of the proteolytic degradation of the extracellular matrix. The inhibition of fibronectin activity strongly reduced the effect of carbachol on the elongation of all the neurites, whereas inhibition of laminin-1 activity reduced the elongation of minor neurites only. Plasminogen activator inhibitor-1 also induced neurite elongation through a direct effect on neurons. Taken together, these results demonstrate that cholinergic muscarinic stimulation of astrocytes induces the release of permissive factors that accelerate neuronal development.

  7. Repeated administration of fresh garlic increases memory retention in rats.

    PubMed

    Haider, Saida; Naz, Nosheen; Khaliq, Saima; Perveen, Tahira; Haleem, Darakhshan J

    2008-12-01

    Garlic (Allium sativum) is regarded as both a food and a medicinal herb. Increasing attention has focused on the biological functions and health benefits of garlic as a potentially major dietary component. Chronic garlic administration has been shown to enhance memory function. Evidence also shows that garlic administration in rats affects brain serotonin (5-hydroxytryptamine [5-HT]) levels. 5-HT, a neurotransmitter involved in a number of physiological functions, is also known to enhance cognitive performance. The present study was designed to investigate the probable neurochemical mechanism responsible for the enhancement of memory following garlic administration. Sixteen adult locally bred male albino Wistar rats were divided into control (n = 8) and test (n = 8) groups. The test group was orally administered 250 mg/kg fresh garlic homogenate (FGH), while control animals received an equal amount of water daily for 21 days. Estimation of plasma free and total tryptophan (TRP) and whole brain TRP, 5-HT, and 5-hydroxyindole acetic acid (5-HIAA) was determined by high-performance liquid chromatography with electrochemical detection. For assessment of memory, a step-through passive avoidance paradigm (electric shock avoidance) was used. The results showed that the levels of plasma free TRP significantly increased (P < .01) and plasma total TRP significantly decreased (P < .01) in garlic-treated rats. Brain TRP, 5-HT, and 5-HIAA levels were also significantly increased following garlic administration. A significant improvement in memory function was exhibited by garlic-treated rats in the passive avoidance test. Increased brain 5-HT levels were associated with improved cognitive performance. The present results, therefore, demonstrate that the memory-enhancing effect of garlic may be associated with increased brain 5-HT metabolism in rats. The results further support the use of garlic as a food supplement for the enhancement of memory. PMID:19053859

  8. Increased brain nitric oxide levels following ethanol administration.

    PubMed

    Finnerty, Niall; O'Riordan, Saidhbhe L; Klamer, Daniel; Lowry, John; Pålsson, Erik

    2015-05-01

    Nitric oxide is a ubiquitous messenger molecule, which at elevated concentrations has been implicated in the pathogenesis of several neurological disorders. Its role in oxidative stress, attributed in particular to the formation of peroxynitrite, proceeds through its high affinity for the superoxide radical. Alcoholism has recently been associated with the induction of oxidative stress, which is generally defined as a shift in equilibrium between pro-oxidant and anti-oxidant species in the direction of the former. Furthermore, its primary metabolite acetaldehyde, has been extensively associated with oxidative damage related toxic effects following alcohol ingestion. The principal objective of this study was the application of long term in vivo electrochemistry (LIVE) to investigate the effect of ethanol (0.125, 0.5 and 2.0 g kg(-1)) and acetaldehyde (12.5, 50 and 200 mg kg(-1)) on NO levels in the nucleus accumbens of freely moving rats. Systemic administrations of ethanol and acetaldehyde resulted in a dose-dependent increases in NO levels, albeit with very differing time courses. Subsequent to this the effect on accumbal NO levels, of subjecting the animal to different drug combinations, was also elucidated. The nitric oxide synthase inhibitor L-NAME (20 mg kg(-1)) and acetaldehyde sequestering agent D-penicillamine (50 mg kg(-1)) both attenuated the increase in NO levels following ethanol (1 g kg(-1)) administration. Conversely, the alcohol dehydrogenase inhibitor 4-methylpyrazole (25 mg kg(-1)) and catalase inhibitor sodium azide (10 mg kg(-1)) potentiated the increase in NO levels following ethanol administration. Finally, dual inhibition of aldehyde dehydrogenase and catalase by cyanamide (25 mg kg(-1)) caused an attenuation of ethanol effects on NO levels. Taken together these data highlight a robust increase in brain NO levels following systemic alcohol administration which is dependent on NO synthase activity and may involve both alcohol- and acetaldehyde

  9. Intranasal administration of oxytocin increases human aggressive behavior.

    PubMed

    Ne'eman, R; Perach-Barzilay, N; Fischer-Shofty, M; Atias, A; Shamay-Tsoory, S G

    2016-04-01

    Considering its role in prosocial behaviors, oxytocin (OT) has been suggested to diminish levels of aggression. Nevertheless, recent findings indicate that oxytocin may have a broader influence on increasing the salience of social stimuli and may therefore, under certain circumstances, increase antisocial behaviors such as aggression. This controversy led to the following speculations: If indeed oxytocin promotes primarily prosocial behavior, administration of OT is expected to diminish levels of aggression. However, if oxytocin mainly acts to increase the salience of social stimuli, it is expected to elevate levels of aggression following provocation. In order to test this assumption we used the Social Orientation Paradigm (SOP), a monetary game played against a fictitious partner that allows measuring three types of responses in the context of provocation: an aggressive response - reducing a point from the fictitious partner, an individualistic response - adding a point to oneself, and a collaborative response - adding half a point to the partner and half a point to oneself. In the current double-blind, placebo-controlled, within-subject study design, 45 participants completed the SOP task following the administration of oxytocin or placebo. The results indicated that among subjects naïve to the procedure oxytocin increased aggressive responses in comparison with placebo. These results support the saliency hypothesis of oxytocin and suggest that oxytocin plays a complex role in the modulation of human behavior. PMID:26862988

  10. INCREASING AND PROLONGING BLOOD PENICILLIN CONCENTRATIONS FOLLOWING INTRAMUSCULAR ADMINISTRATION.

    PubMed

    Bronfenbrenner, J; Favour, C B

    1945-06-29

    (1) Restriction of fluid intake to 1,500 cc and the salt intake to 3 gm a day doubles the penicillin blood level following interrupted intramuscular [See Figure in the PDF file] injections of penicillin. (2) The administration of benzoic acid to a patient on an unrestricted diet Ilay double the penicillin blood level during similar treatment. (3) The combination of these two procedures results in a four- to eight-fold increase in penicillin blood level with a prolonged effective blood concentration.

  11. L-tyrosine administration increases acetylcholinesterase activity in rats.

    PubMed

    Ferreira, Gabriela K; Carvalho-Silva, Milena; Gonçalves, Cinara L; Vieira, Júlia S; Scaini, Giselli; Ghedim, Fernando V; Deroza, Pedro F; Zugno, Alexandra I; Pereira, Talita C B; Oliveira, Giovanna M T; Kist, Luiza W; Bogo, Maurício R; Schuck, Patrícia F; Ferreira, Gustavo C; Streck, Emilio L

    2012-12-01

    Tyrosinemia is a rare genetic disease caused by mutations on genes that codify enzymes responsible for tyrosine metabolism. Considering that tyrosinemics patients usually present symptoms associated with central nervous system alterations that ranges from slight decreases in intelligence to severe mental retardation, we decided to investigate whether acute and chronic administration of L-tyrosine in rats would affect acetylcholinesterase mRNA expression and enzymatic activity during their development. In our acute protocol, Wistar rats (10 and 30 days old) were killed one hour after a single intraperitoneal L-tyrosine injection (500 mg/kg) or saline. Chronic administration consisted of L-tyrosine (500 mg/kg) or saline injections 12 h apart for 24 days in Wistar rats (7 days old) and rats were killed 12 h after last injection. Acetylcholinesterase activity was measured by Ellman's method and acetylcholinesterase expression was carried out by a semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) assay. We observed that acute (10 and 30 days old rats) and chronic L-tyrosine administration increased acetylcholinesterase activity in serum and all tested brain areas (hippocampus, striatum and cerebral cortex) when compared to control group. Moreover, there was a significant decrease in mRNA levels of acetylcholinesterase in hippocampus was observed after acute protocol (10 and 30 days old rats) and in striatum after chronic protocol. In case these alterations also occur in the brain of the patients, our results may explain, at least in part, the neurological sequelae associated with high plasma concentrations of tyrosine seen in patients affected by tyrosinemia type II. PMID:23046746

  12. L-tyrosine administration increases acetylcholinesterase activity in rats.

    PubMed

    Ferreira, Gabriela K; Carvalho-Silva, Milena; Gonçalves, Cinara L; Vieira, Júlia S; Scaini, Giselli; Ghedim, Fernando V; Deroza, Pedro F; Zugno, Alexandra I; Pereira, Talita C B; Oliveira, Giovanna M T; Kist, Luiza W; Bogo, Maurício R; Schuck, Patrícia F; Ferreira, Gustavo C; Streck, Emilio L

    2012-12-01

    Tyrosinemia is a rare genetic disease caused by mutations on genes that codify enzymes responsible for tyrosine metabolism. Considering that tyrosinemics patients usually present symptoms associated with central nervous system alterations that ranges from slight decreases in intelligence to severe mental retardation, we decided to investigate whether acute and chronic administration of L-tyrosine in rats would affect acetylcholinesterase mRNA expression and enzymatic activity during their development. In our acute protocol, Wistar rats (10 and 30 days old) were killed one hour after a single intraperitoneal L-tyrosine injection (500 mg/kg) or saline. Chronic administration consisted of L-tyrosine (500 mg/kg) or saline injections 12 h apart for 24 days in Wistar rats (7 days old) and rats were killed 12 h after last injection. Acetylcholinesterase activity was measured by Ellman's method and acetylcholinesterase expression was carried out by a semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) assay. We observed that acute (10 and 30 days old rats) and chronic L-tyrosine administration increased acetylcholinesterase activity in serum and all tested brain areas (hippocampus, striatum and cerebral cortex) when compared to control group. Moreover, there was a significant decrease in mRNA levels of acetylcholinesterase in hippocampus was observed after acute protocol (10 and 30 days old rats) and in striatum after chronic protocol. In case these alterations also occur in the brain of the patients, our results may explain, at least in part, the neurological sequelae associated with high plasma concentrations of tyrosine seen in patients affected by tyrosinemia type II.

  13. Pharmacological approaches to targeting muscarinic acetylcholine receptors.

    PubMed

    Matera, Carlo; Tata, Ada M

    2014-01-01

    The presence of cholinergic system markers and muscarinic receptor subtypes in several tissues also of nonneuronal type has been largely demonstrated. Acetylcholine, synthesized in the nervous system, can locally contribute to modulate cell proliferation, survival and apoptosis. Considering that the cholinergic system functions are impaired in a number of disorders, the identification of new drugs regulating these functions appears of great clinical relevance. The possible involvement of muscarinic acetylcholine receptors in different pathologies has been proposed in recent years and is becoming an important area of study. However, the lack of selective muscarinic receptor ligands has for long time limited the therapeutic treatment based on muscarinic receptors as targets. To date, some muscarinic ligands such as xanomeline (patent, US5980933) or cevimeline (patents US4855290, US5571918) have been developed for the treatment of several pathologies (Alzheimer's and Sjogren's diseases). The present review will be focused on the potential effects produced by muscarinic receptor activation in different pathologies, including tumors. In fact, the potential use of muscarinic ligands in therapeutic protocols in cancer therapy will be discussed, considering that several muscarinic antagonists, already used in the treatment of genitourinary diseases (e.g. darifenacin, patent, US5096890, US6106864), have also been demonstrated to arrest the tumor growth in vivo. Moreover, the contribution of muscarinic receptors to analgesia is also reviewed. Finally, some of the most significant achievements in the field of bitopic/dualsteric ligands will be discussed and the molecules patented so far will be presented.

  14. Anti-muscarinic adjunct therapy accelerates functional human oligodendrocyte repair.

    PubMed

    Abiraman, Kavitha; Pol, Suyog U; O'Bara, Melanie A; Chen, Guang-Di; Khaku, Zainab M; Wang, Jing; Thorn, David; Vedia, Bansi H; Ekwegbalu, Ezinne C; Li, Jun-Xu; Salvi, Richard J; Sim, Fraser J

    2015-02-25

    Therapeutic repair of myelin disorders may be limited by the relatively slow rate of human oligodendrocyte differentiation. To identify appropriate pharmacological targets with which to accelerate differentiation of human oligodendrocyte progenitors (hOPCs) directly, we used CD140a/O4-based FACS of human forebrain and microarray to hOPC-specific receptors. Among these, we identified CHRM3, a M3R muscarinic acetylcholine receptor, as being restricted to oligodendrocyte-biased CD140a(+)O4(+) cells. Muscarinic agonist treatment of hOPCs resulted in a specific and dose-dependent blockade of oligodendrocyte commitment. Conversely, when hOPCs were cocultured with human neurons, M3R antagonist treatment stimulated oligodendrocytic differentiation. Systemic treatment with solifenacin, an FDA-approved muscarinic receptor antagonist, increased oligodendrocyte differentiation of transplanted hOPCs in hypomyelinated shiverer/rag2 brain. Importantly, solifenacin treatment of engrafted animals reduced auditory brainstem response interpeak latency, indicative of increased conduction velocity and thereby enhanced functional repair. Therefore, solifenacin and other selective muscarinic antagonists represent new adjunct approaches to accelerate repair by engrafted human progenitors.

  15. Anti-Muscarinic Adjunct Therapy Accelerates Functional Human Oligodendrocyte Repair

    PubMed Central

    Abiraman, Kavitha; Pol, Suyog U.; O'Bara, Melanie A.; Chen, Guang-Di; Khaku, Zainab M.; Wang, Jing; Thorn, David; Vedia, Bansi H.; Ekwegbalu, Ezinne C.; Li, Jun-Xu; Salvi, Richard J.

    2015-01-01

    Therapeutic repair of myelin disorders may be limited by the relatively slow rate of human oligodendrocyte differentiation. To identify appropriate pharmacological targets with which to accelerate differentiation of human oligodendrocyte progenitors (hOPCs) directly, we used CD140a/O4-based FACS of human forebrain and microarray to hOPC-specific receptors. Among these, we identified CHRM3, a M3R muscarinic acetylcholine receptor, as being restricted to oligodendrocyte-biased CD140a+O4+ cells. Muscarinic agonist treatment of hOPCs resulted in a specific and dose-dependent blockade of oligodendrocyte commitment. Conversely, when hOPCs were cocultured with human neurons, M3R antagonist treatment stimulated oligodendrocytic differentiation. Systemic treatment with solifenacin, an FDA-approved muscarinic receptor antagonist, increased oligodendrocyte differentiation of transplanted hOPCs in hypomyelinated shiverer/rag2 brain. Importantly, solifenacin treatment of engrafted animals reduced auditory brainstem response interpeak latency, indicative of increased conduction velocity and thereby enhanced functional repair. Therefore, solifenacin and other selective muscarinic antagonists represent new adjunct approaches to accelerate repair by engrafted human progenitors. PMID:25716865

  16. Facilitation of memory storage by the acetylcholine M2 muscarinic receptor antagonist AF-DX 116.

    PubMed

    Baratti, C M; Opezzo, J W; Kopf, S R

    1993-07-01

    Post-training administration of the acetylcholine muscarinic M2 presynaptic receptor antagonist AF-DX 116 (0.1-10.0 mg/kg, ip), facilitated 48 h retention, in male Swiss mice, of a one-trial step-through inhibitory avoidance task. The dose-response curve was an inverted U. AF-DX 116 did not increase the retention latencies of mice that had not received a footshock during training. The influence of AF-DX 116 (1 mg/kg, ip) on retention was time-dependent, which suggests that the drug facilitated memory storage. The memory facilitation induced by AF-DX 116 (1 mg/kg, ip) was prevented by atropine (0.5 mg/kg, ip) administered after training, but 10 min prior to AF-DX 116 treatment. In contrast, neither methylatropine (0.5 mg/kg, ip), a peripherally acting muscarinic receptor blocker, nor mecamylamine (5 mg/kg, ip) or hexamethonium (5 mg/kg, ip), two cholinergic nicotinic receptor antagonists, prevented the effects of post-training AF-DX 116 on retention. Low subeffective doses of the central acting anticholinesterase physostigmine (35 micrograms/kg, ip), administered immediately after training, and AF-DX 116 (0.1 mg/kg, ip), given 10 min after training, acted synergistically to improve retention. The effects of AF-DX 116 (0.1 mg/kg, ip) were not influenced by the peripherally acting anticholinesterase neostigmine (35 micrograms/kg, ip). Considered together, these findings suggest that the activation of a muscarinic cholinergic presynaptic inhibitory mechanism, probably by increasing brain acetylcholine release, may modulate the activity of post-training processes involved in memory storage. PMID:8216161

  17. Facilitation of memory storage by the acetylcholine M2 muscarinic receptor antagonist AF-DX 116.

    PubMed

    Baratti, C M; Opezzo, J W; Kopf, S R

    1993-07-01

    Post-training administration of the acetylcholine muscarinic M2 presynaptic receptor antagonist AF-DX 116 (0.1-10.0 mg/kg, ip), facilitated 48 h retention, in male Swiss mice, of a one-trial step-through inhibitory avoidance task. The dose-response curve was an inverted U. AF-DX 116 did not increase the retention latencies of mice that had not received a footshock during training. The influence of AF-DX 116 (1 mg/kg, ip) on retention was time-dependent, which suggests that the drug facilitated memory storage. The memory facilitation induced by AF-DX 116 (1 mg/kg, ip) was prevented by atropine (0.5 mg/kg, ip) administered after training, but 10 min prior to AF-DX 116 treatment. In contrast, neither methylatropine (0.5 mg/kg, ip), a peripherally acting muscarinic receptor blocker, nor mecamylamine (5 mg/kg, ip) or hexamethonium (5 mg/kg, ip), two cholinergic nicotinic receptor antagonists, prevented the effects of post-training AF-DX 116 on retention. Low subeffective doses of the central acting anticholinesterase physostigmine (35 micrograms/kg, ip), administered immediately after training, and AF-DX 116 (0.1 mg/kg, ip), given 10 min after training, acted synergistically to improve retention. The effects of AF-DX 116 (0.1 mg/kg, ip) were not influenced by the peripherally acting anticholinesterase neostigmine (35 micrograms/kg, ip). Considered together, these findings suggest that the activation of a muscarinic cholinergic presynaptic inhibitory mechanism, probably by increasing brain acetylcholine release, may modulate the activity of post-training processes involved in memory storage.

  18. Chronic ethanol (EtOH) feeding increases muscarinic receptor (mAChR) density in esophagus without parallel change in dose response (D-R) to cholinergic agonists

    SciTech Connect

    Keshavarzian, A.; Gordon, J.H.; Urban, G.; Fields, J.Z. VA Hospital, Hines, IL )

    1991-03-11

    The mAChR/effector pathway for signal transduction is important in the physiology of esophagus and mAChR alterations are involved in EtOH induced changes in several organs. To see if EtOH-induced increases in lower esophageal sphincter pressure (LESP) are due to upregulation of mAChR, the authors evaluated mAChR binding and D-R curves for bethanechol (IV) induced increases in LESP, and compared these values to changes in LESP after acute and chronic EtOH. EtOH was given to cats acutely or chronically. The number of mAChR sites (Bmax) in esophagus was lowered by acute EtOH, withdrawal from chronic EtOH raised Bmax. Acute injection of EtOH to cats in withdrawal reversed this increase in mAChR density. These changes correlated with the earlier data on EtOH-induced changes in LESP. In contrast, the D-R curve for bethanechol shifted to the right. Thus, the withdrawal-associated increase in Bmax is more likely to be a compensatory response to deficits distal to the receptor recognition site than to proximal deficits and doesn't cause LESP hyperactivity. Also, receptor binding changes do not necessarily translate into physiological changes.

  19. Acetylcholine as a mitogen: muscarinic receptor-mediated proliferation of rat astrocytes and human astrocytoma cells.

    PubMed

    Guizzetti, M; Costa, P; Peters, J; Costa, L G

    1996-02-22

    The mitogenic effect of muscarinic receptor agonists in glial cells has been characterized in rat cortical astrocytes and human 132 1N1 astrocytoma cells. The muscarinic receptor agonist carbachol caused a dose- and time-dependent increase in proliferation, as measured by [3H]thymidine incorporation. The mitogenic effect was mimicked by several muscarinic, but not nicotinic receptor agonists, and was blocked by muscarinic receptor antagonists. Reverse transcription-polymerase chain reaction (RT-PCR) experiments indicated the presence of m2, m3 and to a lesser degree, m5 muscarinic receptor mRNA in both astrocytes and astrocytoma cells. Proliferation experiments with subtype-specific muscarinic receptor antagonists suggest that carbachol-induced proliferation is due to activation of muscarinic M3 receptors. The phorbol ester 12-O-tetradecanoyl-phorbol 13-acetate (TPA) also stimulated glial cell proliferation. Down-regulation of protein kinase C, or the protein kinase C antagonist 1,5-(isoquinolynsulfanyl)-2-methylpiperazine dihydrochloride (H7) blocked proliferation induced by either TPA or carbachol. Of other neurotransmitters tested, histamine caused glial cell proliferation, norepinephrine and gamma-aminobutyric acid were ineffective, while serotonin and glutamate inhibited basal or serum-stimulated proliferation. PMID:8666059

  20. Regulation and ontogeny of subtypes of muscarinic receptors and muscarinic receptor-mediated

    SciTech Connect

    Lee, W.

    1989-01-01

    The densities of total and M1 muscarinic receptors were measured using the muscarinic receptor antagonists {sup 3}H-quinuclidinyl benzilate and {sup 3}H-pirenzepine, respectively. Thus, the difference between the density of {sup 3}H-quinuclidinyl benzilate and {sup 3}H-pirenzepine binding sites represents the density of M2 sites. In addition, there is no observable change in either acetylcholine-stimulated phosphoinositide breakdown (suggested to be an M1 receptor-mediated response) or in carbachol-mediated inhibition of cyclic AMP accumulation (suggested to be an M2 receptor-mediated response) in slices of cortex+dorsal hippocampus following chronic atropine administration. In other experiments, it has been shown that the M1 and M2 receptors in rat cortex have different ontogenetic profiles. The M2 receptor is present at adult levels at birth, while the M1 receptor develops slowly from low levels at postnatal week 1 to adult levels at postnatal week 3. The expression of acetylcholine-stimulated phosphoinositide breakdown parallels the development of M1 receptors, while the development of carbachol-mediated inhibition of cyclic AMP accumulation occurs abruptly between weeks 2 and 3 postnatally.

  1. Alkylating derivative of oxotremorine interacts irreversibly with the muscarinic receptor

    SciTech Connect

    Ehlert, F.J.; Jenden, D.J.; Ringdahl, B.

    1984-03-05

    A 2-chloroethylamine derivative of oxotremorine was studied in pharmacological experiments and muscarinic receptor binding assays. The compound, N-(4-(2-chloroethylmethylamino)-2-butynyl)-2-pyrrolidone (BM 123), forms an aziridinium ion in aqueous solution at neutral pH that stimulates contractions of guinea pig ileum with a potency similar to that of oxotremorine. Following the initial stimulation, there is a long lasting period of lack of sensitivity of the guinea pig ileum to muscarinic agonists. BM 123 also produces muscarinic effects in vivo. When homogenates of the rat cerebral cortex were incubated with BM 123 and assayed subsequently in muscarinic receptor binding assays, a loss of binding capacity for the muscarinic antagonist, (/sup 3/H)N-methylscopolamine ((/sup 3/H)NMS), was noted without a change in affinity. Similar observations were made in (/sup 3/H)1-3-quinuclidinyl benzilate ((/sup 3/H)-QNB) binding assays on the forebrains of mice that had been injected with BM 123 24 hr earlier. The loss in receptor capacity for both (/sup 3/H)NMS and (/sup 3/H)-QNB was prevented by atropine treatment. Kinetic studies of the interaction of BM 123 with homogenates of the rat cerebral cortex in vitro showed that the half-time for the loss of (/sup 3/H)-QNB binding sites increased from 10 to 45 min as the concentration of BM 123 decreased from 10 to 1 ..mu..M. In contrast to the aziridinium ion, the parent 2-chloroethylamine compound and the alcoholic hydrolysis product were largely devoid of pharmacological and binding activity.

  2. Differential Muscarinic Modulation in the Olfactory Bulb

    PubMed Central

    Smith, Richard S.; Hu, Ruilong; DeSouza, Andre; Eberly, Christian L.; Krahe, Krista; Chan, Wilson

    2015-01-01

    Neuromodulation of olfactory circuits by acetylcholine (ACh) plays an important role in odor discrimination and learning. Early processing of chemosensory signals occurs in two functionally and anatomically distinct regions, the main and accessory olfactory bulbs (MOB and AOB), which receive extensive cholinergic input from the basal forebrain. Here, we explore the regulation of AOB and MOB circuits by ACh, and how cholinergic modulation influences olfactory-mediated behaviors in mice. Surprisingly, despite the presence of a conserved circuit, activation of muscarinic ACh receptors revealed marked differences in cholinergic modulation of output neurons: excitation in the AOB and inhibition in the MOB. Granule cells (GCs), the most abundant intrinsic neuron in the OB, also exhibited a complex muscarinic response. While GCs in the AOB were excited, MOB GCs exhibited a dual muscarinic action in the form of a hyperpolarization and an increase in excitability uncovered by cell depolarization. Furthermore, ACh influenced the input–output relationship of mitral cells in the AOB and MOB differently showing a net effect on gain in mitral cells of the MOB, but not in the AOB. Interestingly, despite the striking differences in neuromodulatory actions on output neurons, chemogenetic inhibition of cholinergic neurons produced similar perturbations in olfactory behaviors mediated by these two regions. Decreasing ACh in the OB disrupted the natural discrimination of molecularly related odors and the natural investigation of odors associated with social behaviors. Thus, the distinct neuromodulation by ACh in these circuits could underlie different solutions to the processing of general odors and semiochemicals, and the diverse olfactory behaviors they trigger. SIGNIFICANCE STATEMENT State-dependent cholinergic modulation of brain circuits is critical for several high-level cognitive functions, including attention and memory. Here, we provide new evidence that cholinergic

  3. Vasoactive Intestinal Polypeptide and Muscarinic Receptors: Supersensitivity Induced by Long-Term Atropine Treatment

    NASA Astrophysics Data System (ADS)

    Hedlund, Britta; Abens, Janis; Bartfai, Tamas

    1983-04-01

    Long-term treatment of rats with atropine induced large increases in the numbers of muscarinic receptors and receptors for vasoactive intestinal polypeptide in the salivary glands. Since receptors for vasoactive intestinal polypeptide coexist with muscarinic receptors on the same neurons in this preparation, the results suggest that a drug that alters the sensitivity of one receptor may also affect the sensitivity of the receptor for a costored transmitter and in this way contribute to the therapeutic or side effects of the drug.

  4. Muscarinic receptor occupancy by biperiden in living human brain.

    PubMed

    Sudo, Y; Suhara, T; Suzuki, K; Okubo, Y; Yoshikawa, K; Uchida, S; Sassa, T; Okauchi, T; Sasaki, Y; Matsushita, M

    1999-01-01

    Anticholinergic drug is often used to treat extrapyramidal symptoms. We measured muscarinic cholinergic receptor (mAchR) occupancy by the oral administration of biperiden in eight healthy subjects using positron emission tomography (PET) and [11C]N-methyl-4-piperidylbenzilate (NMPB). After the baseline scan each subject underwent one or two post-dose PET scans. mAchR occupancy was 10-45% in the frontal cortex three hours after the oral administration of 4 mg of biperiden. The occupancy correlated with the plasma concentration of biperiden in a curvilinear manner.

  5. Genetic deletion of muscarinic M4 receptors is anxiolytic in the shock-probe burying model.

    PubMed

    Degroot, Aldemar; Nomikos, George G

    2006-02-15

    We used muscarinic M2 and M4 receptor knockout (KO) mice to further explore the role of the cholinergic system in anxiety. Using the shock-probe burying model we were able to both assess anxiety and cognition. In this paradigm, an anxiolytic response is reflected by decreased burying behavior. In addition, retention latency depicts long-term memory performance. Whereas muscarinic M2 receptor KO mice did not differ behaviorally from wild-type mice, muscarinic M4 receptor KO mice showed increased anxiolysis, but normal long-term memory compared to wild-type mice. Therefore, muscarinic M4 receptors are of particular significance in anxiety modulation that seems dissociated from changes in long-term memory.

  6. Muscarinic receptor subtypes as potential targets to modulate oligodendrocyte progenitor survival, proliferation, and differentiation.

    PubMed

    De Angelis, Federica; Bernardo, Antonietta; Magnaghi, Valerio; Minghetti, Luisa; Tata, Ada Maria

    2012-05-01

    Acetylcholine (ACh) is a major neurotransmitter but also an important signaling molecule in neuron-glia interactions. Expression of ACh receptors has been reported in several glial cell populations, including oligodendrocytes (OLs). Nonetheless, the characterization of muscarinic receptors in these cells, as well as the description of the cholinergic effects at different stages of OL development, is still incomplete. In this study, we characterized the pattern of expression of muscarinic receptor subtypes in primary cultures of rat oligodendrocyte progenitor cells (OPC) and mature OLs, at both mRNA and protein levels. We found that muscarinic receptor expression is developmentally regulated. M1, M3, and M4 receptors were the main subtypes expressed in OPC, whereas all receptor subtypes were expressed at low levels in mature OLs. Exposure of OPC to muscarine enhanced cell proliferation, an effect mainly due to M1, M3, and M4 receptor subtypes as demonstrated by pharmacological competition with selective antagonists. Conversely, M2 receptor activation impaired OPC survival. In line with the mitogenic activity, muscarinic receptor activation increased the expression of platelet derived growth factor receptor α. Muscarine stimulation increased CX32 and myelin basic protein expression, left unaffected that of myelin proteolipid protein (PLP), and decreased member of the family of epidermal growth factor receptor (EGFR) ErbB3/ErbB4 receptor expression indicating a predominant role of muscarinic receptors in OPC. These findings suggest that ACh may contribute to the maintenance of an immature proliferating progenitor pool and impair the progression toward mature stage. This hypothesis is further supported by increased expression of Notch-1 in OL on muscarinic activation.

  7. Enhanced muscarinic M1 receptor gene expression in the corpus striatum of streptozotocin-induced diabetic rats

    PubMed Central

    Gireesh, G; Kumar, T Peeyush; Mathew, Jobin; Paulose, CS

    2009-01-01

    Acetylcholine (ACh), the first neurotransmitter to be identified, regulate the activities of central and peripheral functions through interactions with muscarinic receptors. Changes in muscarinic acetylcholine receptor (mAChR) have been implicated in the pathophysiology of many major diseases of the central nervous system (CNS). Previous reports from our laboratory on streptozotocin (STZ) induced diabetic rats showed down regulation of muscarinic M1 receptors in the brainstem, hypothalamus, cerebral cortex and pancreatic islets. In this study, we have investigated the changes of acetylcholine esterase (AChE) enzyme activity, total muscarinic and muscarinic M1 receptor binding and gene expression in the corpus striatum of STZ – diabetic rats and the insulin treated diabetic rats. The striatum, a neuronal nucleus intimately involved in motor behaviour, is one of the brain regions with the highest acetylcholine content. ACh has complex and clinically important actions in the striatum that are mediated predominantly by muscarinic receptors. We observed that insulin treatment brought back the decreased maximal velocity (Vmax) of acetylcholine esterase in the corpus striatum during diabetes to near control state. In diabetic rats there was a decrease in maximal number (Bmax) and affinity (Kd) of total muscarinic receptors whereas muscarinic M1 receptors were increased with decrease in affinity in diabetic rats. We observed that, in all cases, the binding parameters were reversed to near control by the treatment of diabetic rats with insulin. Real-time PCR experiment confirmed the increase in muscarinic M1 receptor gene expression and a similar reversal with insulin treatment. These results suggest the diabetes-induced changes of the cholinergic activity in the corpus striatum and the regulatory role of insulin on binding parameters and gene expression of total and muscarinic M1 receptors. PMID:19344500

  8. Chronic decentralization of the heart differentially remodels canine intrinsic cardiac neuron muscarinic receptors.

    PubMed

    Smith, F M; McGuirt, A S; Hoover, D B; Armour, J A; Ardell, J L

    2001-11-01

    The objective of the study was to determine if chronic interruption of all extrinsic nerve inputs to the heart alters cholinergic-mediated responses within the intrinsic cardiac nervous system (ICN). Extracardiac nerve inputs to the ICN were surgically interrupted (ICN decentralized). Three weeks later, the intrinsic cardiac right atrial ganglionated plexus (RAGP) was removed and intrinsic cardiac neuronal responses were evaluated electrophysiologically. Cholinergic receptor abundance was evaluated using autoradiography. In sham controls and chronic decentralized ICN ganglia, neuronal postsynaptic responses were mediated by acetylcholine, acting at nicotinic and muscarinic receptors. Muscarine- but not nicotine-mediated synaptic responses that were enhanced after chronic ICN decentralization. After chronic decentralization, muscarine facilitation of orthodromic neuronal activation increased. Receptor autoradiography demonstrated that nicotinic and muscarinic receptor density associated with the RAGP was unaffected by decentralization and that muscarinic receptors were tenfold more abundant than nicotinic receptors in the right atrial ganglia in each group. After chronic decentralization of the ICN, intrinsic cardiac neurons remain viable and responsive to cholinergic synaptic inputs. Enhanced muscarinic responsiveness of intrinsic cardiac neurons occurs without changes in receptor abundance.

  9. Comparison of muscarine- and vasopressin-stimulated inositol phospholipid metabolism in the superior cervical ganglion of the rat

    SciTech Connect

    Horwitz, J.; Anderson, C.; Perlman, R.L.

    1986-03-05

    Both muscarine and vasopressin have previously been shown to increase the accumulation of /sup 3/H-inositol phosphates (/sup 3/H-IP) in superior cervical ganglia in which the phospholipids were labeled with /sup 3/H-inositol. They have compared the effects of muscarine and vasopressin on phospholipid metabolism in the ganglion. The effects of these agents on /sup 3/H-IP accumulation are additive. The response to muscarine plateaus after approximately 10 min whereas the response to vasopressin increases for at least 30 min. Decentralization and maintenance in organ culture appear to potentiate the effect of muscarine on /sup 3/H-IP accumulation but do not effect the response of the ganglia to vasopressin. Muscarine and vasopressin also increase the incorporation of /sup 3/H-inositol into phospholipids in the ganglion. Autoradiographic techniques were used to localize the inositol-containing phospholipids in the ganglion. Muscarine increases phospholipid labeling primarily in the cell bodies of the principal ganglionic neurons, whereas vasopressin increases phospholipid labeling primarily in the neuropil. These data are consistent with the hypothesis that muscarine and vasopressin stimulate the metabolism of different pools of phospholipids.

  10. Long-acting muscarinic antagonists.

    PubMed

    Melani, Andrea S

    2015-01-01

    Chronic obstructive pulmonary disease (COPD) is a major cause of death and disability worldwide. Inhaled bronchodilators are the mainstay of COPD pharmacological treatment. Long-acting muscarinic antagonists (LAMAs) are a major class of inhaled bronchodilators. Some LAMA/device systems with different characteristics and dosing schedules are currently approved for maintenance therapy of COPD and a range of other products are being developed. They improve lung function and patient-reported outcomes and reduce acute bronchial exacerbations with good safety. LAMAs are used either alone or associated with long-acting β₂-agonists, eventually in fixed dose combinations. Long-acting β₂-agonist/LAMA combinations assure additional benefits over the individual components alone. The reader will obtain a view of the safety and efficacy of the different LAMA/device systems in COPD patients. PMID:26109098

  11. Role of muscarinic and nicotinic cholinergic receptors in an experimental model of epilepsy-induced analgesia.

    PubMed

    de Freitas, Renato Leonardo; de Oliveira, Rithiele Cristina; de Carvalho, Andressa Daiane; Felippotti, Tatiana Tocchini; Bassi, Gabriel Shimizu; Elias-Filho, Daoud Hibrahim; Coimbra, Norberto Cysne

    2004-10-01

    The blockade of GABA-mediated Cl(-) influx with pentylenetetrazol (PTZ) was used in the present work to induce seizures in animals. The neurotransmission in the postictal period has been the focus of many studies, and there is evidence suggesting antinociceptive mechanisms following tonic-clonic seizures in both animals and men. The aim of this work was to study the involvement of acetylcholine in the antinociception induced by convulsions elicited by peripheral administration of PTZ (64 mg/kg). Analgesia was measured by the tail-flick test in eight albino Wistar rats per group. Convulsions were followed by significant increases in tail-flick latencies (TFLs) at least for 120 min of the postictal period. Peripheral administration of atropine (0.25, 1 and 4 mg/kg) caused a significant dose-dependent decrease in the TFL in seizing animals, as compared to controls. These data were corroborated by peripheral administration of mecamylamine, a nicotinic cholinergic receptor blocker, at the same doses (0.25, 1 and 4 mg/kg) used for the muscarinic cholinergic receptor antagonist. The recruitment of the muscarinic receptor was made 10 min postconvulsions and in subsequent periods of postictal analgesia, whereas the involvement of the nicotinic cholinergic receptor was implicated only after 30 min postseizures. The cholinergic antagonists caused a minimal reduction in body temperature, but did not impair baseline TFL, spontaneous exploration or motor coordination in the rotarod test at the maximal dose of 4 mg/kg. These results indicate that acetylcholine may be involved as a neurotransmitter in postictal analgesia.

  12. Affective Analgesia following Muscarinic Activation of the Ventral Tegmental Area in Rats

    PubMed Central

    Kender, Robert G.; Harte, Steven E.; Munn, Elizabeth M.; Borszcz, George S.

    2009-01-01

    Cholinergic stimulation of dopamine neurons in the ventral tegmental area (VTA) underlies activation of the brain reward circuitry. Activation of this circuit is proposed to preferentially suppress the affective reaction to noxious stimulation. Vocalization afterdischarges (VADs) are a validated model of the affective response of rats to noxious tailshock. The antinociceptive action of the acetylcholine agonist carbachol microinjected into the VTA on VAD threshold was compared to its effect on the thresholds of other tailshock-elicited responses (VDS = vocalizations during shock, and SMR = spinal motor reflexes). Whereas VADs are organized within the forebrain, VDSs and SMRs are organized at medullary and spinal levels of the neuraxis, respectively. Carbachol (1 μg, 2 μg, and 4 μg) injected into VTA produced dose-dependent increases in VAD and VDS thresholds, although increases in VAD threshold were significantly greater than increases in VDS threshold. Administration of carbachol into VTA failed to elevate SMR threshold. Elevations in vocalization thresholds produced by intra-VTA carbachol were reversed in a dose-dependent manner by local administration of the muscarinic receptor antagonist atropine sulfate (30 μg and 60 μg). These results provide the first demonstration of the involvement of the VTA in muscarinic-induced suppression of pain affect. Perspective Cholinergic activation of the brain reward circuit produced a preferential suppression of rats’ affective reaction to noxious stimulation. The neurobiology that relates reinforcement to suppression of pain affect may provide insights into new treatments for pain and its associated affective disorders. PMID:18387853

  13. Functional subtyping of muscarinic receptors on canine esophageal mucosa.

    PubMed

    Lad, R; Donoff, B; Rangachari, P K

    1991-09-01

    Serosal addition of muscarinic agonists elicited rapid changes in electrical parameters across the isolated canine esophageal epithelium set up in vitro. Both carbachol and the M1-selective agonist, McNeil A343 (McN), increased transmucosal potential differences (PDs), decreased transmucosal resistances (R), and increased short-circuit currents (Isc). Carbachol was more potent and more effective than McN. Muscarinic antagonists were used to define the muscarinic receptor involved. The pA2 values obtained with Schild plots were as follows: atropine 9.14, 4-DAMP 8.98, AFDX-116 6.71, and pirenzepine 7.12. Low concentrations of pirenzepine (10(-8) M), produced a rightward shift in the dose-response curve to McN, without inhibiting responses to carbachol. Thus the receptor subtype is clearly not an M2. As in other glandular systems, M3 receptors are present. Whether M1 receptors also exist requires better definition of receptor densities-reserves in this tissue. Carbachol induced net secretion of Na and Cl and converted a predominantly absorptive tissue to a secretory one. PMID:1716057

  14. Intranasal administration of oxytocin increases compassion toward women

    PubMed Central

    Palgi, Sharon; Klein, Ehud

    2015-01-01

    It has been suggested that the degree of compassion—the feeling of warmth, understanding and kindness that motivates the desire to help others, is modulated by observers’ views regarding the target’s vulnerability and suffering. This study tested the hypothesis that as compassion developed to protect vulnerable kinships, hormones such as oxytocin, which have been suggested as playing a key role in ‘tend-and-befriend’ behaviors among women, will enhance compassion toward women but not toward men. Thirty subjects participated in a double-blind, placebo-controlled, within-subject study. Following administration of oxytocin/placebo, participants listened to recordings of different female/male protagonists describing distressful emotional conflicts and were then asked to provide compassionate advice to the protagonist. The participants’ responses were coded according to various components of compassion by two clinical psychologists who were blind to the treatment. The results showed that in women and men participants oxytocin enhanced compassion toward women, but did not affect compassion toward men. These findings indicate that the oxytocinergic system differentially mediates compassion toward women and toward men, emphasizing an evolutionary perspective that views compassion as a caregiving behavior designed to help vulnerable individuals. PMID:24711542

  15. Intranasal administration of oxytocin increases compassion toward women.

    PubMed

    Palgi, Sharon; Klein, Ehud; Shamay-Tsoory, Simone G

    2015-03-01

    It has been suggested that the degree of compassion-the feeling of warmth, understanding and kindness that motivates the desire to help others, is modulated by observers' views regarding the target's vulnerability and suffering. This study tested the hypothesis that as compassion developed to protect vulnerable kinships, hormones such as oxytocin, which have been suggested as playing a key role in 'tend-and-befriend' behaviors among women, will enhance compassion toward women but not toward men. Thirty subjects participated in a double-blind, placebo-controlled, within-subject study. Following administration of oxytocin/placebo, participants listened to recordings of different female/male protagonists describing distressful emotional conflicts and were then asked to provide compassionate advice to the protagonist. The participants' responses were coded according to various components of compassion by two clinical psychologists who were blind to the treatment. The results showed that in women and men participants oxytocin enhanced compassion toward women, but did not affect compassion toward men. These findings indicate that the oxytocinergic system differentially mediates compassion toward women and toward men, emphasizing an evolutionary perspective that views compassion as a caregiving behavior designed to help vulnerable individuals.

  16. Intranasal administration of oxytocin increases compassion toward women.

    PubMed

    Palgi, Sharon; Klein, Ehud; Shamay-Tsoory, Simone G

    2015-03-01

    It has been suggested that the degree of compassion-the feeling of warmth, understanding and kindness that motivates the desire to help others, is modulated by observers' views regarding the target's vulnerability and suffering. This study tested the hypothesis that as compassion developed to protect vulnerable kinships, hormones such as oxytocin, which have been suggested as playing a key role in 'tend-and-befriend' behaviors among women, will enhance compassion toward women but not toward men. Thirty subjects participated in a double-blind, placebo-controlled, within-subject study. Following administration of oxytocin/placebo, participants listened to recordings of different female/male protagonists describing distressful emotional conflicts and were then asked to provide compassionate advice to the protagonist. The participants' responses were coded according to various components of compassion by two clinical psychologists who were blind to the treatment. The results showed that in women and men participants oxytocin enhanced compassion toward women, but did not affect compassion toward men. These findings indicate that the oxytocinergic system differentially mediates compassion toward women and toward men, emphasizing an evolutionary perspective that views compassion as a caregiving behavior designed to help vulnerable individuals. PMID:24711542

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

    PubMed

    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.

  18. Acetylcholine and muscarinic receptor function in cerebral cortex of diabetic young and old male Wistar rats and the role of muscarinic receptors in calcium release from pancreatic islets.

    PubMed

    Savitha, Balakrishnan; Joseph, Binoy; Peeyush Kumar, T; Paulose, C S

    2010-04-01

    We investigated acetylcholine esterase (AChE) activity, acetylcholine and muscarinic M1, M3 receptors kinetics in the cerebral cortex of young and old streptozotocin induced and insulin treated diabetic rats. The role of muscarinic receptors in intracellular calcium release from pancreatic islets was studied in vitro. Wistar rats of 7 and 90-weeks old were used. All studies were done in cerebral cortex. AChE assay was done by spectrophotometric method. Radioreceptor binding assays were done for Acetylcholine, Muscarinic M1 and M3 receptors using specific ligands. Calcium imaging was done using fluo4-AM in pancreatic cells. Ninety-weeks old control rats showed significantly decreased Vmax and increased Km for AChE compared to 7-weeks old control rats. An increased Vmax observed in both 7 and 90-weeks old diabetic groups with significant decrease in Km. Scatchard analysis using specific agonists showed significant decrease in the B (max) and K (d) of acetylcholine and muscarinic M1 receptors in 90-weeks old control rats compared to 7-weeks old control. Binding studies for M3 receptors showed no significant change compared to 7-weeks old control. Acetylcholine, muscarinic M1 and M3 receptor number significantly increased in 90-weeks old diabetic rat groups compared to their respective controls. Insulin treatment significantly reversed the binding parameters to near control compared to diabetic group. In vitro studies showed that acetylcholine through muscarinic M1 and M3 receptors' stimulated calcium release from the pancreatic islets. Thus our studies suggest that Insulin signaling play an important part in differentially regulating pancreatic cholinergic activity, and the diabetes mediated cortical dysfunctions with age.

  19. Risk Factors for Increased Severity of Paediatric Medication Administration Errors

    PubMed Central

    Sears, Kim; Goodman, William M.

    2012-01-01

    Patients' risks from medication errors are widely acknowledged. Yet not all errors, if they occur, have the same risks for severe consequences. Facing resource constraints, policy makers could prioritize factors having the greatest severe–outcome risks. This study assists such prioritization by identifying work-related risk factors most clearly associated with more severe consequences. Data from three Canadian paediatric centres were collected, without identifiers, on actual or potential errors that occurred. Three hundred seventy-two errors were reported, with outcome severities ranging from time delays up to fatalities. Four factors correlated significantly with increased risk for more severe outcomes: insufficient training; overtime; precepting a student; and off-service patient. Factors' impacts on severity also vary with error class: for wrong-time errors, the factors precepting a student or working overtime significantly increase severe-outcomes risk. For other types, caring for an off-service patient has greatest severity risk. To expand such research, better standardization is needed for categorizing outcome severities. PMID:23968607

  20. Peptidergic and muscarinic excitation at amphibian sympathetic synapses.

    PubMed Central

    Kuffler, S W; Sejnowski, T J

    1983-01-01

    A single-electrode voltage clamp was used to study the slow muscarinic and late slow peptidergic excitatory post-synaptic currents (e.p.s.c.s) in B cells of the paravertebral sympathetic ganglia of the bull-frog. Conductance decreases were measured during peptidergic e.p.s.c.s in nearly all cells at clamped potentials near the resting level. In about half of the cells the size of the peptidergic e.p.s.c.s increased with hyperpolarization and in some of these cells conductance increases were found at hyperpolarized levels. In the remaining cells conductance decreases occurred at all levels of membrane potential tested, and in a few of these the polarity of the e.p.s.c.s reversed at hyperpolarized potentials. A similar diversity was observed among muscarinic e.p.s.c.s. At least two simple ionic mechanisms are required to explain the heterogeneous voltage dependencies observed: a conductance decrease primarily to K+ that dominates at depolarized potentials and a conductance increase to other ions that is more prominent at hyperpolarized potentials. The proportion of these two mechanisms appears to differ among B cells. The two slow e.p.s.c.s recorded in the same neurone had the same voltage dependence and were accompanied by the same conductance changes in each of eight cells despite differences between cells. The muscarinic e.p.s.c. was reduced during the peptidergic e.p.s.c. in each of twenty-five neurones tested over a range of membrane potentials. Externally-applied luteinizing hormone releasing hormone (LHRH) produced currents with the same voltage dependence and conductance changes as the nerve-evoked peptidergic e.p.s.c. in each of fifteen cells tested. Bethanechol, a muscarinic agonist, and LHRH produced currents with the same voltage dependence and conductance changes in each of the twelve cells studied. In several cells a saturating response to a prolonged application of LHRH completely occluded the response to bethanechol, and vice versa. Slow currents were

  1. Nucleus accumbens neurotransmission and effort-related choice behavior in food motivation: effects of drugs acting on dopamine, adenosine, and muscarinic acetylcholine receptors.

    PubMed

    Nunes, Eric J; Randall, Patrick A; Podurgiel, Samantha; Correa, Mercè; Salamone, John D

    2013-11-01

    Mesolimbic dopamine (DA) is a critical component of the brain circuitry regulating behavioral activation and effort-related processes. Although nucleus accumbens (NAc) DA depletions or antagonism leave aspects of appetite and primary food motivation intact, rats with impaired DA transmission reallocate their instrumental behavior away from food-reinforced tasks with high response requirements, and instead select less effortful food-seeking behaviors. Previous work showed that adenosine A2A antagonists can reverse the effects of DA D2 antagonists on effort-related choice, and that stimulation of adenosine A2A receptors produces behavioral effects that are similar to those induced by DA antagonism. The present review summarizes the literature on the role of NAc DA and adenosine in effort-related processes, and also presents original data on the effects of local stimulation of muscarinic acetylcholine receptors in NAc core. Local injections of the muscarinic agonist pilocarpine directly into NAc core produces shifts in effort-related choice behavior similar to those induced by DA antagonism or A2A receptor stimulation, decreasing lever pressing but increasing chow intake in rats responding on a concurrent fixed ratio/chow feeding choice task. In contrast, injections into a neostriatal control site dorsal to the NAc were ineffective. The actions of pilocarpine on this task were attenuated by co-administration of the muscarinic antagonist scopolamine. Thus, drugs that act on DA, adenosine A2A, and muscarinic receptors regulate effort-related choice behavior, which may have implications for the treatment of psychiatric symptoms such as psychomotor slowing, fatigue or anergia that can be observed in depression and other disorders.

  2. Memory-improving actions of glucose: involvement of a central cholinergic muscarinic mechanism.

    PubMed

    Kopf, S R; Baratti, C M

    1994-11-01

    Post-training intraperitoneal administration of alpha-D[+]-glucose (10-300 mg/kg) facilitated 24-h retention, in male Swiss mice, of a one-trial step-through inhibitory avoidance task. The dose-response curve was an inverted U. Glucose did not increase the retention latencies of mice that had not received a footshock during training. The effect of glucose (30 mg/kg, ip) on retention was time-dependent, which suggests that the drug facilitated memory storage. The memory facilitation induced by glucose (30 mg/kg, ip) was prevented by atropine (0.5 mg/kg, ip) administered after training, but 10 min prior to glucose treatment. In contrast, neither methylatropine (0.5 mg/kg, ip), a peripherally acting muscarinic receptor blocker, nor mecamylamine (5 mg/kg, ip) or hexamethonium (5 mg/kg, ip), two cholinergic nicotinic receptor antagonists, prevented the effects of post-training glucose on retention. Low subeffective doses of the central acting anticholinesterase physostigmine (35 micrograms/kg, ip), administered immediately after training, and glucose (10 mg/kg, ip), given 10 min after training, acted synergistically to improve retention. The effects of glucose (10 mg/kg, ip) were not influenced by the peripherally acting anticholinesterase neostigmine (35 micrograms/kg, ip). Considered together, these findings suggest that the memory facilitation induced by post-training administration of glucose could result from an enhancement of brain acetylcholine synthesis and/or its release that, in turn, might modulate the activity of muscarinic cholinergic mechanisms that are critically involved in memory storage. PMID:7857246

  3. Memory-improving actions of glucose: involvement of a central cholinergic muscarinic mechanism.

    PubMed

    Kopf, S R; Baratti, C M

    1994-11-01

    Post-training intraperitoneal administration of alpha-D[+]-glucose (10-300 mg/kg) facilitated 24-h retention, in male Swiss mice, of a one-trial step-through inhibitory avoidance task. The dose-response curve was an inverted U. Glucose did not increase the retention latencies of mice that had not received a footshock during training. The effect of glucose (30 mg/kg, ip) on retention was time-dependent, which suggests that the drug facilitated memory storage. The memory facilitation induced by glucose (30 mg/kg, ip) was prevented by atropine (0.5 mg/kg, ip) administered after training, but 10 min prior to glucose treatment. In contrast, neither methylatropine (0.5 mg/kg, ip), a peripherally acting muscarinic receptor blocker, nor mecamylamine (5 mg/kg, ip) or hexamethonium (5 mg/kg, ip), two cholinergic nicotinic receptor antagonists, prevented the effects of post-training glucose on retention. Low subeffective doses of the central acting anticholinesterase physostigmine (35 micrograms/kg, ip), administered immediately after training, and glucose (10 mg/kg, ip), given 10 min after training, acted synergistically to improve retention. The effects of glucose (10 mg/kg, ip) were not influenced by the peripherally acting anticholinesterase neostigmine (35 micrograms/kg, ip). Considered together, these findings suggest that the memory facilitation induced by post-training administration of glucose could result from an enhancement of brain acetylcholine synthesis and/or its release that, in turn, might modulate the activity of muscarinic cholinergic mechanisms that are critically involved in memory storage.

  4. Stimulation of acid secretion and phosphoinositol production by rat parietal cell muscarinic M sub 2 receptors

    SciTech Connect

    Pfeiffer, A.; Rochlitz, H.; Herz, A.; Paumgartner, G. )

    1988-04-01

    The muscarinic receptor system involved in hydrogen production by enriched rat gastric parietal cells was investigated. Muscarinic receptor density determined by (N-methyl-{sup 3}H)scopolamine binding was 8,100/cell. The receptor appeared to be of the M{sub 2} muscarinic receptor subtype, since it had a low affinity (K{sub d} 189 nM) for the M{sub 1} receptor antagonist pirenzepine compared with atropine. Receptor activation by carbachol rapidly augmented levels of polyphosphoinositides, indicating an activation of phospholipase C. The dose-response relations for the increase in inositol phosphates closely paralleled the binding of carbachol to muscarinic receptors. The inositol phosphate response was antagonized by pirenzepine with a K{sub i} of 177 nM. the stimulation of inositol phosphate levels by carbachol correlated well with the stimulation of ({sup 14}C)aminopyrine uptake, determine as an index of acid secretion. The muscarinic agonists oxotremorine, pilocarpine, and bethanechol elicited partial increases in inositol phosphates at maximal drug concentrations, and these partial increases correlated with their ability to stimulate ({sup 14}C)aminopyrine uptake. These data indicate that inositolpolyphosphates may be a second messenger of M{sub 2} receptors stimulating acid secretion.

  5. Imaging muscarinic cholinergic receptors in human brain in vivo with Spect, [123I]4-iododexetimide, and [123I]4-iodolevetimide.

    PubMed

    Müller-Gärtner, H W; Wilson, A A; Dannals, R F; Wagner, H N; Frost, J J

    1992-07-01

    A method to image muscarinic acetylcholine receptors (muscarinic receptors) noninvasively in human brain in vivo was developed using [123I]4-iododexetimide ([123I]IDex), [123I]4-iodolevetimide ([123I]ILev), and single photon emission computed tomography (SPECT). [123I]IDex is a high-affinity muscarinic receptor antagonist. [123I]ILev is its pharmacologically inactive enantiomer and measures nonspecific binding of [123I]IDex in vitro. Regional brain activity after tracer injection was measured in four young normal volunteers for 24 h. Regional [123I]IDex and [123I]ILev activities were correlated early after injection, but not after 1.5 h. [123I]IDex activity increased over 7-12 h in neocortex, neostriatum, and thalamus, but decreased immediately after the injection peak in cerebellum. [123I]IDex activity was highest in neostriatum, followed in rank order by neocortex, thalamus, and cerebellum. [123I]IDex activity correlated with muscarinic receptor concentrations in matching brain regions. In contrast, [123I]ILev activity decreased immediately after the injection peak in all brain regions and did not correspond to muscarinic receptor concentrations. [123I]IDex activity in neocortex and neostriatum during equilibrium was six to seven times higher than [123I]ILev activity. The data demonstrate that [123I]IDex binds specifically to muscarinic receptors in vivo, whereas [123I]ILev represents the nonspecific part of [123I]IDex binding. Subtraction of [123I]ILev from [123I]IDex images on a pixel-by-pixel basis therefore reflects specific [123I]IDex binding to muscarinic receptors. Owing to its high specific binding, [123I]IDex has the potential to measure small changes in muscarinic receptor characteristics in vivo with SPECT. The use of stereoisomerism directly to measure nonspecific binding of [123I]IDex in vivo may reduce complexity in modeling approaches to muscarinic acetylcholine receptors in human brain. PMID:1618935

  6. Impaired vascular responses to parasympathetic nerve stimulation and muscarinic receptor activation in the submandibular gland in nonobese diabetic mice

    PubMed Central

    Berggreen, Ellen; Nyløkken, Krister; Delaleu, Nicolas; Hajdaragic-Ibricevic, Hamijeta; Jonsson, Malin V

    2009-01-01

    Introduction Decreased vascular responses to salivary gland stimulation are observed in Sjögren's syndrome patients. We investigate whether impaired vascular responses to parasympathetic stimulation and muscarinic receptor activation in salivary glands parallels hyposalivation in an experimental model for Sjögren's syndrome. Methods Blood flow responses in the salivary glands were measured by laser Doppler flowmeter. Muscarinic receptor activation was followed by saliva secretion measurements. Nitric oxide synthesis-mediated blood flow responses were studied after administration of a nitric oxide synthase inhibitor. Glandular autonomic nerves and muscarinic 3 receptor distributions were also investigated. Results Maximal blood flow responses to parasympathetic stimulation and muscarinic receptor activation were significantly lower in nonobese diabetic (NOD) mice compared with BALB/c mice, coinciding with impaired saliva secretion in nonobese diabetic mice (P < 0.005). Nitric oxide synthase inhibitor had less effect on blood flow responses after parasympathetic nerve stimulation in nonobese diabetic mice compared with BALB/c mice (P < 0.02). In nonobese diabetic mice, salivary gland parasympathetic nerve fibres were absent in areas of focal infiltrates. Muscarinic 3 receptor might be localized in the blood vessel walls of salivary glands. Conclusions Impaired vasodilatation in response to parasympathetic nerve stimulation and muscarinic receptor activation may contribute to hyposalivation observed in nonobese diabetic mice. Reduced nitric oxide signalling after parasympathetic nerve stimulation may contribute in part to the impaired blood flow responses. The possibility of muscarinic 3 receptor in the vasculature supports the notion that muscarinic 3 receptor autoantibodies present in nonobese diabetic mice might impair the fluid transport required for salivation. Parasympathetic nerves were absent in areas of focal infiltrates, whereas a normal distribution was

  7. Muscarinic receptor agonists stimulate matrix metalloproteinase 1-dependent invasion of human colon cancer cells

    SciTech Connect

    Raufman, Jean-Pierre; Cheng, Kunrong; Saxena, Neeraj; Chahdi, Ahmed; Belo, Angelica; Khurana, Sandeep; Xie, Guofeng

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Muscarinic receptor agonists stimulated robust human colon cancer cell invasion. Black-Right-Pointing-Pointer Anti-matrix metalloproteinase1 antibody pre-treatment blocks cell invasion. Black-Right-Pointing-Pointer Bile acids stimulate MMP1 expression, cell migration and MMP1-dependent invasion. -- Abstract: Mammalian matrix metalloproteinases (MMPs) which degrade extracellular matrix facilitate colon cancer cell invasion into the bloodstream and extra-colonic tissues; in particular, MMP1 expression correlates strongly with advanced colon cancer stage, hematogenous metastasis and poor prognosis. Likewise, muscarinic receptor signaling plays an important role in colon cancer; muscarinic receptors are over-expressed in colon cancer compared to normal colon epithelial cells. Muscarinic receptor activation stimulates proliferation, migration and invasion of human colon cancer cells. In mouse intestinal neoplasia models genetic ablation of muscarinic receptors attenuates carcinogenesis. In the present work, we sought to link these observations by showing that MMP1 expression and activation plays a mechanistic role in muscarinic receptor agonist-induced colon cancer cell invasion. We show that acetylcholine, which robustly increases MMP1 expression, stimulates invasion of HT29 and H508 human colon cancer cells into human umbilical vein endothelial cell monolayers - this was abolished by pre-incubation with atropine, a non-selective muscarinic receptor inhibitor, and by pre-incubation with anti-MMP1 neutralizing antibody. Similar results were obtained using a Matrigel chamber assay and deoxycholyltaurine (DCT), an amidated dihydroxy bile acid associated with colon neoplasia in animal models and humans, and previously shown to interact functionally with muscarinic receptors. DCT treatment of human colon cancer cells resulted in time-dependent, 10-fold increased MMP1 expression, and DCT-induced cell invasion was also blocked by pre

  8. Muscarine binding sites in bovine adrenal medulla.

    PubMed

    Barron, B A; Murrin, L C; Hexum, T D

    1986-03-18

    The presence of muscarinic binding sites in the bovine adrenal medulla was investigated using [3H]QNB and the bovine adrenal medulla. Scatchard analysis combined with computer analysis yielded data consistent with a two binding site configuration. KDs of 0.15 and 14 nM and Bmax s of 29 and 210 fmol/mg protein, respectively, were observed. Displacement of [3H]QNB by various cholinergic agents is, in order of decreasing potency: QNB, dexetimide, atropine, scopolamine, imipramine, desipramine, oxotremorine, pilocarpine, acetylcholine, methacholine and carbachol. These results demonstrate the presence of more than one muscarine binding site in the bovine adrenal gland. PMID:3709656

  9. Muscarinic preferential M(1) receptor antagonists enhance the discriminative-stimulus effects of cocaine in rats.

    PubMed

    Tanda, Gianluigi; Katz, Jonathan L

    2007-10-01

    Previous studies of benztropine analogues have found them to inhibit dopamine uptake like cocaine, but with less effectiveness than cocaine in producing behavioral effects related to drug abuse. Studies have assessed whether nonselective muscarinic antagonists decrease the effects of cocaine because many of the benztropine analogues are also muscarinic antagonists. As previous studies were conducted with nonselective muscarinic antagonists and the benztropine analogues show preferential affinity for the M(1) muscarinic receptor subtype, the present study examined interactions of cocaine and the preferential M(1) antagonists, telenzepine (TZP) and trihexyphenidyl (TXP) on subjective effects in rats trained to discriminate cocaine (10 mg/kg, i.p.) from saline injections. Cocaine dose-dependently increased the percentage of responses on the cocaine-appropriate lever, with full substitution at the training dose. In contrast neither TZP nor TXP produced more than 25% cocaine-appropriate responding at any dose. Both M(1) antagonists produced significant leftward shifts in the cocaine dose-effect curve, TZP at 3.0 and TXP at 0.3 and 1.0 mg/kg. The present results indicate that preferential antagonist actions at muscarinic M(1) receptors enhance rather than attenuate the discriminative-stimulus effects of cocaine, and thus those actions unlikely contribute to the reduced cocaine-like effects of BZT analogues.

  10. Midlevel Administrators' Pay Increases Slightly but Doesn't Match Inflation

    ERIC Educational Resources Information Center

    Fuller, Andrea

    2012-01-01

    Salaries for midlevel administrators rose by a median of 2 percent this year over last year, matching the median pay increase for senior administrators and coming in slightly higher than the 1.9-percent median increase for faculty members, says an annual report released by the College and University Professional Association for Human Resources.…

  11. Organophosphorus Pesticides Decrease M2 Muscarinic Receptor Function in Guinea Pig Airway Nerves via Indirect Mechanisms

    PubMed Central

    Proskocil, Becky J.; Bruun, Donald A.; Thompson, Charles M.; Fryer, Allison D.; Lein, Pamela J.

    2010-01-01

    Background Epidemiological studies link organophosphorus pesticide (OP) exposures to asthma, and we have shown that the OPs chlorpyrifos, diazinon and parathion cause airway hyperreactivity in guinea pigs 24 hr after a single subcutaneous injection. OP-induced airway hyperreactivity involves M2 muscarinic receptor dysfunction on airway nerves independent of acetylcholinesterase (AChE) inhibition, but how OPs inhibit neuronal M2 receptors in airways is not known. In the central nervous system, OPs interact directly with neurons to alter muscarinic receptor function or expression; therefore, in this study we tested whether the OP parathion or its oxon metabolite, paraoxon, might decrease M2 receptor function on peripheral neurons via similar direct mechanisms. Methodology/Principal Findings Intravenous administration of paraoxon, but not parathion, caused acute frequency-dependent potentiation of vagally-induced bronchoconstriction and increased electrical field stimulation (EFS)-induced contractions in isolated trachea independent of AChE inhibition. However, paraoxon had no effect on vagally-induced bradycardia in intact guinea pigs or EFS-induced contractions in isolated ileum, suggesting mechanisms other than pharmacologic antagonism of M2 receptors. Paraoxon did not alter M2 receptor expression in cultured cells at the mRNA or protein level as determined by quantitative RT-PCR and radio-ligand binding assays, respectively. Additionally, a biotin-labeled fluorophosphonate, which was used as a probe to identify molecular targets phosphorylated by OPs, did not phosphorylate proteins in guinea pig cardiac membranes that were recognized by M2 receptor antibodies. Conclusions/Significance These data indicate that neither direct pharmacologic antagonism nor downregulated expression of M2 receptors contributes to OP inhibition of M2 function in airway nerves, adding to the growing evidence of non-cholinergic mechanisms of OP neurotoxicity. PMID:20479945

  12. Molecular properties of muscarinic acetylcholine receptors

    PubMed Central

    HAGA, Tatsuya

    2013-01-01

    Muscarinic acetylcholine receptors, which comprise five subtypes (M1-M5 receptors), are expressed in both the CNS and PNS (particularly the target organs of parasympathetic neurons). M1-M5 receptors are integral membrane proteins with seven transmembrane segments, bind with acetylcholine (ACh) in the extracellular phase, and thereafter interact with and activate GTP-binding regulatory proteins (G proteins) in the intracellular phase: M1, M3, and M5 receptors interact with Gq-type G proteins, and M2 and M4 receptors with Gi/Go-type G proteins. Activated G proteins initiate a number of intracellular signal transduction systems. Agonist-bound muscarinic receptors are phosphorylated by G protein-coupled receptor kinases, which initiate their desensitization through uncoupling from G proteins, receptor internalization, and receptor breakdown (down regulation). Recently the crystal structures of M2 and M3 receptors were determined and are expected to contribute to the development of drugs targeted to muscarinic receptors. This paper summarizes the molecular properties of muscarinic receptors with reference to the historical background and bias to studies performed in our laboratories. PMID:23759942

  13. Batrachotoxin changes the properties of the muscarinic receptor in rat brain and heart: possible interaction(s) between muscarinic receptors and sodium channels.

    PubMed Central

    Cohen-Armon, M; Kloog, Y; Henis, Y I; Sokolovsky, M

    1985-01-01

    The effects of Na+-channel activator batrachotoxin (BTX) on the binding properties of muscarinic receptors in homogenates of rat brain and heart were studied. BTX enhanced the affinity for the binding of the agonists carbamoylcholine and acetylcholine to the muscarinic receptors in brainstem and ventricle, but not in the cerebral cortex. Analysis of the data according to a two-site model for agonist binding indicated that the effect of BTX was to increase the affinity of the agonists to the high-affinity site. Guanyl nucleotides, known to induce interconversion of high-affinity agonist binding sites to the low-affinity state, canceled the effect of BTX on carbamoylcholine and acetylcholine binding. BTX had no effect on the binding of the agonist oxotremorine or on the binding of the antagonist [3H]-N-methyl-4-piperidyl benzilate. The local anesthetics dibucaine and tetracaine antagonized the effect of BTX on the binding of muscarinic agonists at concentrations known to inhibit the activation of Na+ channels by BTX. On the basis of these findings, we propose that in specific tissues the muscarinic receptors may interact with the BTX binding site (Na+ channels). PMID:2582418

  14. Extension Master Gardener Intranet: Automating Administration, Motivating Volunteers, Increasing Efficiency, and Facilitating Impact Reporting

    ERIC Educational Resources Information Center

    Bradley, Lucy K.; Cook, Jonneen; Cook, Chris

    2011-01-01

    North Carolina State University has incorporated many aspects of volunteer program administration and reporting into an on-line solution that integrates impact reporting into daily program management. The Extension Master Gardener Intranet automates many of the administrative tasks associated with volunteer management, increasing efficiency, and…

  15. Binding and functional characterization of the cardioselective muscarinic antagonist methoctramine.

    PubMed

    Giraldo, E; Micheletti, R; Montagna, E; Giachetti, A; Viganò, M A; Ladinsky, H; Melchiorre, C

    1988-03-01

    The antimuscarinic properties of the newly synthetized polymethylene tetramine derivative, methoctramine, were investigated in binding and functional assays. Methoctramine displaced the specific binding of [3H]-N-methylscopolamine [( 3H]NMS) and [3H] pirenzepine from membranes of rat tissues with the following order of affinities: heart = cerebellum greater than cortex greater than submandibular glands, the ratio of the affinities of the compound for the heart and the glands amounting to about 130. Computer fits of binding curves generated in cardiac and cortical membranes were compatible with an interaction at one binding site, whereas those in submandibular glands and cerebellum had slopes significantly lower than 1. Experiments performed in cardiac membranes to investigate the effect of methoctramine on the dissociation kinetics of [3H]-NMS showed that concentrations of compound up to 1 microM did not affect the dissociation of [3H]-NMS elicited by an excess of NMS. At greater concentrations (10-100 microM), methoctramine dose dependently inhibited [3H]-NMS dissociation, thus revealing an allosteric interaction. In in vitro functional assays, methoctramine displayed more than 100 times greater affinity for the muscarinic receptors mediating negative inotropic and chronotropic effects in guinea pig atria than for those responsible for tracheal contraction. Similarly, the compound was a more potent antagonist of the bradycardial response to bethanechol than of the bladder tonus increase, saliva secretion and hypotension induced by the muscarinic agonist in anesthetized cats. Finally, in the pithed rat, methoctramine preferentially inhibited cardiac M2 (vagal bradycardia) over ganglionic M1 (McN-A-343-induced hypertension) responses. The evidence appears to characterize methoctramine as being the most selective M2 muscarinic antagonist described to date.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3252019

  16. Coenzyme Q10 Administration Increases Brain Mitochondrial Concentrations and Exerts Neuroprotective Effects

    NASA Astrophysics Data System (ADS)

    Matthews, Russell T.; Yang, Lichuan; Browne, Susan; Baik, Myong; Flint Beal, M.

    1998-07-01

    Coenzyme Q10 is an essential cofactor of the electron transport chain as well as a potent free radical scavenger in lipid and mitochondrial membranes. Feeding with coenzyme Q10 increased cerebral cortex concentrations in 12- and 24-month-old rats. In 12-month-old rats administration of coenzyme Q10 resulted in significant increases in cerebral cortex mitochondrial concentrations of coenzyme Q10. Oral administration of coenzyme Q10 markedly attenuated striatal lesions produced by systemic administration of 3-nitropropionic acid and significantly increased life span in a transgenic mouse model of familial amyotrophic lateral sclerosis. These results show that oral administration of coenzyme Q10 increases both brain and brain mitochondrial concentrations. They provide further evidence that coenzyme Q10 can exert neuroprotective effects that might be useful in the treatment of neurodegenerative diseases.

  17. Synaptic muscarinic response types in hippocampal CA1 interneurons depend on different levels of presynaptic activity and different muscarinic receptor subtypes.

    PubMed

    Bell, L Andrew; Bell, Karen A; McQuiston, A Rory

    2013-10-01

    Depolarizing, hyperpolarizing and biphasic muscarinic responses have been described in hippocampal inhibitory interneurons, but the receptor subtypes and activity patterns required to synaptically activate muscarinic responses in interneurons have not been completely characterized. Using optogenetics combined with whole cell patch clamp recordings in acute slices, we measured muscarinic responses produced by endogenously released acetylcholine (ACh) from cholinergic medial septum/diagonal bands of Broca inputs in hippocampal CA1. We found that depolarizing responses required more cholinergic terminal stimulation than hyperpolarizing ones. Furthermore, elevating extracellular ACh with the acetylcholinesterase inhibitor physostigmine had a larger effect on depolarizing versus hyperpolarizing responses. Another subpopulation of interneurons responded biphasically, and periodic release of ACh entrained some of these interneurons to rhythmically burst. M4 receptors mediated hyperpolarizing responses by activating inwardly rectifying K(+) channels, whereas the depolarizing responses were inhibited by the nonselective muscarinic antagonist atropine but were unaffected by M1, M4 or M5 receptor modulators. In addition, activation of M4 receptors significantly altered biphasic interneuron firing patterns. Anatomically, interneuron soma location appeared predictive of muscarinic response types but response types did not correlate with interneuron morphological subclasses. Together these observations suggest that the hippocampal CA1 interneuron network will be differentially affected by cholinergic input activity levels. Low levels of cholinergic activity will preferentially suppress some interneurons via hyperpolarization and increased activity will recruit other interneurons to depolarize, possibly because of elevated extracellular ACh concentrations. These data provide important information for understanding how cholinergic therapies will affect hippocampal network function

  18. Allosteric activation of M4 muscarinic receptors improve behavioral and physiological alterations in early symptomatic YAC128 mice

    PubMed Central

    Pancani, Tristano; Foster, Daniel J.; Moehle, Mark S.; Bichell, Terry Jo; Bradley, Emma; Bridges, Thomas M.; Klar, Rebecca; Poslusney, Mike; Rook, Jerri M.; Daniels, J. Scott; Niswender, Colleen M.; Jones, Carrie K.; Wood, Michael R.; Bowman, Aaron B.; Lindsley, Craig W.; Xiang, Zixiu; Conn, P. Jeffrey

    2015-01-01

    Mutations that lead to Huntington’s disease (HD) result in increased transmission at glutamatergic corticostriatal synapses at early presymptomatic stages that have been postulated to set the stage for pathological changes and symptoms that are observed at later ages. Based on this, pharmacological interventions that reverse excessive corticostriatal transmission may provide a novel approach for reducing early physiological changes and motor symptoms observed in HD. We report that activation of the M4 subtype of muscarinic acetylcholine receptor reduces transmission at corticostriatal synapses and that this effect is dramatically enhanced in presymptomatic YAC128 HD and BACHD relative to wild-type mice. Furthermore, chronic administration of a novel highly selective M4 positive allosteric modulator (PAM) beginning at presymptomatic ages improves motor and synaptic deficits in 5-mo-old YAC128 mice. These data raise the exciting possibility that selective M4 PAMs could provide a therapeutic strategy for the treatment of HD. PMID:26508634

  19. External Imaging of Cerebral Muscarinic Acetylcholine Receptors

    NASA Astrophysics Data System (ADS)

    Eckelman, William C.; Reba, Richard C.; Rzeszotarski, Waclaw J.; Gibson, Raymond E.; Hill, Thomas; Holman, B. Leonard; Budinger, Thomas; Conklin, James J.; Eng, Robert; Grissom, Michael P.

    1984-01-01

    A radioiodinated ligand that binds to muscarinic acetylcholine receptors was shown to distribute in the brain by a receptor-mediated process. With single-photon-emission imaging techniques, radioactivity was detected in the cerebrum but not in the cerebellum, whereas with a flow-limited radiotracer, radioactivity was detected in cerebrum and cerebellum. Single-photon-emission computed tomography showed good definition of the caudate putamen and cortex in man.

  20. External imaging of cerebral muscarinic acetylcholine receptors

    SciTech Connect

    Eckelman, W.C.; Reba, R.C.; Rzeszotarski, W.J.; Gibson, R.E.; Hill, T.; Holman, B.L.; Budinger, T.; Conklin, J.J.; Eng, R.; Grissom, M.P.

    1984-01-20

    A radioiodinated ligand that binds to muscarinic acetylcholine receptors was shown to distribute in the brain by a receptor-mediated process. With single-photon-emission imaging techniques, radioactivity was detected in the cerebrum but not in the cerebellum, whereas with a flow-limited radiotracer, radioactivity was detected in cerebrum and cerebellum. Single-photon-emission computed tomography showed good definition of the caudate putamen and cortex in man.

  1. Carbachol dimers as homobivalent modulators of muscarinic receptors.

    PubMed

    Matucci, Rosanna; Nesi, Marta; Martino, Maria Vittoria; Bellucci, Cristina; Manetti, Dina; Ciuti, Elisa; Mazzolari, Angelica; Dei, Silvia; Guandalini, Luca; Teodori, Elisabetta; Vistoli, Giulio; Romanelli, Maria Novella

    2016-05-15

    A series of homodimers of the well-known cholinergic agonist carbachol have been synthesized, showing the two agonist units symmetrically connected through a methylene chain of variable length. The new compounds have been tested on the five cloned muscarinic receptors (hM1-5) expressed in CHO cells by means of equilibrium binding studies, showing an increase in affinity by rising the number of methylene units up to 7 and 9. Functional experiments on guinea-pig ileum and assessment of ERK1/2 phosphorylation on hM1, hM2 and hM3 on CHO cells have shown that the new compounds are endowed with muscarinic antagonistic properties. Kinetic binding studies have revealed that some of the tested compounds are able to slow the rate of dissociation of NMS, suggesting a bitopic behavior. Docking simulations, performed on the hM1 and hM2 receptors, give a sound rationalization of the experimental data revealing how these compounds are able to interact with both orthosteric and allosteric binding sites depending on the length of their connecting chain. PMID:26996304

  2. Ethanol inhibits neuritogenesis induced by astrocyte muscarinic receptors.

    PubMed

    Guizzetti, Marina; Moore, Nadia H; Giordano, Gennaro; VanDeMark, Kathryn L; Costa, Lucio G

    2010-09-01

    In utero alcohol exposure can lead to fetal alcohol spectrum disorders, characterized by cognitive and behavioral deficits. In vivo and in vitro studies have shown that ethanol alters neuronal development. We have recently shown that stimulation of M(3) muscarinic receptors in astrocytes increases the synthesis and release of fibronectin, laminin, and plasminogen activator inhibitor-1, causing neurite outgrowth in hippocampal neurons. As M(3) muscarinic receptor signaling in astroglial cells is strongly inhibited by ethanol, we hypothesized that ethanol may also inhibit neuritogenesis in hippocampal neurons induced by carbachol-stimulated astrocytes. In the present study, we report that the effect of carbachol-stimulated astrocytes on hippocampal neuron neurite outgrowth was inhibited in a concentration-dependent manner (25-100 mM) by ethanol. This effect was because of the inhibition of the release of fibronectin, laminin, and plasminogen activator inhibitor-1. Similar effects on neuritogenesis and on the release of astrocyte extracellular proteins were observed after the incubation of astrocytes with carbachol in the presence of 1-butanol, another short-chain alcohol, which like ethanol is a competitive substrate for phospholipase D, but not by tert-butanol, its analog that is not a substrate for this enzyme. This study identifies a potential novel mechanism involved in the developmental effects of ethanol mediated by the interaction of ethanol with cell signaling in astrocytes, leading to an impairment in neuron-astrocyte communication.

  3. Homology modeling of human muscarinic acetylcholine receptors.

    PubMed

    Thomas, Trayder; McLean, Kimberley C; McRobb, Fiona M; Manallack, David T; Chalmers, David K; Yuriev, Elizabeth

    2014-01-27

    We have developed homology models of the acetylcholine muscarinic receptors M₁R-M₅R, based on the β₂-adrenergic receptor crystal as the template. This is the first report of homology modeling of all five subtypes of acetylcholine muscarinic receptors with binding sites optimized for ligand binding. The models were evaluated for their ability to discriminate between muscarinic antagonists and decoy compounds using virtual screening using enrichment factors, area under the ROC curve (AUC), and an early enrichment measure, LogAUC. The models produce rational binding modes of docked ligands as well as good enrichment capacity when tested against property-matched decoy libraries, which demonstrates their unbiased predictive ability. To test the relative effects of homology model template selection and the binding site optimization procedure, we generated and evaluated a naïve M₂R model, using the M₃R crystal structure as a template. Our results confirm previous findings that binding site optimization using ligand(s) active at a particular receptor, i.e. including functional knowledge into the model building process, has a more pronounced effect on model quality than target-template sequence similarity. The optimized M₁R-M₅R homology models are made available as part of the Supporting Information to allow researchers to use these structures, compare them to their own results, and thus advance the development of better modeling approaches.

  4. Muscarinic receptor subtypes mediating the mucosal response to neural stimulation of guinea pig ileum

    SciTech Connect

    Carey, H.V.; Tien, X.Y.; Wallace, L.J.; Cooke, H.J.

    1987-09-01

    Muscarinic receptors involved in the secretory response evoked by electrical stimulation of submucosal neutrons were investigated in muscle-stripped flat sheets of guinea pig ileum set up in flux chambers. Neural stimulation produced a biphasic increase in short-circuit current due to active chloride secretion. Atropine and 4-diphenylacetoxy-N-methylpiperadine methiodide (4-DAMP) (10/sup -7/ M) were more potent inhibitors of the cholinergic phase of the response than was pirenzepine. Dose-dependent increases in base-line short-circuit current were evoked by carbachol and bethanechol; 4-hydroxy-2-butynyl trimethylammonium chloride (McN A343) produced a much smaller effect. Tetrodotoxin abolished the effects of McN A343 but did not alter the responses of carbachol and bethanechol. McN A343 significantly reduced the cholinergic phase of the neurally evoked response and caused a rightward shift of the carbachol dose-response curve. All muscarinic compounds inhibited (/sup 3/H)quinuclidinyl benzilate binding to membranes from muscosal scrapings, with a rank order of potency of 4-DAMP > pirenzepine > McN A343 > carbachol > bethanechol. These results suggest that acetylcholine released from submucosal neurons mediates chloride secretion by interacting with muscarinic cholinergic receptors that display a high binding affinity for 4-DAMP. Activation of neural muscarinic receptors makes a relatively small contribution to the overall secretory response.

  5. Reduction of ventricular M2 muscarinic receptors in cardiomyopathic hamster (CHF 147) at the necrotic stage of the myopathy.

    PubMed

    Wilkinson, M; Horackova, M; Giles, A

    1994-04-01

    We have previously demonstrated that isolated ventricular myocytes from cardiomyopathic hamsters (CHF 147) during the necrotic stage (70-100 days) exhibit an attenuated contractile response to muscarinic stimulation. In the present study we have investigated whether this dysfunction may be related to a change in the density (or affinity) of cardiac muscarinic receptors. Thus, we have characterized and quantified the binding of the muscarinic antagonist [3H]-N-methyl scopolamine (NMS) to M2 muscarinic receptors in cardiac micropunches and in suspensions of isolated intact cardiomyocytes obtained from cardiomyopathic (CHF 147) and Golden Syrian hamsters. The hamsters were either 70-100 days old, when the cardiomyopathy had reached the cytolytic and necrotic stage or 30 days old, i.e. before the onset of the cardiomyopathy. In both preparations (micropunches and dissociated cardiomyocytes) the specific binding of [3H]-NMS was stereospecific, reversible, saturable, of high affinity and linearly dependent upon increasing amounts of tissue and cells. The binding site also possessed the drug specificity typical of an M2 muscarinic receptor. Saturation binding analysis revealed that the hearts of the older CHF 147 hamsters contain significantly fewer M2 muscarinic receptors than the control Golden Syrian hamsters while the affinity (Kd) was not altered. This reduction of M2 receptor number was not observed in CHF 147 hamsters at 30 days. Further, we found no differences in beta-adrenergic or in alpha 1-adrenergic binding in the two strains of hamster at either age. Thus, our results indicate that the parasympathetic regulation of cardiac function in CHF 147 hamsters may be compromised by a decreased number of muscarinic receptors at the necrotic stage of the cardiomyopathy.

  6. Muscarinic Ca2+ responses resistant to muscarinic antagonists at perisynaptic Schwann cells of the frog neuromuscular junction.

    PubMed Central

    Robitaille, R; Jahromi, B S; Charlton, M P

    1997-01-01

    1. Acetylcholine causes a rise of intracellular Ca2+ in perisynaptic Schwann cells (PSCs) of the frog neuromuscular junction. The signalling pathway was characterized using the fluorescent Ca2+ indicator fluo-3 and fluorescence microscopy. 2. Nicotinic antagonists had no effect on Ca2+ responses evoked by ACh and no Ca2+ responses were evoked with the nicotinic agonist nicotine. The muscarinic agonists muscarine and oxotremorine-M induced Ca2+ signals in PSCs. 3. Ca2+ responses remained unchanged when extracellular Ca2+ was removed, indicating that they are due to the release of Ca2+ from internal stores. Incubation with pertussis toxin did not alter the Ca2+ signals induced by muscarine, but did block depression of transmitter release induced by adenosine and prevented Ca2+ responses in PSCs induced by adenosine. 4. The general muscarinic antagonists atropine, quinuclidinyl benzilate and N-methyl-scopolamine failed to block Ca2+ responses to muscarinic agonists. Atropine (at 20,000-fold excess concentration) also failed to reduce the proportion of cells responding to a threshold muscarine concentration sufficient to cause responses in less than 50% of cells. Only the allosteric, non-specific blocker, gallamine (1-10 microM) was effective in blocking muscarine-induced Ca2+ responses. 5. In preparations denervated 7 days prior to experiments, low concentrations of atropine reversibly and completely blocked Ca2+ responses to muscarine. 6. The lack of blockade by general muscarinic antagonists in innervated, in situ preparations suggests that muscarinic Ca2+ responses at PSCs are not mediated by any of the five known muscarinic receptors or that post-translational modification prevented antagonist binding. Images Figure 2 Figure 3 Figure 4 Figure 6 Figure 7 PMID:9365908

  7. Type 3 Muscarinic Receptors Contribute to Clearance of Citrobacter rodentium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although the alpha 7 nicotinic receptor exerts anti-inflammatory effects on immune cells, the role of muscarinic receptors in mucosal homeostasis, response to enteric pathogens, and modulation of immune cell function is undefined. The contribution of type 3 muscarinic receptor (M3R) to mucosal homeo...

  8. The Michigan Model Pilot: Increasing the Number of Female Administrators in Michigan Public Schools.

    ERIC Educational Resources Information Center

    Michigan State Dept. of Education, Lansing.

    The Michigan Model provides a 3-year plan for school districts to increase the number of women in administration. Nineteen objectives address six key role groups--three external to the school district (professional educational organizations, local community groups, and parent/community members) and three internal to the district…

  9. The Leadership Roles of Distance Learning Administrators (DLAs) in Increasing Educational Value and Quality Perceptions

    ERIC Educational Resources Information Center

    McFarlane, Donovan A.

    2011-01-01

    This paper examines the leadership roles of distance learning administrators (DLAs) in light of the demand and need for value and quality in educational distance learning programs and schools. The author explores the development of distance learning using available and emerging technologies in relation to increased demand for education, training,…

  10. Alcohol administration increases cocaine craving but not cocaine cue attentional bias

    PubMed Central

    Marks, Katherine R.; Pike, Erika; Stoops, William W.; Rush, Craig R.

    2015-01-01

    Background Alcohol consumption is a known antecedent to cocaine relapse. Through associative conditioning, it is hypothesized that alcohol increases incentive motivation for cocaine and thus the salience of cocaine-related cues, which are important in maintaining drug-taking behavior. Cocaine-using individuals display a robust cocaine cue attentional bias as measured by fixation time during the visual probe task. The purpose of the present study was to evaluate the influence of alcohol administration on cocaine cue attentional bias using eye-tracking technology to directly measure attentional allocation. Methods Twenty current cocaine users completed a double-blind, placebo-controlled, within-subjects study that tested the effect of three doses of alcohol (0.00, 0.325, 0.65 g/kg alcohol) on cocaine cue attentional bias using the visual probe task with eye-tracking technology. The participant-rated and physiological effects of alcohol were also assessed. Results Participants displayed a robust cocaine cue attentional bias following both placebo and alcohol administration as measured by fixation time, but not response time. Alcohol administration did not influence cocaine cue attentional bias, but increased craving for cocaine in a dose dependent manner. Alcohol produced prototypic psychomotor and participant-rated effects. Conclusions Alcohol administration increases cocaine craving but not cocaine cue attentional bias. Alcohol-induced cocaine craving suggests that alcohol increases incentive motivation for cocaine but not the salience of cocaine-related cues. PMID:26331880

  11. Direct muscarinic and nicotinic receptor-mediated excitation of rat medial vestibular nucleus neurons in vitro

    NASA Technical Reports Server (NTRS)

    Phelan, K. D.; Gallagher, J. P.

    1992-01-01

    We have utilized intracellular recording techniques to investigate the cholinoceptivity of rat medial vestibular nucleus (MVN) neurons in a submerged brain slice preparation. Exogenous application of the mixed cholinergic agonists, acetylcholine (ACh) or carbachol (CCh), produced predominantly membrane depolarization, induction of action potential firing, and decreased input resistance. Application of the selective muscarinic receptor agonist muscarine (MUSC), or the selective nicotinic receptor agonists nicotine (NIC) or 1,1-dimethyl-4-phenylpiperazinium (DMPP) also produced membrane depolarizations. The MUSC-induced depolarization was accompanied by decreased conductance, while an increase in conductance appeared to underlie the NIC- and DMPP-induced depolarizations. The muscarinic and nicotinic receptor mediated depolarizations persisted in tetrodotoxin and/or low Ca2+/high Mg2+ containing media, suggesting direct postsynaptic receptor activation. The MUSC-induced depolarization could be reversibly blocked by the selective muscarinic-receptor antagonist, atropine, while the DMPP-induced depolarization could be reversibly suppressed by the selective ganglionic nicotinic-receptor antagonist, mecamylamine. Some neurons exhibited a transient membrane hyperpolarization during the depolarizing response to CCh or MUSC application. This transient inhibition could be reversibly blocked by the gamma-aminobutyric acid (GABA) antagonist, bicuculline, suggesting that the underlying hyperpolarization results indirectly from the endogenous release of GABA acting at GABA receptors. This study confirms the cholinoceptivity of MVN neurons and establishes that individual MVN cells possess muscarinic as well as nicotinic receptors. The data provide support for a prominent role of cholinergic mechanisms in the direct and indirect regulation of the excitability of MVN neurons.

  12. Nicotine decreases ethanol-induced dopamine signaling and increases self-administration via stress hormones.

    PubMed

    Doyon, William M; Dong, Yu; Ostroumov, Alexey; Thomas, Alyse M; Zhang, Tao A; Dani, John A

    2013-08-01

    Tobacco smoking is a well-known risk factor for subsequent alcohol abuse, but the neural events underlying this risk remain largely unknown. Alcohol and nicotine reinforcement involve common neural circuitry, including the mesolimbic dopamine system. We demonstrate in rodents that pre-exposure to nicotine increases alcohol self-administration and decreases alcohol-induced dopamine responses. The blunted dopamine response was due to increased inhibitory synaptic transmission onto dopamine neurons. Blocking stress hormone receptors prior to nicotine exposure prevented all interactions with alcohol that we measured, including the increased inhibition onto dopamine neurons, the decreased dopamine responses, and the increased alcohol self-administration. These results indicate that nicotine recruits neuroendocrine systems to influence neurotransmission and behavior associated with alcohol reinforcement.

  13. Characterization of muscarinic receptors in rat kidney.

    PubMed

    Blankesteijn, W M; Siero, H L; Rodrigues de Miranda, J F; van Megen, Y J; Russel, F G

    1993-01-01

    Muscarinic receptors in mammalian kidney seem to be involved in diuresis. In this study we give a detailed characterization of receptors in rat kidney. Specific binding of [3H](-)-quinuclidinylbenzilate ([3H]QNB) to membranes of rat kidney cortex was saturable and of high affinity. A dissociation constant of 0.063 +/- 0.003 nM and a receptor density of 1.46 +/- 0.07 pmol/g wet weight were obtained. The dissociation kinetics could be best described by assuming a mono-exponential function (k-1 = (0.52 +/- 0.1) x 10(-4) s-1). The binding of [3H]QNB reached a maximum in 60 min at 0.6 nM at 37 degrees C. Competition experiments with the enantiomers of benzetimide confirmed the muscarinic nature of the [3H]QNB binding sites. The inhibition constants of pirenzepine (0.23 +/- 0.02 microM), (+-)-hexahydrosiladifenidol (0.040 +/- 0.002 microM), AF-DX 116 (1.45 +/- 0.07 microM), methoctramine (1.67 +/- 0.02 microM) and gallamine (78 +/- 3 microM) classified this receptor as an M3 receptor. Inhibition of [3H]QNB binding by the agonists methylfurtrethonium, arecoline, isoarecoline methiodide, arecaidine propargyl ester and McN-A-343 displayed monophasic inhibition curves. With (+/-)-cis-2-methyl-4-dimethylaminomethyl-1,3- dioxolane methiodide in two out of four experiments a small (11%) population of high affinity agonist sites could be detected. The potassium sparing diuretic amiloride inhibited [3H]QNB binding (36 +/- 3 microM). Although in a way related to the amiloride binding site, the muscarinic receptors in rat kidney are unlikely to be the primary target of diuretic action of this drug. PMID:8420789

  14. Cholinergic muscarinic receptors in rat cochlea.

    PubMed

    van Megen, Y J; Klaassen, A B; Rodrigues de Miranda, J F; Kuijpers, W

    1988-11-22

    Specific 3H-1-quinuclidinylbenzilate (3H-1-QNB) binding to rat cochlea homogenates occurs to a homogeneous class of binding sites with Kd = 0.13 +/- 0.01 nM and Bmax = 0.57 +/- 0.07 fmol per cochlea. Binding is stereoselectively inhibited by benzetimide enantiomers. Dexetimide was more effective than levetimide in displacing 3H-1-QNB from its binding sites (Ki = 4 x 10(-10) M and 6.5 x 10(-6) M, respectively). Pirenzepine inhibits 3H-1-QNB binding with low affinity (Ki = 2 x 10(-6) M), classifying the binding sites as muscarinic M2 receptors. PMID:3214711

  15. Muscarinic Preferential M1 Receptor Antagonists Minimally Alter the Discriminative-Stimulus Effects of Cocaine in Rats

    PubMed Central

    Tanda, Gianluigi; Katz, Jonathan L.

    2009-01-01

    Previous studies of benztropine analogues have found them to inhibit dopamine uptake like cocaine, but with less effectiveness than cocaine in producing behavioral effects related to drug abuse. Studies have assessed whether nonselective muscarinic antagonists decrease the effects of cocaine because many of the benztropine analogues are also muscarinic antagonists. As previous studies were conducted with nonselective muscarinic antagonists and the benztropine analogues show preferential affinity for the M1 muscarinic receptor subtype, the present study examined interactions of cocaine and the preferential M1 antagonists, telenzepine (TZP) and trihexyphenidyl (TXP) on subjective effects in rats trained to discriminate cocaine (10 mg/kg, i.p.) from saline injections. Cocaine dose-dependently increased the percentage of responses on the cocaine-appropriate lever, with full substitution at the training dose. In contrast neither TZP nor TXP produced more than 25% cocaine-appropriate responding at any dose. Both M1 antagonists produced significant leftward shifts in the cocaine dose-effect curve, TZP at 3.0 and TXP at 0.3 and 1.0 mg/kg. The present results indicate that preferential antagonist actions at muscarinic M1 receptors enhance rather than attenuate the discriminative-stimulus effects of cocaine, and thus those actions unlikely contribute to the reduced cocaine-like effects of BZT analogues. PMID:17631384

  16. Chronic administration of nalmefene leads to increased food intake and body weight gain in mice.

    PubMed

    Chen, Richard Z; Huang, Ruey-Ruey C; Shen, Chun-Pyn; MacNeil, Douglas J; Fong, Tung M

    2004-07-01

    Nalmefene is an orally available opioid receptor antagonist that has been shown to suppress appetite in humans, but its effects on chronic food intake and body weight remain unclear. Here, we report that chronic (21-day) oral administration of nalmefene at 2 or 10 mg/kg/day in diet-induced obese (DIO) mice led to significant increases (9-11%) in cumulative food intake. Mice in the nalmefene-treated groups also gained body weight at a rate faster than the control. Body composition analysis showed that the extra body weight gains in the treated animals were mostly due to increased fat accumulation. Since acute nalmefene treatment showed a trend toward a decrease rather than an increase in food intake, it is possible that the orexigenic effect of chronic oral administration of nalmefene was caused by pharmacologically active metabolites rather than the drug itself. Our results argue against the potential use of nalmefene for treating human obesity.

  17. A Single Oral Administration of Theaflavins Increases Energy Expenditure and the Expression of Metabolic Genes

    PubMed Central

    Kudo, Naoto; Arai, Yasunori; Suhara, Yoshitomo; Ishii, Takeshi; Nakayama, Tsutomu; Osakabe, Naomi

    2015-01-01

    Theaflavins are polyphenols found in black tea, whose physiological activities are not well understood. This study on mice evaluated the influence of a single oral administration of theaflavins on energy metabolism by monitoring the initial metabolic changess in skeletal muscle and brown adipose tissue (BAT). Oxygen consumption (VO2) and energy expenditure (EE) were increased significantly in mice treated with theaflavin rich fraction (TF) compared with the group administered vehicle alone. There was no difference in locomotor activity. Fasting mice were euthanized under anesthesia before and 2 and 5, 20-hr after treatment with TF or vehicle. The mRNA levels of uncoupling protein-1 (UCP-1) and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in BAT were increased significantly 2-hr after administration ofTF. The levels of UCP-3 and PGC-1α in the gastrocnemius muscle were increased significantly 2 and 5-hr after administration of TF. The concentration of phosphorylated AMP-activated protein kinase (AMPK) 1α was also increased significantly in the gastrocnemius 2 and 5-hr after treatment with TF. These results indicate that TF significantly enhances systemic energy expenditure, as evidenced by an increase in expression of metabolic genes. PMID:26375960

  18. A Single Oral Administration of Theaflavins Increases Energy Expenditure and the Expression of Metabolic Genes.

    PubMed

    Kudo, Naoto; Arai, Yasunori; Suhara, Yoshitomo; Ishii, Takeshi; Nakayama, Tsutomu; Osakabe, Naomi

    2015-01-01

    Theaflavins are polyphenols found in black tea, whose physiological activities are not well understood. This study on mice evaluated the influence of a single oral administration of theaflavins on energy metabolism by monitoring the initial metabolic changess in skeletal muscle and brown adipose tissue (BAT). Oxygen consumption (VO2) and energy expenditure (EE) were increased significantly in mice treated with theaflavin rich fraction (TF) compared with the group administered vehicle alone. There was no difference in locomotor activity. Fasting mice were euthanized under anesthesia before and 2 and 5, 20-hr after treatment with TF or vehicle. The mRNA levels of uncoupling protein-1 (UCP-1) and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in BAT were increased significantly 2-hr after administration ofTF. The levels of UCP-3 and PGC-1α in the gastrocnemius muscle were increased significantly 2 and 5-hr after administration of TF. The concentration of phosphorylated AMP-activated protein kinase (AMPK) 1α was also increased significantly in the gastrocnemius 2 and 5-hr after treatment with TF. These results indicate that TF significantly enhances systemic energy expenditure, as evidenced by an increase in expression of metabolic genes. PMID:26375960

  19. Imaging muscarinic cholinergic receptors using I-123-paraiododexetimide and emission computed tomography

    SciTech Connect

    Frost, J.J.; Wilson, A.A.; Dannals, R.F.; Ravert, H.T.; Smith, A.C.; Wagner, H.N. Jr.

    1985-05-01

    Dexetimide is a high affinity muscarinic cholinergic receptor (MR) antagonist which binds preferentially to MR in vivo. The corresponding optical isomer of dexetimide is levetimide (LEV) which binds to MR with a thousand-fold lower affinity. Paraiododexetimide (IDEX) has an affinity slightly less than dexetimide (KD = 2.7 nM). Following the administration of I-125 IDEX (10 ..mu..g/kg,IV) to mice high radioactivity concentrations in the striatum and cerebral cortex and a low radioactivity concentration in the cerebellum were observed; this distribution corresponds to the known distribution of MR in the brain. Following the injection of I-125-ILEV a low and uniform radioactivity concentration was observed throughout the brain. Preliminary imaging studies using an Anger camera and I-123-IDEX in a baboon showed high radioactivity in the brain and parotid gland. The radioactivity in the parotid gland diminished with time whereas the radioactivity in the brain increased over the 90 min. imaging period, which suggests high affinity binding to MR in the brain. These results demonstrate the feasibility of imaging MR using single photon emission computed tomography and I-123-IDEX.

  20. Acute and subchronic administration of anandamide or oleamide increases REM sleep in rats.

    PubMed

    Herrera-Solís, Andrea; Vásquez, Khalil Guzmán; Prospéro-García, Oscar

    2010-03-01

    Anandamide and oleamide, induce sleep when administered acutely, via the CB1 receptor. Their subchronic administration must be tested to demonstrate the absence of tolerance to this effect, and that the sudden withdrawal of these endocannabinoids (eCBs) does not affect sleep negatively. The sleep-waking cycle of rats was evaluated for 24h, under the effect of an acute or subchronic administration of eCBs, and during sudden eCBs withdrawal. AM251, a CB1 receptor antagonist (CB1Ra) was utilized to block eCBs effects. Our results indicated that both acute and subchronic administration of eCBs increase REMS. During eCBs withdrawal, rats lack the expression of an abstinence-like syndrome. AM251 was efficacious to prevent REMS increase caused by both acute and subchronic administration of these eCBs, suggesting that this effect is mediated by the CB1 receptor. Our data further support a role of the eCBs in REMS regulation.

  1. Exogenous t-PA Administration Increases Hippocampal Mature BDNF Levels. Plasmin- or NMDA-Dependent Mechanism?

    PubMed Central

    Rodier, Marion; Prigent-Tessier, Anne; Béjot, Yannick; Jacquin, Agnès; Mossiat, Claude; Marie, Christine; Garnier, Philippe

    2014-01-01

    Brain-derived neurotrophic factor (BDNF) through TrkB activation is central for brain functioning. Since the demonstration that plasmin is able to process pro-BDNF to mature BDNF and that these two forms have opposite effects on neuronal survival and plasticity, a particular attention has been paid to the link between tissue plasminogen activator (tPA)/plasmin system and BDNF metabolism. However, t-PA via its action on different N-methyl-D-aspartate (NMDA) receptor subunits is also considered as a neuromodulator of glutamatergic transmission. In this context, the aim of our study was to investigate the effect of recombinant (r)t-PA administration on brain BDNF metabolism in rats. In the hippocampus, we found that rt-PA (10 mg/kg) administration induced a progressive increase in mature BDNF levels associated with TrkB activation. In order to delineate the mechanistic involved, plasmin activity was assessed and its inhibition was attempted using tranexamic acid (30 or 300 mg/kg, i.v.) while NMDA receptors were antagonized with MK801 (0.3 or 3 mg/kg, i.p.) in combination with rt-PA treatment. Our results showed that despite a rise in rt-PA activity, rt-PA administration failed to increase hippocampal plasmin activity suggesting that the plasminogen/plasmin system is not involved whereas MK801 abrogated the augmentation in mature BDNF levels observed after rt-PA administration. All together, our results show that rt-PA administration induces increase in hippocampal mature BDNF expression and suggests that rt-PA contributes to the control of brain BDNF synthesis through a plasmin-independent potentiation of NMDA receptors signaling. PMID:24670989

  2. Exogenous t-PA administration increases hippocampal mature BDNF levels. plasmin- or NMDA-dependent mechanism?

    PubMed

    Rodier, Marion; Prigent-Tessier, Anne; Béjot, Yannick; Jacquin, Agnès; Mossiat, Claude; Marie, Christine; Garnier, Philippe

    2014-01-01

    Brain-derived neurotrophic factor (BDNF) through TrkB activation is central for brain functioning. Since the demonstration that plasmin is able to process pro-BDNF to mature BDNF and that these two forms have opposite effects on neuronal survival and plasticity, a particular attention has been paid to the link between tissue plasminogen activator (tPA)/plasmin system and BDNF metabolism. However, t-PA via its action on different N-methyl-D-aspartate (NMDA) receptor subunits is also considered as a neuromodulator of glutamatergic transmission. In this context, the aim of our study was to investigate the effect of recombinant (r)t-PA administration on brain BDNF metabolism in rats. In the hippocampus, we found that rt-PA (10 mg/kg) administration induced a progressive increase in mature BDNF levels associated with TrkB activation. In order to delineate the mechanistic involved, plasmin activity was assessed and its inhibition was attempted using tranexamic acid (30 or 300 mg/kg, i.v.) while NMDA receptors were antagonized with MK801 (0.3 or 3 mg/kg, i.p.) in combination with rt-PA treatment. Our results showed that despite a rise in rt-PA activity, rt-PA administration failed to increase hippocampal plasmin activity suggesting that the plasminogen/plasmin system is not involved whereas MK801 abrogated the augmentation in mature BDNF levels observed after rt-PA administration. All together, our results show that rt-PA administration induces increase in hippocampal mature BDNF expression and suggests that rt-PA contributes to the control of brain BDNF synthesis through a plasmin-independent potentiation of NMDA receptors signaling.

  3. Laminaria japonica increases plasma exposure of glycyrrhetinic acid following oral administration of Liquorice extract in rats.

    PubMed

    Zhao, Wei-Man; Jiang, Shu-Wen; Chen, Yang; Zhong, Ze-Yu; Wang, Zhong-Jian; Zhang, Mian; Li, Ying; Xu, Ping; Liu, Li; Liu, Xiao-Dong

    2015-07-01

    The present study was designed to investigate the effects of Laminaria japonica (Laminaria) on pharmacokinetics of glycyrrhetinic acid (GA) following oral administration of Liquorice extract in rats. Following oral administrations of single-dose and multi-dose Liquorice extract and Liquorice-Laminaria extract, respectively, plasma samples were obtained at various times and the concentrations of GA, liquiritigenin, and isoliquiritigenin were measured by LC-MS. The effects of Laminaria extract on pharmacokinetics of GA were also investigated, following single-dose and multidose of glycyrrhizic acid (GL). The effects of Laminaria extract on intestinal absorption of GA and GL were studied using the in situ single-pass intestinal perfusion model. The metabolism of GL to GA in the contents of small and large intestines was also studied. The results showed Liquorice-Laminaria extract markedly increased the plasma concentration of GA, accompanied by a shorter Tmax. Similar alteration was observed following multidose administration. However, pharmacokinetics of neither liquiritigenin nor isoliquiritigenin was affected by Laminaria. Similarly, Laminaria markedly increased concentration and decreased Tmax of GA following oral GL were observed. The data from the intestinal perfusion model showed that Laminaria markedly increased GL absorption in duodenum and jejunum, but did not affect the intestinal absorption of GA. It was found that Laminaria enhanced the metabolism of GL to GA in large intestine. In conclusion, Laminaria increased plasma exposures of GA following oral administration of liquorice or GL, which partly resulted from increased intestinal absorption of GL and metabolism of GL to GA in large intestine.

  4. Muscarinic cholinergic and alpha/sub 1/ adrenergic receptors in murine atria: phosphatidylinositol breakdown and receptor interaction

    SciTech Connect

    Scherer, R.W.

    1987-01-01

    Upon stimulation of muscarinic cholinergic receptors, there is a decrease in the force of contraction rate of firing in heart, while stimulation of ..cap alpha.. adrenergic receptors causes an increase in the force of contraction with no change in the heart rate. Yet both receptors stimulate the breakdown of phosphatidylinositol (PI). Therefore, the breakdown of PI was examined to determine how the process differed between the two receptor systems. Murine atria, prelabelled with (/sup 3/H)inositol, were stimulated with the muscarinic cholinergic agonists, carbamylcholine (CARB), and oxotremorine (OXO); and with the ..cap alpha.. adrenergic agonists, norepinephrine (NE) and phenylephrine (PE); either singly or in combination. Breakdown of PI was assessed by measurement of individual inositol phosphates by anion exchange chromatography. Binding of CARB to atrial muscarinic receptors was measured by competition with (/sup 3/H)quinuclidinyl benzilate.

  5. Transient increase in alcohol self-administration following a period of chronic exposure to corticosterone.

    PubMed

    Besheer, Joyce; Fisher, Kristen R; Lindsay, Tessa G; Cannady, Reginald

    2013-09-01

    Stressful life events and chronic stressors have been associated with escalations in alcohol drinking. Stress exposure leads to the secretion of glucocorticoids (cortisol in the human; corticosterone (CORT) in the rodent). To model a period of heightened elevations in CORT, the present work assessed the effects of chronic exposure to the stress hormone CORT on alcohol self-administration. Male Long Evans rats were trained to self-administer a sweetened alcohol solution (2% sucrose/15% alcohol) resulting in moderate levels of daily alcohol intake (0.5-0.7 g/kg). Following stable baseline operant self-administration, rats received CORT in the drinking water for 7 days. A transient increase in alcohol self-administration was observed on the first self-administration session following CORT exposure, and behavior returned to control levels by the second session. Control experiments determined that this increase in alcohol self-administration was specific to alcohol, unrelated to general motor activation, and functionally dissociated from decreased CORT levels at the time of testing. These results indicate that repeated exposure to heightened levels of stress hormone (e.g., as may be experienced during stressful episodes) has the potential to lead to exacerbated alcohol intake in low to moderate drinkers. Given that maladaptive drinking patterns, such as escalated alcohol drinking following stressful episodes, have the potential to put an individual at risk for future drinking disorders, utilization of this model will be important for examination of neuroadaptations that occur as a consequence of CORT exposure in order to better understand escalated drinking following stressful episodes in nondependent individuals. PMID:23643750

  6. Oral administration of antimicrobials increase antimicrobial resistance in E. coli from chicken--a systematic review.

    PubMed

    Simoneit, C; Burow, E; Tenhagen, B-A; Käsbohrer, A

    2015-01-01

    Antimicrobials play an important role in animal and human health care. It was the aim of this systematic review to assess the effects of oral administration of antimicrobials on the development of antimicrobial resistance (AMR) in Escherichia coli (E. coli) from chickens. Moreover, the effects of the administration of more than one antimicrobial and of different dosages were studied. Literature was searched in November 2012 from the electronic databases ISI Web of Science, PubMed, Scopus and a national literature database (DIMDI) as well as the database ProQuest LLC. The search was updated in March 2014. Original studies describing a treatment (A) and a control group of either non-treatment (C) or initial value (0) and determining AMR in E. coli at different sample points (SP) were included. The literature search resulted in 35 full text articles on the topic, seven (20%) of which contained sufficient information on the administered antimicrobial and the impact of treatment on AMR. Most papers described the use of more than one antimicrobial, several dosages, controls (non-treatment or pre-treatment) and measured AMR at different SPs leading to a total of 227 SPs on the impact of the use of antimicrobials on AMR in chickens. 74% of the SPs (168/227) described a higher AMR-rate in E. coli from treated animals than from controls. After the administration of a single antimicrobial, AMR increased at 72% of the SPs. Administration of more than one antimicrobial increased AMR at 82% of the SPs. Higher dosages were associated with similar or higher AMR rates. The limited number of studies for each antimicrobial agent and the high variability in the resistance effect call for more well designed studies on the impact of oral administration on AMR development and spread.

  7. Increased loss and decreased synthesis of hepatic glutathione after acute ethanol administration. Turnover studies.

    PubMed Central

    Speisky, H; MacDonald, A; Giles, G; Orrego, H; Israel, Y

    1985-01-01

    The effect of acute ethanol administration on rates of synthesis and utilization of hepatic glutathione (GSH) was studied in rats after a pulse of [35S]cysteine. A 35% decrease in hepatic GSH content 5h after administration of 4 g of ethanol/kg body wt. was accompanied by a 33% increase in the rate of GSH utilization. The decrease occurred without increases in hepatic oxidized glutathione (GSSG) or in the GSH/GSSG ratio. The rate of non-enzymic condensation of GSH with acetaldehyde could account for only 6% of the rate of hepatic GSH disappearance. The increased loss of [35S]GSH induced by ethanol was not accompanied by an increased turnover; rather, a 30% inhibition of GSH synthesis balanced the increased rate of loss, leaving the turnover rate unchanged. The rate of acetaldehyde condensation with cysteine in vitro occurred at about one-third of the rate of GSH loss in ethanol-treated animals. However, ethanol induced only a minor decrease in liver cysteine content, which did not precede, but followed, the decrease in GSH. The characteristics of 2-methylthiazolidine-4-carboxylic acid, the condensation product between acetaldehyde and cysteine, were studied and methodologies were developed to determine its presence in tissues. It was not found in the liver of ethanol-treated animals. Ethanol administration led to a marked increase (47%) in plasma GSH in the post-hepatic inferior vena cava, but not in its pre-hepatic segment. Data suggest that an increased loss of GSH from the liver constitutes an important mechanism for the decrease in GSH induced by ethanol. In addition, an inhibition of GSH synthesis is observed. PMID:3977847

  8. Prophylactic Chronic Zinc Administration Increases Neuroinflammation in a Hypoxia-Ischemia Model.

    PubMed

    Tomas-Sanchez, Constantino; Blanco-Alvarez, Victor Manuel; Gonzalez-Barrios, Juan Antonio; Martinez-Fong, Daniel; Garcia-Robles, Guadalupe; Soto-Rodriguez, Guadalupe; Brambila, Eduardo; Torres-Soto, Maricela; Gonzalez-Vazquez, Alejandro; Aguilar-Peralta, Ana Karina; Garate-Morales, José-Luis; Aguilar-Carrasco, Luis-Angel; Limón, Daniel I; Cebada, Jorge; Leon-Chavez, Bertha Alicia

    2016-01-01

    Acute and subacute administration of zinc exert neuroprotective effects in hypoxia-ischemia animal models; yet the effect of chronic administration of zinc still remains unknown. We addressed this issue by injecting zinc at a tolerable dose (0.5 mg/kg weight, i.p.) for 14 days before common carotid artery occlusion (CCAO) in a rat. After CCAO, the level of zinc was measured by atomic absorption spectrophotometry, nitrites were determined by Griess method, lipoperoxidation was measured by Gerard-Monnier assay, and mRNA expression of 84 genes coding for cytokines, chemokines, and their receptors was measured by qRT-PCR, whereas nitrotyrosine, chemokines, and their receptors were assessed by ELISA and histopathological changes in the temporoparietal cortex-hippocampus at different time points. Long-term memory was evaluated using Morris water maze. Following CCAO, a significant increase in nitrosative stress, inflammatory chemokines/receptors, and cell death was observed after 8 h, and a 2.5-fold increase in zinc levels was detected after 7 days. Although CXCL12 and FGF2 protein levels were significantly increased, the long-term memory was impaired 12 days after reperfusion in the Zn+CCAO group. Our data suggest that the chronic administration of zinc at tolerable doses causes nitrosative stress, toxic zinc accumulation, and neuroinflammation, which might account for the neuronal death and cerebral dysfunction after CCAO. PMID:27635404

  9. Oral Administration of Recombinant Lactococcus lactis Expressing the Cellulase Gene Increases Digestibility of Fiber in Geese.

    PubMed

    Zhou, Haizhu; Gao, Yunhang; Gao, Guang; Lou, Yujie

    2015-12-01

    Enhancing cellulose digestibility in animals is important for improving the utilization of forage, which can decrease the amount of food used in animal production. The aim of the present study was to achieve recombinant expression of the cellulase gene in Lactococcus lactis and evaluate the effects of oral administration of the recombinant L. lactis on fiber digestibility in geese. Cellulase (Cell) and green fluorescent protein (GFP) genes were cloned into a L. lactis expression vector (pNZ8149) to construct the recombinant expression plasmid (pNZ8149-GFP-Cell). Then, the recombinant expression plasmid was transformed into L. lactis (NZ3900) competent cells by electroporation to obtain recombinant L. lactis (pNZ8149-GFP-Cell/NZ3900) in which protein expression was induced by Nisin. Expression of GFP and Cell by the recombinant L. lactis was confirmed using SDS-PAGE, fluorescence detection, and Congo red assays. A feeding experiment showed that oral administration of pNZ8149-GFP-Cell/NZ3900 significantly increased the digestibility of dietary fiber in geese fed either a maize stalk diet or a rice chaff diet. Therefore, oral administration of recombinant L. lactis cells expressing the cellulase gene increases fiber digestibility in geese, offering a way to increase the utilization of dietary fiber in geese.

  10. Oral Administration of Recombinant Lactococcus lactis Expressing the Cellulase Gene Increases Digestibility of Fiber in Geese.

    PubMed

    Zhou, Haizhu; Gao, Yunhang; Gao, Guang; Lou, Yujie

    2015-12-01

    Enhancing cellulose digestibility in animals is important for improving the utilization of forage, which can decrease the amount of food used in animal production. The aim of the present study was to achieve recombinant expression of the cellulase gene in Lactococcus lactis and evaluate the effects of oral administration of the recombinant L. lactis on fiber digestibility in geese. Cellulase (Cell) and green fluorescent protein (GFP) genes were cloned into a L. lactis expression vector (pNZ8149) to construct the recombinant expression plasmid (pNZ8149-GFP-Cell). Then, the recombinant expression plasmid was transformed into L. lactis (NZ3900) competent cells by electroporation to obtain recombinant L. lactis (pNZ8149-GFP-Cell/NZ3900) in which protein expression was induced by Nisin. Expression of GFP and Cell by the recombinant L. lactis was confirmed using SDS-PAGE, fluorescence detection, and Congo red assays. A feeding experiment showed that oral administration of pNZ8149-GFP-Cell/NZ3900 significantly increased the digestibility of dietary fiber in geese fed either a maize stalk diet or a rice chaff diet. Therefore, oral administration of recombinant L. lactis cells expressing the cellulase gene increases fiber digestibility in geese, offering a way to increase the utilization of dietary fiber in geese. PMID:26341925

  11. Prophylactic Chronic Zinc Administration Increases Neuroinflammation in a Hypoxia-Ischemia Model

    PubMed Central

    Tomas-Sanchez, Constantino; Blanco-Alvarez, Victor Manuel; Gonzalez-Barrios, Juan Antonio; Martinez-Fong, Daniel; Garcia-Robles, Guadalupe; Soto-Rodriguez, Guadalupe; Torres-Soto, Maricela; Gonzalez-Vazquez, Alejandro; Aguilar-Peralta, Ana Karina; Garate-Morales, José-Luis; Aguilar-Carrasco, Luis-Angel; Limón, Daniel I.; Cebada, Jorge

    2016-01-01

    Acute and subacute administration of zinc exert neuroprotective effects in hypoxia-ischemia animal models; yet the effect of chronic administration of zinc still remains unknown. We addressed this issue by injecting zinc at a tolerable dose (0.5 mg/kg weight, i.p.) for 14 days before common carotid artery occlusion (CCAO) in a rat. After CCAO, the level of zinc was measured by atomic absorption spectrophotometry, nitrites were determined by Griess method, lipoperoxidation was measured by Gerard-Monnier assay, and mRNA expression of 84 genes coding for cytokines, chemokines, and their receptors was measured by qRT-PCR, whereas nitrotyrosine, chemokines, and their receptors were assessed by ELISA and histopathological changes in the temporoparietal cortex-hippocampus at different time points. Long-term memory was evaluated using Morris water maze. Following CCAO, a significant increase in nitrosative stress, inflammatory chemokines/receptors, and cell death was observed after 8 h, and a 2.5-fold increase in zinc levels was detected after 7 days. Although CXCL12 and FGF2 protein levels were significantly increased, the long-term memory was impaired 12 days after reperfusion in the Zn+CCAO group. Our data suggest that the chronic administration of zinc at tolerable doses causes nitrosative stress, toxic zinc accumulation, and neuroinflammation, which might account for the neuronal death and cerebral dysfunction after CCAO.

  12. Prophylactic Chronic Zinc Administration Increases Neuroinflammation in a Hypoxia-Ischemia Model

    PubMed Central

    Tomas-Sanchez, Constantino; Blanco-Alvarez, Victor Manuel; Gonzalez-Barrios, Juan Antonio; Martinez-Fong, Daniel; Garcia-Robles, Guadalupe; Soto-Rodriguez, Guadalupe; Torres-Soto, Maricela; Gonzalez-Vazquez, Alejandro; Aguilar-Peralta, Ana Karina; Garate-Morales, José-Luis; Aguilar-Carrasco, Luis-Angel; Limón, Daniel I.; Cebada, Jorge

    2016-01-01

    Acute and subacute administration of zinc exert neuroprotective effects in hypoxia-ischemia animal models; yet the effect of chronic administration of zinc still remains unknown. We addressed this issue by injecting zinc at a tolerable dose (0.5 mg/kg weight, i.p.) for 14 days before common carotid artery occlusion (CCAO) in a rat. After CCAO, the level of zinc was measured by atomic absorption spectrophotometry, nitrites were determined by Griess method, lipoperoxidation was measured by Gerard-Monnier assay, and mRNA expression of 84 genes coding for cytokines, chemokines, and their receptors was measured by qRT-PCR, whereas nitrotyrosine, chemokines, and their receptors were assessed by ELISA and histopathological changes in the temporoparietal cortex-hippocampus at different time points. Long-term memory was evaluated using Morris water maze. Following CCAO, a significant increase in nitrosative stress, inflammatory chemokines/receptors, and cell death was observed after 8 h, and a 2.5-fold increase in zinc levels was detected after 7 days. Although CXCL12 and FGF2 protein levels were significantly increased, the long-term memory was impaired 12 days after reperfusion in the Zn+CCAO group. Our data suggest that the chronic administration of zinc at tolerable doses causes nitrosative stress, toxic zinc accumulation, and neuroinflammation, which might account for the neuronal death and cerebral dysfunction after CCAO. PMID:27635404

  13. Social stress increases the acquisition of cocaine self-administration in male and female rats.

    PubMed

    Haney, M; Maccari, S; Le Moal, M; Simon, H; Piazza, P V

    1995-11-01

    The effect of social stress on the vulnerability to commence cocaine self-administration was examined in Sprague-Dawley rats repeatedly exposed to aggressive attack by a same-sex opponent. Both sexes were studied, since the factors influencing the acquisition of drug self-administration in females have not been defined. Male and female rats encountered an aggressive male or lactating female opponent on four separate occasions over the course of one week. Control male and female rats were not exposed to attack. All animals were implanted with jugular catheters, and six days later placed into the self-administration box, where a nose-poke in the designated 'active hole' resulted in a 20 microliters injection of cocaine (0.32 mg/kg). Nose-pokes in an 'inactive' hole had no effect. Male and female rats that had experienced social stress self-administered more cocaine than non-defeated controls. The difference between the stressed and non-stressed animals in the number of cocaine injections was not present during the first few days of exposure to cocaine, but became more pronounced over time. Social stress increased the number of responses for cocaine, but did not alter the number of non-specific responses. Sex differences in self-administration were not significant. Therefore, social status appears to be a potent influence in the onset of drug taking behavior in both male and female rats. PMID:8581502

  14. Administration of Escherichia coli endotoxin to rat increases liver mass and hepatocyte volume in vivo.

    PubMed Central

    Qian, D; Brosnan, J T

    1996-01-01

    We have established, in vivo, an increase in liver mass and hepatocyte volume after a single intraperitoneal administration, to fasted rats, of Escherichia coli lipopolysaccharide (0127:B8) at 3 mg/kg. The phenomenon was time- and dose-dependent and could be prevented by treatment with polyclonal antiserum against tumour necrosis factor-alpha (TNF-alpha) before the endotoxin injection. Endotoxin caused an increase of 26% in the hepatic mass compared with fasted controls at 24 h. An increase of 27% in the hepatic water content underlay the altered hepatic mass which could not be accounted for by a change in the volume of hepatic blood and/or interstitial fluid (measured in vivo), suggesting an expansion in the hepatocellular volume. This is supported by an increase of 25% in the K+ content of the endotoxic livers. Morphometric study confirmed a 15% increase in hepatocyte volume after endotoxin administration. The data are discussed in the light of possible metabolic effects of increased hepatocyte volume. PMID:8573081

  15. Effect of the combination of the benzodiazepine tranquilizer medazepam and the nootropic agent meclofenoxate on the activity of rat brain muscarinic receptors.

    PubMed

    Popova, J S; Petkov, V D

    1990-01-01

    1. The effect of 7-day treatment with the benzodiazepine tranquilizer medazepam (5 mg/kg), the nootropic agent meclofenoxate (100 mg/kg) and their combination in the same doses on the binding activity of muscarinic receptors in four rat brain structures (cerebral cortex, striatum, hippocampus and hypothalamus) were studied using the antagonist [3H]-1-quinuclidinyl benzylate [( 3H]-QNB) as radio-ligand. 2. Medazepam treatment caused significant decrease of muscarinic receptor binding affinity (Kd) and of the receptor binding capacity (Bmax) in the brain structures studied. The number of muscarinic binding sites was unsignificantly decreased only in the hippocampus. 3. Meclofenoxate treatment caused an increase of muscarinic receptor affinity and a decrease of the binding capacity in the cerebral cortex and hypothalamus and an increase of the binding affinity in the striatum and hippocampus. 4. The combination of medazepam and meclofenoxate caused no significant changes of both muscarinic receptor characteristics in the hippocampus and of the receptor affinity in the striatum and hypothalamus in comparison with control rats. The Bmax values were decreased in the cerebral cortex, striatum and hypothalamus when compared with control animals. The differences observed were slighter than those determined after the comparison of medazepam treated rats with control rats. 5. The results obtained afford an opportunity to suggest that the nootropic agent meclofenoxate acts to moderate the effect of the benzodiazepine tranquilizer medazepam on the activity of rat brain muscarinic receptors.

  16. [Interaction of chagasic autoantibodies with the third extracellular domain of the human heart muscarinic receptor. Functional and pathological implications].

    PubMed

    Goin, J C; Pérez Leirós, C; Borda, E; Sterin-Borda, L

    1996-01-01

    Herein we demonstrate by ELISA and immunoblotting the presence in the sera of chagasic patients of circulating autoantibodies against the third extracellular domain of human muscarinic acetylcholine receptors by using a synthetic peptide corresponding to the sequence 169-192 of the receptor. Immunoaffinity purified antipeptide antibodies displayed cardiac muscarinic activity as decreased contractility and cAMP production and increased cGMP levels. These effects were specifically blocked by the synthetic peptide and by atropine. A strong association between the existence of circulating autoantibodies and the presence of dysautonomia was shown, making these autoantibodies an appropriate marker of heart autonomic dysfunction.

  17. Effects of muscarine on single rat adrenal chromaffin cells.

    PubMed Central

    Neely, A; Lingle, C J

    1992-01-01

    1. The action of muscarine on membrane currents and cytosolic calcium (Ca2+) of dissociated rat adrenal chromaffin cells was investigated using standard whole-cell voltage-clamp techniques and microfluorimetry of unclamped single cells. 2. In cells held at a constant holding potential negative to -40 mV, brief (5-10 s) applications of muscarine produced a transient activation of outward current. The activation of this current by muscarine also occurs in the presence of 5 mM-Co2+. 3. The outward current activated by muscarine at holding potentials negative to about -40 mV is blocked over 90% by either 200 microM-curare or 200 nM-apamin. One millimolar TEA produces variable blocking effects at such potentials. 4. The outward current activated by muscarine is transient even in the continuing presence of muscarine. Complete recovery between pairs of muscarine applications occurs over a 1-2 min period. If sufficient time was allowed for recovery between muscarine applications, the muscarine-activated outward current could be reliably elicited in dialysed cells for periods of 20-30 min. 5. Voltage ramps were used to examine effects of muscarine on currents over a range of membrane potentials. Over all potentials, muscarine activates a relatively voltage-independent component which is blocked almost completely by 200 nM-apamin and by 200 microM-curare. At potentials negative to about -40 mV, the apamin- and curare-sensitive current accounts for virtually all muscarine-activated current. This current appears to correspond to a Ca(2+)-activated, voltage-independent current found in these cells. Effects of muscarine on currents activated at potentials positive to 0 mV are complex. At potentials above 0 mV, muscarine can produce either an activation or an inhibition of outward current. The outward current activated at positive potentials was primarily voltage dependent and blocked by 1 mM-TEA. However, in some cells activation of voltage-dependent current was not observed and

  18. Enhanced AMPA receptor activity increases operant alcohol self-administration and cue-induced reinstatement.

    PubMed

    Cannady, Reginald; Fisher, Kristen R; Durant, Brandon; Besheer, Joyce; Hodge, Clyde W

    2013-01-01

    Long-term alcohol exposure produces neuroadaptations that contribute to the progression of alcohol abuse disorders. Chronic alcohol consumption results in strengthened excitatory neurotransmission and increased α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors (AMPA) receptor signaling in animal models. However, the mechanistic role of enhanced AMPA receptor activity in alcohol-reinforcement and alcohol-seeking behavior remains unclear. This study examined the role of enhanced AMPA receptor function using the selective positive allosteric modulator, aniracetam, in modulating operant alcohol self-administration and cue-induced reinstatement. Male alcohol-preferring (P-) rats, trained to self-administer alcohol (15%, v/v) versus water were pre-treated with aniracetam to assess effects on maintenance of alcohol self-administration. To determine reinforcer specificity, P-rats were trained to self-administer sucrose (0.8%, w/v) versus water, and effects of aniracetam were tested. The role of aniracetam in modulating relapse of alcohol-seeking was assessed using a response contingent cue-induced reinstatement procedure in P-rats trained to self-administer 15% alcohol. Aniracetam pre-treatment significantly increased alcohol-reinforced responses relative to vehicle treatment. This increase was not attributed to aniracetam-induced hyperactivity as aniracetam pre-treatment did not alter locomotor activity. AMPA receptor involvement was confirmed because 6,7-dinitroquinoxaline-2,3-dione (AMPA receptor antagonist) blocked the aniracetam-induced increase in alcohol self-administration. Aniracetam did not alter sucrose-reinforced responses in sucrose-trained P-rats, suggesting that enhanced AMPA receptor activity is selective in modulating the reinforcing function of alcohol. Finally, aniracetam pre-treatment potentiated cue-induced reinstatement of alcohol-seeking behavior versus vehicle-treated P-rats. These data suggest that enhanced glutamate activity at AMPA

  19. Agonist mediated conformational changes of solubilized calf forebrain muscarinic acetylcholine receptors.

    PubMed

    Vanderheyden, P; Andre, C; de Backer, J P; Vauquelin, G

    1984-10-01

    Muscarinic receptors in calf forebrain membranes can be identified by the specific binding of the radiolabelled antagonist [3H]dexetimide. These receptors (2.8 pM/mg protein) comprise two non-interconvertible subpopulations with respectively high and low agonist affinity but with the same antagonist affinity. For all the agonists tested the low affinity sites represent 85 +/- 5% of the total receptor population. 0.5% Digitonin solubilized extracts contain 0.8 pM muscarinic receptor/mg protein. In contrast with the membranes, these extracts contain only sites with low agonist affinity. The alkylating reagent N-ethylmaleimide causes an increase of the acetylcholine affinity for the low affinity sites in membranes as well as for the solubilized sites. This effect is time dependent until a maximal 3-fold increase in affinity is attained. The rate of N-ethylmaleimide action is enhanced by the concomitant presence of agonists. In contrast, N-ethylmaleimide does not affect antagonist binding. This suggests that agonists mediate a conformational change of both the membrane bound low affinity muscarinic sites and of the solubilized sites, resulting in their increased susceptibility towards NEM alkylation. PMID:6487351

  20. Cocaine treatment causes early and long-lasting changes in muscarinic and dopaminergic receptors.

    PubMed

    Macêdo, D S; Correia, E E; Vasconcelos, S M M; Aguiar, L M V; Viana, G S B; Sousa, F C F

    2004-02-01

    1. The study of changes that persist after drug discontinuation could be fundamental to understand the mechanisms involved in craving and relapse. 2. In this work the changes occurring in muscarinic, D1- and D2-like receptors after 30 min (immediate), 1 day (early), 5 and 30 days (late) withdrawal periods were studied, in the striatum of rats treated once a day for 7 days with cocaine 20 and 30 mg/kg, i.p. 3. Binding assays were performed in 10% homogenates and ligands used were [3H]-N-methylscopolamine, [3H]-SCH 23390, and [3H]-spiroperidol for muscarinic (M1 + M2-like), D1-, and D2-like receptors, respectively. 4. Muscarinic receptors presented a downregulation at all doses and discontinuation times, while the dissociation constant (Kd) for this receptor decreased after 30 min, 5 and 30 days abstinence times. In relation to D1-like receptors we found an antagonistic effect with 100% increase in receptor number 30 min after the last cocaine injection, but after 1-day withdrawal a downregulation was observed with both doses that persisted up to 30 days, only with the higher dose. The dissociation constant value (Kd) for this receptor showed a decrease only with 5 and 30 days withdrawal. An increase occurred with D2-like receptors at all doses and withdrawal periods studied, while Kd increased in 30-min, 5, and 30 days withdrawal. 5. In this work we found that the subchronic cocaine treatment produces early and long-lasting modifications in cholinergic muscarinic as well in dopaminergic receptors that persist up to 30 days of cocaine withdrawal. PMID:15049517

  1. Persistent Sodium Current Drives Conditional Pacemaking in CA1 Pyramidal Neurons under Muscarinic Stimulation

    PubMed Central

    Yamada-Hanff, Jason

    2013-01-01

    Hippocampal CA1 pyramidal neurons are normally quiescent but can fire spontaneously when stimulated by muscarinic agonists. In brain slice recordings from mouse CA1 pyramidal neurons, we examined the ionic basis of this activity using interleaved current-clamp and voltage-clamp experiments. Both in control and after muscarinic stimulation, the steady-state current–voltage curve was dominated by inward TTX-sensitive persistent sodium current (INaP) that activated near −75 mV and increased steeply with depolarization. In control, total membrane current was net outward (hyperpolarizing) near −70 mV so that cells had a stable resting potential. Muscarinic stimulation activated a small nonselective cation current so that total membrane current near −70 mV shifted to become barely net inward (depolarizing). The small depolarization triggers regenerative activation of INaP, which then depolarizes the cell from −70 mV to spike threshold. We quantified the relative contributions of INaP, hyperpolarization-activated cation current (Ih), and calcium current to pacemaking by using the cell's own firing as a voltage command along with specific blockers. TTX-sensitive sodium current was substantial throughout the entire interspike interval, increasing as the membrane potential approached threshold, while both Ih and calcium current were minimal. Thus, spontaneous activity is driven primarily by activation of INaP in a positive feedback loop starting near −70 mV and providing increasing inward current to threshold. These results show that the pacemaking “engine” from INaP is an inherent property of CA1 pyramidal neurons that can be engaged or disengaged by small shifts in net membrane current near −70 mV, as by muscarinic stimulation. PMID:24048831

  2. Muscarinic cholinergic receptors in pancreatic acinar carcinoma of rat.

    PubMed

    Taton, G; Delhaye, M; Swillens, S; Morisset, J; Larose, L; Longnecker, D S; Poirier, G G

    1985-04-15

    The active enantiomer of tritiated quinuclidinyl benzilate (3H(-)QNB) was used as a ligand to evaluate the muscarinic receptors. The 3H(-)QNB binding characteristics of muscarinic cholinergic receptors obtained from normal and neoplastic tissues were studied to determine changes in receptor properties during neoplastic transformation. Saturable and stereospecific binding sites for 3H(-)QNB are present in homogenates of rat pancreatic adenocarcinoma. The proportions of high- and low-affinity agonist binding sites are similar for neoplastic and normal tissues. The density of muscarinic receptors is higher in neoplastic (200 femtomoles/mg protein) than in normal pancreatic homogenates (80 femtomoles/mg protein). The muscarinic binding sites of the neoplastic and fetal pancreas show similar KD values which are higher than those observed for normal pancreas. PMID:2580801

  3. Progesterone inhibits behavioral responses and estrogen increases corticosterone levels after acute cocaine administration.

    PubMed

    Niyomchai, Tipyamol; Russo, Scott J; Festa, Eugene D; Akhavan, Alaleh; Jenab, Shirzad; Quiñones-Jenab, Vanya

    2005-04-01

    Accumulating evidence suggests that estrogen and progesterone contribute to the sexually dimorphic behavioral response to cocaine. In this study, we tested the hypothesis that varying the level of estrogen or progesterone affects cocaine-induced locomotive behavior in female rats. Ovariectomized (OVX) rats received estrogen (0, 5, 10, 20, or 50 microg) 48 h or progesterone (0, 50, 100, 250, or 500 microg) 24 h before acute saline or cocaine (15 mg/kg) administration. Although estrogen did not affect cocaine-induced ambulatory and rearing behaviors, it affected stereotypic behaviors regardless of cocaine administration (animals receiving 50 microg had higher stereotypic counts than did the OVX group). In contrast, progesterone affected rearing activity dose-dependently: 50 and 500 microg of progesterone inhibited, whereas 100 microg and 250 microg stimulated, rearing in response to cocaine. That estrogen and progesterone did not affect overall baseline behavioral activity suggests their effects are mediated in part through interactions with cocaine. Progesterone administration did not affect corticosterone levels in saline- or cocaine-treated rats. Estrogen administration, however, affected levels of corticosterone both at baseline and after cocaine treatment. After accounting for baseline differences, we found that rats receiving 5 or 10 microg of estrogen and cocaine had higher percentage increases in serum corticosterone levels than did the control group that did not receive estrogen. On the basis of these observations, we suggest that progesterone fluctuations during the estrous cycle impact cocaine-induced behavioral responses, whereas estrogen may affect activity in the hypothalamic-pituitary-adrenal axis. Thus, dose-dependent effects of gonadal hormones may underlie some of the reported sex differences and reproductive cycle effects of cocaine.

  4. Heat increases 3,4-methylenedioxymethamphetamine self-administration and social effects in rats.

    PubMed

    Cornish, Jennifer L; Shahnawaz, Zeeshan; Thompson, Murray R; Wong, Suzanna; Morley, Kirsten C; Hunt, Glenn E; McGregor, Iain S

    2003-12-15

    3,4-Methylenedioxymethamphetamine (MDMA, "Ecstasy") is a drug frequently used under hot conditions in nightclubs. In rats tested in the social interaction paradigm, greater prosocial effects of MDMA (5.0 mg/kg) were seen at a hot temperature (30 degrees C) relative to normal laboratory temperature (21 degrees C). In the intravenous drug self-administration paradigm, hot temperature (30 degrees C) increased the number of MDMA infusions (0.1, 0.3 or 1.0 mg/kg/infusion) self-administered by rats. Hot temperatures thus appear to affect both the social and reinforcing effects of MDMA. PMID:14660042

  5. Role for M5 muscarinic acetylcholine receptors in cocaine addiction.

    PubMed

    Fink-Jensen, Anders; Fedorova, Irina; Wörtwein, Gitta; Woldbye, David P D; Rasmussen, Thøger; Thomsen, Morgane; Bolwig, Tom G; Knitowski, Karen M; McKinzie, David L; Yamada, Masahisa; Wess, Jürgen; Basile, Anthony

    2003-10-01

    Muscarinic cholinergic receptors of the M5 subtype are expressed by dopamine-containing neurons of the ventral tegmentum. These M5 receptors modulate the activity of midbrain dopaminergic neurons, which play an important role in mediating reinforcing properties of abused psychostimulants like cocaine. The potential role of M5 receptors in the reinforcing effects of cocaine was investigated using M5 receptor-deficient mice in a model of acute cocaine self-administration. The M5-deficient mice self-administered cocaine at a significantly lower rate than wild-type controls. In the conditioned place preference procedure, a classic test for evaluating the rewarding properties of drugs, M5-deficient mice spent significantly less time in the cocaine-paired compartment than control mice. Moreover, the severity of the cocaine withdrawal syndrome (withdrawal-associated anxiety measured in the elevated plus-maze) was significantly attenuated in mice lacking the M5 receptor. These results demonstrate that M5 receptors play an important role in mediating both cocaine-associated reinforcement and withdrawal.

  6. Type 3 muscarinic receptors contribute to intestinal mucosal homeostasis and clearance of nippostrongylus brasiliensis through induction of Th2 cytokines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Despite increased appreciation for the role of nicotinic receptors in the modulation of and response to inflammation, the contribution of muscarinic receptors to mucosal homeostasis, clearance of enteric pathogens, and modulation of immune cell function remains relatively undefined. Uninfected and N...

  7. Constitutive Knockout of Kalirin-7 Leads to Increased Rates of Cocaine Self-Administration

    PubMed Central

    Kiraly, Drew D.; Nemirovsky, Natali E.; LaRese, Taylor P.; Tomek, Seven E.; Yahn, Stephanie L.; Olive, M. Foster; Eipper, Betty A.

    2013-01-01

    Kalirin-7 (Kal7) is a Rho-guanine nucleotide exchange factor that is localized in neuronal postsynaptic densities. Kal7 interacts with the NR2B subunit of the NMDA receptor and regulates aspects of dendritic spine dynamics both in vitro and in vivo. Chronic treatment with cocaine increases dendritic spine density in the nucleus accumbens (NAc) of rodents and primates. Kal7 mRNA and protein are upregulated in the NAc following cocaine treatment, and the presence of Kal7 is necessary for the normal proliferation of dendritic spines following cocaine use. Mice that constitutively lack Kal7 [Kalirin-7 knockout mice (Kal7KO)] demonstrate increased locomotor sensitization to cocaine and a decreased place preference for cocaine. Here, using an intravenous cocaine self-administration paradigm, Kal7KO mice exhibit increased administration of cocaine at lower doses as compared with wild-type (Wt) mice. Analyses of mRNA transcript levels from the NAc of mice that self-administered saline or cocaine reveal that larger splice variants of the Kalrn gene are increased by cocaine more dramatically in Kal7KO mice than in Wt mice. Additionally, transcripts encoding the NR2B subunit of the NMDA receptor increased in Wt mice that self-administered cocaine but were unchanged in similarly experienced Kal7KO mice. These findings suggest that Kal7 participates in the reinforcing effects of cocaine, and that Kal7 and cocaine interact to alter the expression of genes related to critical glutamatergic signaling pathways in the NAc. PMID:23894151

  8. Hydralazine administration activates sympathetic preganglionic neurons whose activity mobilizes glucose and increases cardiovascular function.

    PubMed

    Parker, Lindsay M; Damanhuri, Hanafi A; Fletcher, Sophie P S; Goodchild, Ann K

    2015-04-16

    Hypotensive drugs have been used to identify central neurons that mediate compensatory baroreceptor reflex responses. Such drugs also increase blood glucose. Our aim was to identify the neurochemical phenotypes of sympathetic preganglionic neurons (SPN) and adrenal chromaffin cells activated following hydralazine (HDZ; 10mg/kg) administration in rats, and utilize this and SPN target organ destination to ascribe their function as cardiovascular or glucose regulating. Blood glucose was measured and adrenal chromaffin cell activation was assessed using c-Fos immunoreactivity (-ir) and phosphorylation of tyrosine hydroxylase, respectively. The activation and neurochemical phenotype of SPN innervating the adrenal glands and celiac ganglia were determined using the retrograde tracer cholera toxin B subunit, in combination with in situ hybridization and immunohistochemistry. Blood glucose was elevated at multiple time points following HDZ administration but little evidence of chromaffin cell activation was seen suggesting non-adrenal mechanisms contribute to the sustained hyperglycemia. 16±0.1% of T4-T11 SPN contained c-Fos and of these: 24.3±1.4% projected to adrenal glands and 29±5.5% projected to celiac ganglia with the rest innervating other targets. 62.8±1.4% of SPN innervating adrenal glands were activated and 29.9±3.3% expressed PPE mRNA whereas 53.2±8.6% of SPN innervating celiac ganglia were activated and 31.2±8.8% expressed PPE mRNA. CART-ir SPN innervating each target were also activated and did not co-express PPE mRNA. Neurochemical coding reveals that HDZ administration activates both PPE+SPN, whose activity increase glucose mobilization causing hyperglycemia, as well as CART+SPN whose activity drive vasomotor responses mediated by baroreceptor unloading to raise vascular tone and heart rate.

  9. Role of the increased noradrenergic neurotransmission in drug self-administration.

    PubMed

    Wee, Sunmee; Wang, Zhixia; He, Rong; Zhou, Jia; Kozikowski, Alan P; Woolverton, William L

    2006-04-28

    Psychostimulants increase extracellular monoamine concentrations in the CNS. While the contributions of dopamine (DA) and serotonin (5-HT) to the reinforcing effect of psychostimulants have been examined, less is known about the involvement of norepinephrine (NE). In the present study, cocaine, desipramine (DMI) and JZ-III-84 were made available to rhesus monkeys (n=4) responding under a progressive-ratio (PR) schedule. These compounds vary in their in vitro selectivities for blocking NE uptake relative to DA from high (DMI) to modest (JZ-III-84) to non-selective (cocaine). Additionally, cocaine mixed with DMI in mg/kg dose-ratios of 1:1 to 1:3 was made available for self-administration. NE uptake inhibition by the mixture of cocaine and DMI at a ratio of 1:3 was evaluated in an ex vivo uptake assay. Cocaine (0.01-0.1 mg/(kg injection)) and JZ-III-84 (0.001-0.1 mg/(kg injection)) functioned as positive reinforcers with sigmoidal or biphasic dose-response functions, whereas DMI failed to do so. The addition of DMI to cocaine did not systemically alter self-administration of cocaine. In the ex vivo uptake assay, the addition of DMI to cocaine significantly increased the NE uptake inhibition compared to cocaine. These results support the conclusion that CNS NE is not involved in the reinforcing mechanism of psychostimulants. PMID:16213110

  10. M3-subtype muscarinic receptor that controls intracellular calcium release and inositol phosphate accumulation in gastric parietal cells.

    PubMed

    Leonard, A; Cuq, P; Magous, R; Bali, J P

    1991-07-25

    The muscarinic receptor subtype which triggers acid secretion was investigated in isolated rabbit gastric parietal cells. Cytosolic free Ca2+ concentration ([Ca2+]i), measured with the fluorescent indicator FURA-2, increased rapidly after full agonist (carbachol) stimulation (6-8 sec), then returned to an intermediate sustained value. Other M2-agonists, oxotremorine and arecoline, produced a partial [Ca2+]i increase, whereas M1-agonists, pilocarpine and [4-m-chlorophenylcarbamoyloxyl]-2-butynyl-trimethylammonium, were without any significant effect. [Ca2+]i rise was inhibited by selective muscarinic antagonists: atropine greater than 4-diphenylacetoxy-N-methyl-piperidine methbromide greater than quinuclidinylbenzilate (QNB) greater than pirenzepine greater than 11-[[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5,11-dihydro-6H- pyrido[2,3-b][1,4]benzodiazepine-6-one, this sequence being characteristic of the involvement of an M3-subtype. This inhibition was shown to be stereoselective; dexetimide and (-)QNB were more potent than levetimide and (+)QNB. The IC50 values for inhibition of [Ca2+]i increase by muscarinic antagonists were in good agreement with those obtained for inhibition of phospholipase C activation. In conclusion, the muscarinic receptor that controls acid secretion appears to be of the M3-subtype and the biochemical events coupled to the activation of this receptor system are also controlled through the same subtype. PMID:1651079

  11. Modulation of muscarinic and micotinic cholinergic receptor mediated catecholamine secretion in guinea pig chromaffin cells by phorbol esters

    SciTech Connect

    Figueiredo, J.C.; Fisher, S.K.; Horowitz, M.I.

    1986-05-01

    Isolated guinea pig chromaffin cells possess both nicotinic (nAChR) and muscarinic (mAChR) cholinergic receptors that are positively coupled to catecholamine (CA) release. Sixty to 70% of CA release is mediated by nAChRs and 30-40% by mAChRs. In the absence of added calcium, nAChR mediated CA release was reduced by 65% whereas the muscarinic response was unaffected. The addition of 100nM 12-0-tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C (PKC), also resulted in an increased CA release. Temporally and quantitatively, this response resembled that of mAChR activation. Addition of optimal concentrations of nicotine (50..mu..M) and TPA (100nM) induced a synergistic increase in CA release. Addition of muscarine (1mM) and TPA resulted in an additive response despite a 40-60% inhibition of mAChR mediated inositol phosphate release by TPA. Thus, in guinea pig chromaffin cells, it appears that PKC activation alone is a sufficient stimulus for CA release and that activation of both nicotinic and muscarinic receptors may further increase this enzyme's activity.

  12. Muscarinic receptor subtypes controlling the cationic current in guinea-pig ileal smooth muscle

    PubMed Central

    Zholos, Alexander V; Bolton, Thomas B

    1997-01-01

    The effects of muscarinic antagonists on cationic current evoked by activating muscarinic receptors with the stable agonist carbachol were studied by use of patch-clamp recording techniques in guinea-pig single ileal smooth muscle cells. Ascending concentrations of carbachol (3–300 μM) activated the cationic conductance in a concentration-dependent manner with conductance at a maximally effective carbachol concentration (Gmax) of 27.4±1.4 nS and a mean −log EC50 of 5.12±0.03 (mean±s.e.mean) (n=114). Muscarinic antagonists with higher affinity for the M2 receptor, methoctramine, himbacine and tripitramine, produced a parallel shift of the carbachol concentration-effect curve to the right in a concentration-dependent manner with pA2 values of 8.1, 8.0 and 9.1, respectively. All M3 selective muscarinic antagonists tested, 4-DAMP, p-F-HHSiD and zamifenacin, reduced the maximal response in a concentration-dependent and non-competitive manner. This effect could be observed even at concentrations which did not produce any increase in the EC50 for carbachol. At higher concentrations M3 antagonists shifted the agonist curve to the right, increasing the EC50, and depressed the maximum conductance response. Atropine, a non-selective antagonist, produced both reduction in Gmax (M3 effect) and significant increase in the EC50 (M2 effect) in the same concentration range. The depression of the conductance by 4-DAMP, zamifenacin and atropine could not be explained by channel block as cationic current evoked by adding GTPγS to the pipette (without application of carbachol) was unaffected. The results support the hypothesis that carbachol activates M2 muscarinic receptors so initiating the opening of cationic channels which cause depolarization; this effect is potentiated by an unknown mechanism when carbachol activates M3 receptors. As an increasing fraction of M3 receptors are blocked by an antagonist, the effects on cationic current of an increasing proportion of

  13. Enhanced AMPA receptor activity increases operant alcohol self-administration and cue-induced reinstatement.

    PubMed

    Cannady, Reginald; Fisher, Kristen R; Durant, Brandon; Besheer, Joyce; Hodge, Clyde W

    2013-01-01

    Long-term alcohol exposure produces neuroadaptations that contribute to the progression of alcohol abuse disorders. Chronic alcohol consumption results in strengthened excitatory neurotransmission and increased α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors (AMPA) receptor signaling in animal models. However, the mechanistic role of enhanced AMPA receptor activity in alcohol-reinforcement and alcohol-seeking behavior remains unclear. This study examined the role of enhanced AMPA receptor function using the selective positive allosteric modulator, aniracetam, in modulating operant alcohol self-administration and cue-induced reinstatement. Male alcohol-preferring (P-) rats, trained to self-administer alcohol (15%, v/v) versus water were pre-treated with aniracetam to assess effects on maintenance of alcohol self-administration. To determine reinforcer specificity, P-rats were trained to self-administer sucrose (0.8%, w/v) versus water, and effects of aniracetam were tested. The role of aniracetam in modulating relapse of alcohol-seeking was assessed using a response contingent cue-induced reinstatement procedure in P-rats trained to self-administer 15% alcohol. Aniracetam pre-treatment significantly increased alcohol-reinforced responses relative to vehicle treatment. This increase was not attributed to aniracetam-induced hyperactivity as aniracetam pre-treatment did not alter locomotor activity. AMPA receptor involvement was confirmed because 6,7-dinitroquinoxaline-2,3-dione (AMPA receptor antagonist) blocked the aniracetam-induced increase in alcohol self-administration. Aniracetam did not alter sucrose-reinforced responses in sucrose-trained P-rats, suggesting that enhanced AMPA receptor activity is selective in modulating the reinforcing function of alcohol. Finally, aniracetam pre-treatment potentiated cue-induced reinstatement of alcohol-seeking behavior versus vehicle-treated P-rats. These data suggest that enhanced glutamate activity at AMPA

  14. Airway and lung remodelling in chronic pulmonary obstructive disease: a role for muscarinic receptor antagonists?

    PubMed

    Roth, Michael

    2015-01-01

    Lung tissue remodelling in chronic inflammatory lung diseases has long been regarded as a follow-up event to inflammation. Recent studies have indicated that, although airway and lung tissue remodelling is often independent of inflammation, it precedes or causes inflammation. None of the available therapies has a significant effect on airway and lung tissue remodelling in asthma, bronchiectasis, fibrosis and chronic obstructive pulmonary disease (COPD). The goal of stopping or reversing lung tissue remodelling is difficult, as the term summarizes the net effect of independent events, including (1) cell proliferation, (2) cell volume increase, (3) cell migration, (4) modified deposition and metabolism of specific extracellular matrix components, and (5) local action of infiltrated inflammatory cells. The extracellular matrix of the lung has a very high turnover, and thus small changes may accumulate to significant structural pathologies, which seem to be irreversible. The most important question is 'why are pathological changes of the lung structure irreversible and resistant to drugs?' Many drugs have the potential to reduce remodelling mechanisms in vitro but fail in clinical trials. New evidence suggests that muscarinic receptor inhibitors have the potential to improve lung function through modifying tissue remodelling. However, the role of muscarinic receptors in lung remodelling, especially their supportive role for other remodelling driving factors, needs to be further investigated. The focus of this review is the role of muscarinic receptors in lung tissue remodelling as it has been reported in the human lung.

  15. Oral administration of lactoferrin increases hemoglobin and total serum iron in pregnant women.

    PubMed

    Paesano, Rosalba; Torcia, Francesco; Berlutti, Francesca; Pacifici, Enrica; Ebano, Valeria; Moscarini, Massimo; Valenti, Piera

    2006-06-01

    Iron deficiency anemia (IDA) during pregnancy continues to be of world-wide concern. IDA is a risk factor for preterm delivery and subsequent low birth weight, and possibly for poor neonatal health. Iron supplementation in pregnancy is a widely recommended practice, yet intervention programs have met with many controversies. In our study, 300 women at different trimesters of pregnancy were enrolled in a trial of oral administration of ferrous sulfate (520 mg once a day) or 30% iron-saturated bovine lactoferrin (bLf) (100 mg twice a day). Pregnant women refusing treatment represented the control group. In this group hemoglobin and total serum iron values measured after 30 d without treatment decreased significantly, especially in women at 18-31 weeks of pregnancy. In contrast, after 30 d of oral administration of bLf, hemoglobin and total serum iron values increased and to a greater extent than those observed in women treated orally for 30 d with ferrous sulfate, independently of the trimester of pregnancy. Unlike ferrous sulfate, bLf did not result in any side effects. These findings lead us to hypothesize that lactoferrin could influence iron homeostasis directly or through other proteins involved in iron transport out of the intestinal cells into the blood.

  16. Muscarinic receptor binding and muscarinic receptor-mediated inhibition of adenylate cyclase in rat brain myelin

    SciTech Connect

    Larocca, J.N.; Ledeen, R.W.; Dvorkin, B.; Makman, M.H.

    1987-12-01

    High-affinity muscarinic cholinergic receptors were detected in myelin purified from rat brain stem with use of the radioligands /sup 3/H-N-methylscopolamine (/sup 3/H-NMS), /sup 3/H-quinuclidinyl benzilate (/sup 3/H-QNB), and /sup 3/H-pirenzepine. /sup 3/H-NMS binding was also present in myelin isolated from corpus callosum. In contrast, several other receptor types, including alpha 1- and alpha 2-adrenergic receptors, present in the starting brain stem, were not detected in myelin. Based on Bmax values from Scatchard analyses, /sup 3/H-pirenzepine, a putative M1 selective ligand, bound to about 25% of the sites in myelin labeled by /sup 3/H-NMS, a nonselective ligand that binds to both M1 and M2 receptor subtypes. Agonist affinity for /sup 3/H-NMS binding sites in myelin was markedly decreased by Gpp(NH)p, indicating that a major portion of these receptors may be linked to a second messenger system via a guanine-nucleotide regulatory protein. Purified myelin also contained adenylate cyclase activity; this activity was stimulated several fold by forskolin and to small but significant extents by prostaglandin E1 and the beta-adrenergic agonist isoproterenol. Myelin adenylate cyclase activity was inhibited by carbachol and other muscarinic agonists; this inhibition was blocked by the antagonist atropine. Levels in myelin of muscarinic receptors were 20-25% and those of forskolin-stimulated adenylate cyclase 10% of the values for total particulate fraction of whole brain stem. These levels in myelin are appreciably greater than would be predicted on the basis of contamination. Also, additional receptors and adenylate cyclase, added by mixing nonmyelin tissue with whole brain stem, were quantitatively removed during the purification procedure.

  17. Heterogeneity of muscarinic receptors in lamb isolated coronary resistance arteries.

    PubMed Central

    Simonsen, U.; Prieto, D.; Rivera, L.; Hernández, M.; Mulvany, M. J.; García-Sacristán, A.

    1993-01-01

    1. In vitro experiments in a microvascular myograph were designed to characterize postjunctional muscarinic receptors producing contraction both in the presence and absence of the endothelium in coronary resistance arteries (normalized diameter of 150-450 microns), isolated from the left ventricle of hearts from 3-6 month old lambs. Preferential muscarinic receptor antagonists were used to determine the receptor subtype: pirenzepine (M1 receptor), AFDX 116 (M2 receptor), 4-DAMP and pFHHSiD (M3 receptor). 2. The rank order of potency for muscarinic agonist-induced increases in tension in endothelium-intact preparations was oxotremorine-M = methacholine = acetylcholine (ACh) > carbachol. Removal of the endothelium increased the potency of ACh, but this procedure did not change either the sensitivity or maximal response to carbachol. 3. The contractile response to ACh was reproducible. Incubation with 3 x 10(-7)-3 x 10(-6) M pirenzepine induced non-parallel rightward shifts and depressed the maximum of the concentration-response curve to ACh in endothelium-intact arteries. The slope by Schild analysis was 2.9 +/- 0.8 (P < 0.05, n = 7). Atropine, AFDX 116, 4-DAMP and pFHHSiD produced parallel rightward shifts of the curves to ACh and the slopes of the Schild plots were not significantly different from unity. The pKB values for the antagonists from plots constrained to unity in endothelium-intact segments were: atropine (9.4), 4-DAMP (9.0), pFHHSiD (7.9) and AFDX 116 (6.2). 4. In endothelium-denuded arteries, pirenzepine, AFDX 116 and pFHHSiD caused concentration-dependent, parallel rightward displacements of the concentration-response curves to ACh and the slopes of the Schild plots were not significantly different from unity. The plots constrained to a slope of unity gave the following pKB values: pFHHSiD (8.7), pirenzepine (7.5) and AFDX 116 (6.2). 5. In the presence of the endothelium, low concentrations of pirenzepine (10(-9)-10(-7) M) produced leftward shifts of

  18. Prolonged withdrawal following cocaine self-administration increases resistance to punishment in a cocaine binge

    PubMed Central

    Gancarz-Kausch, Amy M.; Adank, Danielle N.; Dietz, David M.

    2014-01-01

    Drug addiction is characterized by compulsive drug-taking behaviors and a high propensity to relapse following drug cessation. Drug craving and seeking can increase during a period of abstinence, but this phenomenon is not observed in drug-induced reinstatement models. To investigate the effect of withdrawal on cocaine relapse, rats were exposed to extended-access cocaine self-administration and subjected to either 1 or 30 d of withdrawal. When tested during 12 h unlimited access to cocaine (binge), the duration of the withdrawal did not influence cocaine intake. However, using a histamine punishment procedure that greatly suppresses drug-taking behavior, we demonstrate that longer periods of abstinence from cocaine induce a greater persistence in responding for drug in the face of negative consequences. PMID:25363133

  19. NOAA Would Receive an 11% Increase Under Obama Administration's Proposed Budget

    NASA Astrophysics Data System (ADS)

    Showstack, Randy

    2013-05-01

    The White House's proposed fiscal year (FY) 2014 budget for the National Oceanic and Atmospheric Administration (NOAA) would provide the agency with 5.45 billion, 11% above the FY 2012 spend plan of 4.91 billion (see Table ). The proposal, which was sent to Congress on 10 April, would increase funding for operations, research, and facilities to 3.41 billion (up 7.97% over FY 2012) and for procurement, acquisition, and construction to 2.12 billion (up 17.51%). The budget proposal uses the FY 2012 spend plan as a comparison because Congress approved the FY 2013 appropriations only a few weeks before the FY 2014 proposal was released.

  20. Stress- and pharmacologically-induced behavioral sensitization increases vulnerability to acquisition of amphetamine self-administration.

    PubMed

    Piazza, P V; Deminiere, J M; le Moal, M; Simon, H

    1990-04-23

    Individual vulnerability to drug addiction may be an important factor in the prognosis of this pathological behavior in man. However, experimental investigations have largely neglected the psychobiological substrate of predisposition to addiction. In this study, we show using a self-administration (SA) acquisition paradigm that previous repeated exposure to a stressful experience (tail-pinch) or to amphetamine, increase the locomotor response to this drug (behavioral sensitization) and enhance vulnerability to acquire amphetamine SA. These results show that vulnerability to develop amphetamine SA may be influenced by stressful experiences, and that previous contact with the drug may enhance a predisposition to amphetamine-taking behavior. As tail-pinch and amphetamine sensitization affect both the dopamine (DA) neural system and the propensity to self-administer amphetamine (behavior also modulated by DA activity), stress may influence SA via an action on the DA system. PMID:2357527

  1. Prolonged withdrawal following cocaine self-administration increases resistance to punishment in a cocaine binge.

    PubMed

    Gancarz-Kausch, Amy M; Adank, Danielle N; Dietz, David M

    2014-11-03

    Drug addiction is characterized by compulsive drug-taking behaviors and a high propensity to relapse following drug cessation. Drug craving and seeking can increase during a period of abstinence, but this phenomenon is not observed in drug-induced reinstatement models. To investigate the effect of withdrawal on cocaine relapse, rats were exposed to extended-access cocaine self-administration and subjected to either 1 or 30 d of withdrawal. When tested during 12 h unlimited access to cocaine (binge), the duration of the withdrawal did not influence cocaine intake. However, using a histamine punishment procedure that greatly suppresses drug-taking behavior, we demonstrate that longer periods of abstinence from cocaine induce a greater persistence in responding for drug in the face of negative consequences.

  2. Increasing Access to Health Administrative Data with ICES Data & Analytic Services.

    PubMed

    Ishiguro, Lisa; Saskin, Refik; Vermeulen, Marian J; Yates, Erika; Gunraj, Nadia; Victor, J Charles

    2016-01-01

    The Institute for Clinical Evaluative Sciences (ICES) is one of only a few organizations in Ontario permitted to access, link and analyze health administrative data for the purpose of generating evidence to inform decisions in policy and practice. Although ICES is a leading research institute, its access to the data has historically been restricted to scientists with an ICES affiliation. This requirement, designed to meet ICES' data privacy and security obligations, created barriers with respect to the widespread use of Ontario's data assets. In 2014, as part of the government's commitment to the Strategy for Patient-Oriented Research, ICES launched the Data & Analytic Services platform, which is aimed at increasing access to data and analytic services to investigators external to ICES. In making the data widely available to the broader research community, this initiative engages investigators involved in front-line care, stimulates new avenues of research and fosters collaboration that was previously challenging or unfeasible. PMID:27133600

  3. Oscillatory chloride current evoked by temperature jumps during muscarinic and serotonergic activation in Xenopus oocyte.

    PubMed Central

    Miledi, R; Parker, I; Sumikawa, K

    1987-01-01

    1. Membrane currents were recorded from voltage-clamped oocytes of Xenopus laevis, during temperature jumps imposed by a heating light. Resting oocytes usually showed little response, but large oscillatory membrane currents developed in response to cooling steps applied during activation of 'native' muscarinic receptors. 2. Similar temperature jump (Tjump) currents were seen during activation of oscillatory chloride currents mediated by muscarinic acetylcholine (ACh), serotonin, glutamate and noradrenaline receptors, expressed in the oocyte following injection with messenger ribonucleic acid (mRNA) from rat brain. The Tjump response during muscarinic activation was selectively blocked by atropine, and that during serotonergic activation by methysergide. In contrast, the 'smooth' membrane currents elicited by nicotinic ACh, kainate and gamma-aminobutyric acid (GABA) were not accompanied by Tjump responses. 3. Rapid cooling of the oocyte gave larger Tjump currents than a gradual cooling over a few seconds. The size of the Tjump current elicited by a fixed cooling step increased linearly with the preceding time of warming, becoming maximal at intervals greater than about 100 s. 4. The Tjump current was inward at a clamp potential of -60 mV and reversed direction at about -22 mV, which corresponds to the chloride equilibrium potential in the oocyte. In low-chloride solution the reversal potential was shifted to more positive potentials, but it was almost unchanged by changes in potassium and sodium concentration. The size of the Tjump current decreased as the membrane potential was made more negative than about -40 mV. 5. The period of oscillation of the Tjump current increased with decreasing temperature, following a Q10 of 3.15. Depolarization also caused a small increase in period. 6. The Tjump current was not abolished in calcium-free solution, or by addition of manganese or lanthanum to the bathing solution. However, it was abolished by intracellular injection of

  4. Inflammation induced by increased frequency of intermittent hypoxia is attenuated by tempol administration.

    PubMed

    Zhang, J; Zheng, L; Cao, J; Chen, B; Jin, D

    2015-12-01

    The levels of serum inflammatory cytokines and the activation of nuclear factor kappa B (NF-κB) and hypoxia inducible factor-1α (HIF-1α) in heart tissues in response to different frequencies of intermittent hypoxia (IH) and the antioxidant tempol were evaluated. Wistar rats (64 males, 200-220 g) were randomly divided into 6 experimental groups and 2 control groups. Four groups were exposed to IH 10, 20, 30, or 40 times/h. The other 2 experimental groups were challenged with IH (30 times/h) plus tempol, either beginning on day 0 (IH30T0) or on day 29 (IH30T29). After 6 weeks of challenge, serum levels of tumor necrosis factor (TNF)-α, intracellular adhesion molecule (ICAM)-1, and interleukin-10 were measured, and western blot analysis was used to detect NF-κB p65 and HIF-1α in myocardial tissues. Serum levels of TNF-α and ICAM-1 and myocardial expression of NF-κB p65 and HIF-1α were all significantly higher in IH rats than in controls (P<0.001). Increased IH frequency resulted in more significant changes. Administration of tempol in IH rats significantly reduced levels of TNF-α, ICAM-1, NF-κB and HIF-1α compared with the non-tempol-treated group (F=16.936, P<0.001). IH induced an inflammatory response in a frequency-dependent manner. Additionally, HIF-1α and NF-κB were increased following IH administration. Importantly, tempol treatment attenuated this effect. PMID:26397969

  5. The cholinergic immune regulation mediated by a novel muscarinic acetylcholine receptor through TNF pathway in oyster Crassostrea gigas.

    PubMed

    Liu, Zhaoqun; Zhou, Zhi; Wang, Lingling; Dong, Wenjing; Qiu, Limei; Song, Linsheng

    2016-12-01

    Muscarinic receptors, which selectively take muscarine as their ligand, are critical for the immunological and physiological processes in animals. In the present study, the open region frame (ORF) of a homologue of muscarinic acetylcholine (ACh) receptor (mAChR) was amplified from oyster Crassostrea gigas (named as CgmAChR-1), whose full length was 1983 bp and the protein it encoded contained 660 amino acids with a seven transmembrane region. Phylogeny analysis suggested that CgmAChR-1 shared homology with M5 muscarinic receptor found in invertebrates including Habropoda laboriosa, Acromyrmex echinatior and Echinococcus granulosus. After cell transfection of CgmAChR-1 into HEK293T cells and ACh incubation, the level of intracellular Ca(2+) and cAMP increased significantly (p < 0.05). Such trend could be reverted with the addition of M3 and M5 muscarinic receptor antagonists DAMP and DAR. The CgmAChR-1 transcripts were ubiquitously detectable in seven different tissues with the maximal expression level in adductor muscle. When the oysters received LPS stimulation, CgmAChR-1 mRNA expression in haemocyte was increased to the highest level (6.05-fold, p < 0.05) at 24 h, while blocking CgmAChR-1 using receptor antagonists before LPS stimulation promoted the expression of oyster TNF, resulting in the increase of haemocyte apoptosis index. These results suggested that CgmAChR-1 was the key molecule in cholinergic neuroendocrine-immune system contributing to the regulation of TNF expression and apoptosis process. PMID:27394930

  6. Amnesic effects of the anticholinergic drugs, trihexyphenidyl and biperiden: differences in binding properties to the brain muscarinic receptor.

    PubMed

    Kimura, Y; Ohue, M; Kitaura, T; Kihira, K

    1999-07-10

    An amnesic effect of anticholinergic drugs was previously described from several behavioral studies. We examined this effect induced by trihexyphenidyl and biperiden, clinically used in the parkinsonism and schizophrenic patients, by using passive avoidance tasks. Both of these drugs (0.1-10 mg/kg, s.c.) showed dose-dependent amnesic effects in the acquisition and retrieval phases. However, the effect induced by trihexyphenidyl was transient, whereas that of biperiden was long-lasting. To clarify the reason for the different duration of the amnesic activity, binding to the muscarinic receptor was examined. In the Scatchard analysis, trihexyphenidyl competed with [(3)H]quinuclidinyl benzilate ([(3)H]QNB) on the muscarinic receptor (showed increased K(d) and unchanged B(max) value), while biperiden decreased [(3)H]QNB binding (B(max) value) significantly. Furthermore, in an exchange assay for receptor inactivation, trihexyphenidyl binding to muscarinic receptors was exchanged by [(3)H]QNB completely, but biperiden decreased the exchangeable binding of [(3)H]QNB in a dose dependent manner (0.1-100 nM). These results suggested that the binding of trihexyphenidyl and biperiden to muscarinic receptor might be completely reversible and partially irreversible, respectively, whereas the K(i) values of these two drugs were similar. In conclusion, this difference in binding property may explain the difference in the time-course of the amnesic effect induced by trihexyphenidyl and biperiden.

  7. General pharmacological profile of the novel muscarinic receptor agonist SNI-2011, a drug for xerostomia in Sjögren's syndrome. 2nd communication: effects on somatic nervous system and on autonomic nervous system and smooth muscle.

    PubMed

    Arisawa, Hirohiko; Fukui, Kenji; Fujise, Nobuaki; Masunaga, Hiroaki

    2002-01-01

    A novel muscarinic receptor agonist SNI-2011 ((+/-)-cis-2-methylspirol[1,3-oxathiolane-5,3'-quinuclidine] monohydrochloride hemihydrate, cevimeline, CAS 153504-70-2), is a candidate therapeutic drug for xerostomia in Sjögren's syndrome. The general pharmacological properties of this drug on the somatic nervous system and on the autonomic nervous system and smooth muscle were investigated in mice, rats, guinea pigs, rabbits and cats. 1. Somatic nervous system: SNI-2011 had no effect on the neuromuscular junction in rats and no muscle relaxant effect in mice. No surface anesthetic effect was observed in guinea pigs, but infiltration anesthetic effect was found after intracutaneous injection of solution (1% or higher). 2. Autonomic nervous system and smooth muscle: SNI-2011 tended to cause mydriasis at 3 mg/kg i.v. or higher in rabbits and dose-dependently caused mydriasis at 10 mg/kg p.o. or higher in rats. Mydriasis in rats was also observed by ophthalmic instillation, caused via the peripheral muscarinic acetylcholine receptors. SNI-2011 elevated the base line tension of nictitating membrane in cats when it was injected intravenously at 3 mg/kg or higher. In the smooth muscle, SNI-2011 increased the spontaneous movement of isolated rabbit ileum (1 x 10(-6) mol/l or higher), contractions of isolated guinea pig ileum (1 x 10(-6) mol/l or higher) and isolated guinea pig trachea (3 x 10(-6) mol/l or higher). SNI-2011 relaxed the histamine- and noradrenaline-induced contractions of isolated guinea pig aorta and augmented noradrenaline- and phenylephrine-induced contractions of isolated rat vas deferens. These effects were induced by relatively higher concentrations only i.e. 1 x 10(-5) mol/l or higher. From these results, SNI-2011 has muscarinic side effects on the somatic nervous system and on the autonomic nervous system and smooth muscle, however, in the case of oral administration, that is clinical administration route, SNI-2011 caused no muscarinic side effect at

  8. Circadian activity of corticosterone in an animal model of depression: response to muscarinic cholinergic stimulation.

    PubMed

    Bonilla-Jaime, H; Retana-Márquez, S; Arteaga-Silva, M; Hernández-González, M; Vázquez-Palacios, G

    2010-06-16

    Neonatal treatment with clomipramine (CMI) in rats induces multiple behavioral alterations during adulthood that resemble certain symptoms of human depression, such as impairments of pleasure-seeking behaviors. CMI may also induce permanent changes in the reactivity of the hypothalamic-pituitary-adrenocortical axis (HPA) to different stimuli; however, the endocrinal changes induced by this treatment are still a matter of debate. In the present study, we evaluated the levels of corticosterone in rats treated in the neonatal period with CMI in basal conditions (0, 6, 12 and 18 h after lights on) and after treatment with the antidepressant fluoxetine (FLX; 5mg/kg for 14 days). To evaluate the response of the HPA axis to a cholinergic agonist, we analyzed the effect of oxotremorine administration (OXO; 0.4, 0.8 mg/kg) on plasma levels of corticosterone. Administration of OXO took place at the beginning of each one of the two phases of the light-dark cycle (time points 0 and 12h, respectively). Results showed an increase in basal plasma levels of corticosterone in CMI-treated rats at time point zero and at 6h after the onset of the light period. While treatment with FLX reversed the increase in corticosterone plasma levels in CMI-treated rats, the results regarding cholinergic stimulation indicate that those rats do not respond to the administration of a low dose of OXO (0.4 mg/kg) at the onset of the dark phase (time point 12h). In conclusion, this study supports the hypothesis that neonatal treatment with CMI induces a hypersecretion of corticosterone in adulthood that was reversed through treatment with the antidepressant FLX. The CMI-treated rats showed a hyporesponse to cholinergic stimulation with OXO at low doses and at the beginning of the dark phase. Thus, the present results do not support the assumption that an increased sensitivity of the muscarinic cholinergic system is one of the possible correlates of the behavioral alterations seen in CMI-treated rats.

  9. Chronic nerve growth factor administration increases the peripheral exocytotic activity of capsaicin-sensitive cutaneous neurons.

    PubMed

    Bowles, Walter R; Sabino, Ma'lou; Harding-Rose, Catherine; Hargreaves, Kenneth M

    2006-08-01

    Nerve growth factor (NGF) plays an important role in inflammation and pain and has been suggested to regulate the responsiveness and sensitivity of nociceptive fibers. However, no study has evaluated whether chronic NGF alters the exocytotic capacity of peripheral terminals of peptidergic fibers. To test this hypothesis, rats were injected subcutaneously every other day with either murine recombinant NGF (mNGF; 1.0 mg/kg) or vehicle for 7 days; or mNGF (0.1 mg/kg), mNGF (1 mg/kg) or vehicle every other day for 13 days. Treatment of rats with NGF over a 13-day period produced a significant increase in capsaicin-evoked iCGRP release from isolated biopsies of hindpaw skin, as assessed by in vitro superfusion and RIA. This effect was dose-dependent and exhibited a temporal requirement, because the enhancement was only observed after 13 days of treatment and was not evident after 7 days of treatment. This NGF enhancement of capsaicin-evoked iCGRP release was not due solely to increases in peripheral iCGRP content since only the 1mg/kg dose of NGF elevated cutaneous pools of iCGRP, whereas both doses significantly increased capsaicin-evoked peptide release. Moreover, NGF also enhanced capsaicin-evoked thermal hyperalgesia under similar dose- and time-related conditions. Collectively, the chronic administration of NGF not only increases capsaicin-evoked hyperalgesia, but also significantly primes peripheral fibers to enhanced peptidergic exocytosis following activation of the capsaicin receptor. Collectively, these data are consistent with the hypothesis that persistently elevated NGF levels may contribute to enhanced neurogenic regulation of inflammatory and wound healing processes in injured tissue.

  10. Heterogeneity of muscarinic receptor subtypes in cerebral blood vessels

    SciTech Connect

    Garcia-Villalon, A.L.; Krause, D.N.; Ehlert, F.J.; Duckles, S.P. )

    1991-07-01

    The identity and distribution of muscarinic cholinergic receptor subtypes and associated signal transduction mechanisms was characterized for the cerebral circulation using correlated functional and biochemical investigations. Subtypes were distinguished by the relative affinities of a panel of muscarinic antagonists, pirenzepine, AF-DX 116 (11-2-((2-(diethylaminomethyl)- 1-piperidinyl)acetyl)-5,11-dihydro-6H- pyrido(2,3-b)(1,4)benzodiazepine-6-one), hexahydrosiladifenidol, methoctramine, 4-diphenylacetoxy-N-methylpiperidine methobromide, dicyclomine, para-fluoro-hexahydrosiladifenidol and atropine. Muscarinic receptors characterized by inhibition of (3H)quinuclidinylbenzilate binding in membranes of bovine pial arteries were of the M2 subtype. In contrast pharmacological analysis of (3H)-quinuclidinylbenzilate binding in bovine intracerebral microvessels suggests the presence of an M4 subtype. Receptors mediating endothelium-dependent vasodilation in rabbit pial arteries were of the M3 subtype, whereas muscarinic receptors stimulating endothelium-independent phosphoinositide hydrolysis in bovine pial arteries were of the M1 subtype. These findings suggest that characteristics of muscarinic receptors in cerebral blood vessels vary depending on the type of vessel, cellular location and function mediated.

  11. The effects of sigma (σ1) receptor-selective ligands on muscarinic receptor antagonist-induced cognitive deficits in mice

    PubMed Central

    Malik, Maninder; Rangel-Barajas, Claudia; Sumien, Nathalie; Su, Chang; Singh, Meharvan; Chen, Zhenglan; Huang, Ren-Qi; Meunier, Johann; Maurice, Tangui; Mach, Robert H; Luedtke, Robert R

    2015-01-01

    Background and Purpose Cognitive deficits in patients with Alzheimer's disease, Parkinson's disease, traumatic brain injury and stroke often involve alterations in cholinergic signalling. Currently available therapeutic drugs provide only symptomatic relief. Therefore, novel therapeutic strategies are needed to retard and/or arrest the progressive loss of memory. Experimental Approach Scopolamine-induced memory impairment provides a rapid and reversible phenotypic screening paradigm for cognition enhancement drug discovery. Male C57BL/6J mice given scopolamine (1 mg·kg−1) were used to evaluate the ability of LS-1–137, a novel sigma (σ1) receptor-selective agonist, to improve the cognitive deficits associated with muscarinic antagonist administration. Key Results LS-1–137 is a high-affinity (Ki = 3.2 nM) σ1 receptor agonist that is 80-fold selective for σ1, compared with σ2 receptors. LS-1–137 binds with low affinity at D2-like (D2, D3 and D4) dopamine and muscarinic receptors. LS-1–137 was found to partially reverse the learning deficits associated with scopolamine administration using a water maze test and an active avoidance task. LS-1–137 treatment was also found to trigger the release of brain-derived neurotrophic factor from rat astrocytes. Conclusions and Implications The σ1 receptor-selective compound LS-1–137 may represent a novel candidate cognitive enhancer for the treatment of muscarinic receptor-dependent cognitive deficits. PMID:25573298

  12. Effects of nicotinic and muscarinic compounds on biting attack in the cat.

    PubMed

    Berntson, G G; Beattie, M S; Walker, J M

    1976-09-01

    Predatory-like biting attack on a rat, as well as hissing, growling, and other threat behaviors, could be induced in normally non-aggressive cats by systemic administration of the muscarinic agonist, arecoline (7-12 mg/kg). In contrast to arecoline, nicotine was found to suppress aggressive behaviors. Systemic administration of nicotine (0.5 mg/kg) prior to arecoline injection resulted in a significant reduction in elicited attack and threat behaviors. Furthermore, nicotine (0.075-0.500 mg/kg) was found to produce a dose-dependent suppression of natural predatory behavior as well. This nicotine-produced suppression of attack did not appear to be due to the induction of general malaise, since attack suppression could be seen in the absence of general behavioral inhibition, and doses of nicotine resulting in complete suppression of attack had little effect on food intake. Results indicate that muscarinic and nicotinic compounds can exert antagonistic control over some types of aggressive behaviors. PMID:1033569

  13. Tramadol state-dependent memory: involvement of dorsal hippocampal muscarinic acetylcholine receptors.

    PubMed

    Jafari-Sabet, Majid; Jafari-Sabet, Ali-Reza; Dizaji-Ghadim, Ali

    2016-08-01

    The effects on tramadol state-dependent memory of bilateral intradorsal hippocampal (intra-CA1) injections of physostigmine, an acetylcholinesterase inhibitor, and atropine, a muscarinic acetylcholine receptor antagonist, were examined in adult male NMRI mice. A single-trial step-down passive avoidance task was used for the assessment of memory retention. Post-training intra-CA1 administration of an atypical μ-opioid receptor agonist, tramadol (0.5 and 1 μg/mouse), dose dependently impaired memory retention. Pretest injection of tramadol (0.5 and 1 μg/mouse, intra-CA1) induced state-dependent retrieval of the memory acquired under the influence of post-training tramadol (1 μg/mouse, intra-CA1). A pretest intra-CA1 injection of physostigmine (1 μg/mouse) reversed the memory impairment induced by post-training administration of tramadol (1 μg/mouse, intra-CA1). Moreover, pretest administration of physostigmine (0.5 and 1 μg/mouse, intra-CA1) with an ineffective dose of tramadol (0.25 μg/mouse, intra-CA1) also significantly restored retrieval. Pretest administration of physostigmine (0.25, 0.5, and 1 μg/mouse, intra-CA1) by itself did not affect memory retention. A pretest intra-CA1 injection of the atropine (1 and 2 μg/mouse) 5 min before the administration of tramadol (1 μg/mouse, intra-CA1) dose dependently inhibited tramadol state-dependent memory. Pretest administration of atropine (0.5, 1, and 2 μg/mouse, intra-CA1) by itself did not affect memory retention. It can be concluded that dorsal hippocampal muscarinic acetylcholine receptor mechanisms play an important role in the modulation of tramadol state-dependent memory.

  14. Increased ventilatory response to carbon dioxide in COPD patients following vitamin C administration

    PubMed Central

    Hartmann, Sara E.; Kissel, Christine K.; Szabo, Lian; Walker, Brandie L.; Leigh, Richard; Anderson, Todd J.

    2015-01-01

    Patients with chronic obstructive pulmonary disease (COPD) have decreased ventilatory and cerebrovascular responses to hypercapnia. Antioxidants increase the ventilatory response to hypercapnia in healthy humans. Cerebral blood flow is an important determinant of carbon dioxide/hydrogen ion concentration at the central chemoreceptors and may be affected by antioxidants. It is unknown whether antioxidants can improve the ventilatory and cerebral blood flow response in individuals in whom these are diminished. Thus, we aimed to determine the effect of vitamin C administration on the ventilatory and cerebrovascular responses to hypercapnia during healthy ageing and in COPD. Using transcranial Doppler ultrasound, we measured the ventilatory and cerebral blood flow responses to hyperoxic hypercapnia before and after an intravenous vitamin C infusion in healthy young (Younger) and older (Older) subjects and in moderate COPD. Vitamin C increased the ventilatory response in COPD patients (mean (95% CI) 1.1 (0.9–1.1) versus 1.5 (1.1–2.0) L·min−1·mmHg−1, p<0.05) but not in Younger (2.5 (1.9–3.1) versus 2.4 (1.9–2.9) L·min−1·mmHg−1, p>0.05) or Older (1.3 (1.0–1.7) versus 1.3 (1.0–1.7) L·min−1·mmHg−1, p>0.05) healthy subjects. Vitamin C did not affect the cerebral blood flow response in the young or older healthy subjects or COPD subjects (p>0.05). Vitamin C increases the ventilatory but not cerebrovascular response to hyperoxic hypercapnia in patients with moderate COPD. PMID:27730137

  15. 77 FR 5027 - Food and Drug Administration Transparency Initiative: Exploratory Program To Increase Access to...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-01

    ... HUMAN SERVICES Food and Drug Administration Food and Drug Administration Transparency Initiative... and Human Services responding to a January 18, 2011, Presidential Memorandum on Regulatory Compliance, (76 FR 3825, January 21, 2011), FDA recounted the actions it had already implemented, as well as...

  16. Characterization of muscarinic receptor subtypes in human tissues

    SciTech Connect

    Giraldo, E.; Martos, F.; Gomez, A.; Garcia, A.; Vigano, M.A.; Ladinsky, H.; Sanchez de La Cuesta, F.

    1988-01-01

    The affinities of selective, pirenzepine and AF-DX 116, and classical, N-methylscopolamine and atropine, muscarinic cholinergic receptor antagonists were investigated in displacement binding experiments with (/sup 3/H)Pirenzepine and (/sup 3/H)N-methylscopolamine in membranes from human autoptic tissues (forebrain, cerebellum, atria, ventricle and submaxillary salivary glands). Affinity estimates of N-methylscopolamine and atropine indicated a non-selective profile. Pirenzepine showed differentiation between the M/sub 1/ neuronal receptor of the forebrain and the receptors in other tissues while AF-DX 116 clearly discriminated between muscarinic receptors of heart and glands. The results in human tissues confirm the previously described selectivity profiles of pirenzepine and AF-DX 116 in rat tissues. These findings thus reveal the presence also in man of three distinct muscarinic receptor subtypes: the neuronal M/sub 1/, the cardiac M/sub 2/ and the glandular M/sub 3/.

  17. Systemic amitriptyline administration does not prevent the increased thermal response induced by paradoxical sleep deprivation.

    PubMed

    Damasceno, Fabio; Skinner, Gabriela O; Gomes, Aline; Araújo, Paulo C; de Almeida, Olga M M S

    2009-11-01

    Sleep deprivation has been associated with hyperalgesia in humans and in animal models. The tricyclic antidepressant amitriptyline is used as an analgesic drug in patients and in animal models of chronic pain, including that associated with spinal nerve injury. Pain hypersensitivity following paradoxical sleep deprivation (PSD) and that following peripheral nerve injury seem to share common spinal mechanisms. Accordingly, we evaluated the effects of amitriptyline (acutely and chronically administered) on the increased thermal response observed in PSD rats (72 or 96 h). Rats were evaluated for thermal sensitivity using a hot plate (52 degrees C or 46 degrees C) at 1 or 24 h after the last administration of the drug. Following the hot plate test, motor behavior was analyzed in an open field arena for a period of 5 min. Paw withdrawal latency response to temperatures of 46 degrees C and 52 degrees C was significantly lower in PSD and in 24-hour post-PSD rats than in controls and it was not modified by amitriptyline (3, 10 and 30 mg/kg). Analgesic effects and reduced motor behavior were only observed in control groups. Overall, these findings indicate that a period of PSD can influence pain modulatory mechanisms, and that amitriptyline action is insufficient to reduce PSD-enhanced thermal sensitivity.

  18. Planning and Delivering Instruction with Increasing Class Sizes in Educational Administration Program Coursework: Modeling Leadership Skills for New Professors Transitioning from K-12 Administration

    ERIC Educational Resources Information Center

    Stebbins, Gary

    2009-01-01

    Increased class sizes and advising responsibilities are the new realities in California's graduate programs of Educational Administration. In order to effectively meet new challenges, professors must make adjustments in venue, plan meticulously, utilize technology, distribute leadership, and implement alternative grading systems. This is a…

  19. The new E.U. Animal Transport Regulation: improved welfare and health or increased administration?

    PubMed

    Hartung, J

    2006-03-01

    There is public discussion of the new E.U. Animal Transport Regulation No 1/2005 of Dec. 2004 and its advantages and draw-backs. This Regulation is no longer a Directive, so that it is directly applicable in the Members States. Although the Regulation is recognised to have great potential to improve welfare and health of transported animals, it will also increase administrative work. Most improvements will come through better education and the increased responsibilities of animal attendants, drivers, keepers and transport organisers, and through the stricter control mechanisms (log book, training, instructions etc.) and the introduction of the GPS control systems to further enhance the transparency of animal movements. The formats of the transport certificates used in all Member States will be harmonised. Technical records will be kept on air temperature and water consumption. Contact offices in all member states for transport affairs will improve the exchange of data between the responsible authorities and harmonise control and surveillance practice. Specific regulations are now in place for horses (broken, unbroken, registered) and for the transport age of young animals (piglets, lambs, calves, foals). In spite of some substantial improvements there are still significant gaps in our knowledge of both normal and long transports, for example optimal journey times, food and water supply on long transports, environmental factors such as vibration, motion, light and ventilation requirements in different European geographical regions. The same is true for the epidemiological aspects of the prevention of disease transmission; for example, very little is known about the bacterial and particulate emissions of the animal transport vehicles which travel across Europe. A serious drawback of the regulation is the fact that it does not abolish the unloading of animals on long transports to rest for 24 h at staging points, so that the concomitant risks to health and welfare

  20. Chronic ethanol feeding produces a muscarinic receptor upregulation, but not a muscarinic supersensitivity in lower esophageal sphincter muscle.

    PubMed

    Keshavarzian, A; Gordon, J H; Willson, C; Urban, G; Fields, J Z

    1992-02-01

    Muscarinic acetylcholine receptors (mAChR) are important in esophageal physiology, and mAChR alterations may be involved in ethanol-induced esophageal dysfunction. We previously demonstrated that acute ethanol decreases lower esophageal sphincter pressure (LESP), whereas withdrawal from chronic ethanol results in pressure increases which are reversible by acute ethanol. To see if this increase in LESP is due to upregulation of mAChR, we evaluated both mAChR binding and dose-response curves for bethanechol and atropine-induced changes in LESP before and after acute and chronic ethanol exposure. The number of mAChR sites (Bmax) in LES (3.4 fmol/mg tissue) was lowered by acute ethanol (1.72, -50%); withdrawal from chronic ethanol raised Bmax (5.2, +54%). Acute injection of ethanol into cats in withdrawal reversed this increase in mAChR density (3.1, -10%). These changes correlated with our earlier data on ethanol-induced changes in LESP. However, the dose-response curve for bethanechol-induced pressure increases shifted to the right [ED25 (micrograms/kg); control, 8.6; withdrawal, 21.3], paralleled by an increase in the number of low-affinity agonist binding sites. Thus, 1) the withdrawal-associated increase in Bmax (up-regulation) is more likely to be a compensatory response to deficits (functional subsensitivity) distal to the receptor recognition site than to proximal deficits; 2) the increase in Bmax does not cause LESP hyperactivity; and 3) receptor binding changes do not necessarily translate into physiological changes. PMID:1346638

  1. Muscarinic receptors transform NIH 3T3 cells through a Ras-dependent signalling pathway inhibited by the Ras-GTPase-activating protein SH3 domain.

    PubMed Central

    Mattingly, R R; Sorisky, A; Brann, M R; Macara, I G

    1994-01-01

    Expression of certain subtypes of human muscarinic receptors in NIH 3T3 cells provides an agonist-dependent model of cellular transformation by formation of foci in response to carbachol. Although focus formation correlates with the ability of the muscarinic receptors to activate phospholipase C, the actual mitogenic signal transduction pathway is unknown. Through cotransfection experiments and measurement of the activation state of native and epitope-tagged Ras proteins, the contributions of Ras and Ras GTPase-activating protein (Ras-GAP) to muscarinic receptor-dependent transformation were defined. Transforming muscarinic receptors were able to activate Ras, and such activation was required for transformation because focus formation was inhibited by coexpression of either Ras with a dominant-negative mutation or constructs of Ras-GAP that include the catalytic domain. Coexpression of the N-terminal region of GAP or of its isolated SH3 (Src homology 3) domain, but not its SH2 domain, was also sufficient to suppress muscarinic receptor-dependent focus formation. Point mutations at conserved residues in the Ras-GAP SH3 domain reversed its action, leading to an increase in carbachol-dependent transformation. The inhibitory effect of expression of the Ras-GAP SH3 domain occurs proximal to Ras activation and is selective for the mitogenic pathway activated by carbachol, as cellular transformation by either v-Ras or trkA/nerve growth factor is unaffected. Images PMID:7969134

  2. Pontine Reticular Formation (PnO) Administration of Hypocretin-1 Increases PnO GABA Levels and Wakefulness

    PubMed Central

    Watson, Christopher J.; Soto-Calderon, Haideliza; Lydic, Ralph; Baghdoyan, Helen A.

    2008-01-01

    Study Objectives: GABAergic transmission in the oral part of the pontine reticular formation (PnO) increases wakefulness. The hypothalamic peptide hypocretin-1 (orexin A) promotes wakefulness, and the PnO receives hypocretinergic input. The present study tested the hypothesis that PnO administration of hypocretin-1 increases PnO GABA levels and increases wakefulness. This study also tested the hypothesis that wakefulness is either increased or decreased, respectively, by PnO administration of drugs known to selectively increase or decrease GABA levels. Design: A within-subjects design was used for microdialysis and microinjection experiments. Setting: University of Michigan. Patients or Participants: Experiments were performed using adult male Crl:CD® (SD)IGS BR (Sprague-Dawley) rats (n = 46). Interventions: PnO administration of hypocretin-1, nipecotic acid (a GABA uptake inhibitor that increases extracellular GABA levels), 3-mercaptopropionic acid (a GABA synthesis inhibitor that decreases extracellular GABA levels; 3-MPA), and Ringer solution (vehicle control). Measurements and Results: Dialysis administration of hypocretin-1 to the PnO caused a statistically significant, concentration-dependent increase in PnO GABA levels. PnO microinjection of hypocretin-1 or nipecotic acid caused a significant increase in wakefulness and a significant decrease in non-rapid eye movement (NREM) sleep and REM sleep. Microinjecting 3-MPA into the PnO caused a significant increase in NREM sleep and REM sleep and a significant decrease in wakefulness. Conclusions: An increase or a decrease in PnO GABA levels causes an increase or decrease, respectively, in wakefulness. Hypocretin-1 may promote wakefulness, at least in part, by increasing GABAergic transmission in the PnO. Citation: Watson CJ; Soto-Calderon H; Lydic R; Baghdoyan HA. Pontine reticular formation (PnO) administration of hypocretin-1 increases PnO GABA levels and wakefulness. SLEEP 2008;31(4):453-464. PMID:18457232

  3. Effect of central muscarinic receptors on passive-avoidance learning deficits induced by prenatal pentylenetetrazol kindling in male offspring.

    PubMed

    Pourmotabbed, A; Mahmoodi, G; Mahmoodi, S; Mohammadi-Farani, A; Nedaei, S E; Pourmotabbed, T; Pourmotabbed, T

    2014-10-24

    Occurrence of the epileptic seizures during gestation might affect the neurodevelopment of the fetus resulting in cognitive problems for the child later in life. We have previously reported that prenatal pentylenetetrazol (PTZ)-kindling induces learning and memory deficits in the children born to kindled mothers, later in life but the mechanisms involved in this processes are unknown. The cholinergic system plays a major role in learning and memory. The present study was performed to investigate the possible involvement of central muscarinic cholinergic receptors on learning and memory deficits induced by prenatal PTZ-kindling in male offspring. Pregnant Wistar rats were kindled by repetitive i.p. injection of 25mg/kg of PTZ on day 13 of their pregnancy. The effect of intracerebroventricular (ICV) microinjection of scopolamine and pilocarpine, muscarinic cholinergic receptors antagonist and agonist, respectively on passive-avoidance learning of pups were tested at 12weeks of age using shuttle-box apparatus. Our data showed that the retention latencies of pups that received scopolamine (2 or 3μg) were significantly reduced compared to those received normal saline (p<0.05). Interestingly, post training ICV administration of pilocarpine (2μg) retrieved pups' memory deficits (p<0.001). These results demonstrate for the first time, the importance of the central muscarinic cholinergic receptors in learning and memory deficits in pups born to kindled dams and suggest a central mechanism for the cognitive and memory dysfunction, associated with seizures during pregnancy.

  4. Supraphysiologic glucocorticoid administration increased biomechanical bone strength of rats' vertebral body

    PubMed Central

    Najar, Azam; Fridoni, Mohammadjavad; Rezaei, Fatemesadat; Bayat, Saba

    2015-01-01

    The aim of this study is to assess the effects of different glucocorticoid administration protocols on biomechanical properties of the first lumbar vertebral body in rats. We divided 40 male rats into the following groups: control, dexamethasone (7 mg/week), dexamethasone (0.7 mg/week), methylprednisolone (7 mg/kg/week), methylprednisolone (5 mg/kg twice weekly), dexamethasone (7 mg/kg three times per week), dexamethasone (0.7 mg/kg three times per week, and low-level laser treated rats. Lumbar vertebrae in rats were exposed to the pulsed laser. We conducted a biomechanical test to examine the mechanical properties of vertebral body in rats' lumbar bone. Supraphysiologic glucocorticoid administration protocols did not impair the biomechanical properties of rats' vertebral bodies compared to control and laser-treated rats. Supraphysiologic glucocorticoid administration caused an anabolic effect on the vertebral bodies. PMID:26755921

  5. An Antibody Biosensor Establishes the Activation of the M1 Muscarinic Acetylcholine Receptor during Learning and Memory*♦

    PubMed Central

    Butcher, Adrian J.; Bradley, Sophie J.; Prihandoko, Rudi; Brooke, Simon M.; Mogg, Adrian; Bourgognon, Julie-Myrtille; Macedo-Hatch, Timothy; Edwards, Jennifer M.; Bottrill, Andrew R.; Challiss, R. A. John; Broad, Lisa M.; Felder, Christian C.; Tobin, Andrew B.

    2016-01-01

    Establishing the in vivo activation status of G protein-coupled receptors would not only indicate physiological roles of G protein-coupled receptors but would also aid drug discovery by establishing drug/receptor engagement. Here, we develop a phospho-specific antibody-based biosensor to detect activation of the M1 muscarinic acetylcholine receptor (M1 mAChR) in vitro and in vivo. Mass spectrometry phosphoproteomics identified 14 sites of phosphorylation on the M1 mAChR. Phospho-specific antibodies to four of these sites established that serine at position 228 (Ser228) on the M1 mAChR showed extremely low levels of basal phosphorylation that were significantly up-regulated by orthosteric agonist stimulation. In addition, the M1 mAChR-positive allosteric modulator, 1-(4-methoxybenzyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid, enhanced acetylcholine-mediated phosphorylation at Ser228. These data supported the hypothesis that phosphorylation at Ser228 was an indicator of M1 mAChR activation. This was further supported in vivo by the identification of phosphorylated Ser228 on the M1 mAChR in the hippocampus of mice following administration of the muscarinic ligands xanomeline and 1-(4-methoxybenzyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid. Finally, Ser228 phosphorylation was seen to increase in the CA1 region of the hippocampus following memory acquisition, a response that correlated closely with up-regulation of CA1 neuronal activity. Thus, determining the phosphorylation status of the M1 mAChR at Ser228 not only provides a means of establishing receptor activation following drug treatment both in vitro and in vivo but also allows for the mapping of the activation status of the M1 mAChR in the hippocampus following memory acquisition thereby establishing a link between M1 mAChR activation and hippocampus-based memory and learning. PMID:26826123

  6. In vivo visualization of central muscarinic receptors using [11C]quinuclidinyl benzilate and positron emission tomography in baboons.

    PubMed

    Varastet, M; Brouillet, E; Chavoix, C; Prenant, C; Crouzel, C; Stulzaft, O; Bottlaender, M; Cayla, J; Mazière, B; Mazière, M

    1992-03-24

    The muscarinic antagonist, quinuclidinyl benzilate (QNB), labeled with carbon 11 was used as a radioligand to visualize in vivo by positron emission tomography (PET) the central muscarinic acetylcholine receptors (mAChR) in baboons (Papio papio). The binding characteristics of [11C]QNB showed its specific binding to central mAChR. [11C]QNB brain uptake was high in cerebral cortex and striatum, areas that are rich in mAChR, whereas it decreased rapidly in cerebellum, evidencing non-specific binding in this structure that is almost devoid of mAChR. These results are consistent with the known cerebral distribution of mAChR in primates. [11C]QNB specific cerebral binding was enhanced by pretreatment with methyl-QNB, a peripherally acting muscarinic antagonist. Specifically labeled binding sites alone were blocked by prior administration of dexetimide, a muscarinic antagonist. Specific radioactivity was driven out from mAChR-rich regions by atropine and dexetimide, drugs with high affinity for mAChR. This competition was stereospecific since only dexetimide, the pharmacologically active isomer of benzetimide, was able to compete with the radioligand on its binding sites. A relationship between the occupancy of [11C]QNB-labeled receptors by atropine or dexetimide and the concomitant induction of a pharmacological effect was also detected by simultaneous PET scanning and electroencephalographic recording. Since mAChR form an important part of choline receptors in the central nervous system, [11C]QNB appears to be a suitable radiotracer to monitor cerebral physiological or pathological phenomena linked to the cholinergic system in living subjects. PMID:1521561

  7. Perirhinal Cortex Muscarinic Receptor Blockade Impairs Taste Recognition Memory Formation

    ERIC Educational Resources Information Center

    Gutierrez, Ranier; De la Cruz, Vanesa; Rodriguez-Ortiz, Carlos J.; Bermudez-Rattoni, Federico

    2004-01-01

    The relevance of perirhinal cortical cholinergic and glutamatergic neurotransmission for taste recognition memory and learned taste aversion was assessed by microinfusions of muscarinic (scopolamine), NMDA (AP-5), and AMPA (NBQX) receptor antagonists. Infusions of scopolamine, but not AP5 or NBQX, prevented the consolidation of taste recognition…

  8. Increasing the Transfer of Learning through Problem-Based Learning in Educational Administration.

    ERIC Educational Resources Information Center

    Cordiero, Paula A.; Campbell, Barbara

    Since the 1950s some educational researchers have argued for a strong focus on problem solving in administrator-preparation programs. This paper discusses two types of problem-based learning (PBL)--simulated and authentic. It discusses various PBL concepts and presents two vignettes used during the 1995 and 1996 academic years at the University of…

  9. U.S. Public Administration Programs: Increasing Academic Achievement by Identifying and Utilizing Student Learning Styles

    ERIC Educational Resources Information Center

    Naylor, Lorenda A; Wooldridge, Blue; Lyles, Alan

    2014-01-01

    Global economic shifts are forcing universities to become more competitive and operationally efficient. As a result, universities emphasize access, affordability, and achievement. More specifically, U.S. universities have responded by emphasizing course assessment, retention rates, and graduation rates. Both university administrators and faculty…

  10. Ethanol impairs muscarinic receptor-induced neuritogenesis in rat hippocampal slices: Role of astrocytes and extracellular matrix proteins.

    PubMed

    Giordano, Gennaro; Guizzetti, Marina; Dao, Khoi; Mattison, Hayley A; Costa, Lucio G

    2011-12-01

    In an in vitro co-culture system of astrocytes and neurons, stimulation of cholinergic muscarinic receptors in astrocytes had been shown to cause neuritogenesis in hippocampal neurons, and this effect was inhibited by ethanol. The present study sought to confirm these earlier findings in a more complex system, in vitro rat hippocampal slices in culture. Exposure of hippocampal slices to the cholinergic agonist carbachol (1mM for 24h) induced neurite outgrowth in hippocampal pyramidal neurons, which was mediated by activation of muscarinic M3 receptors. Specifically, carbachol induced a >4-fold increase in the length of the longest neurite, and a 4-fold increase in the length of minor neurites and in the number of branches. Co-incubation of carbachol with ethanol (50mM) resulted in significant inhibition of the effects induced by carbachol on all parameters measured. Neurite outgrowth in CNS neurons is dependent on various permissive factors that are produced and released by glial cells. In hippocampal slices carbachol increased the levels of two extracellular matrix protein, fibronectin and laminin-1, by 1.6-fold, as measured by Western blot. Co-incubation of carbachol with ethanol significantly inhibited these increases. Carbachol-induced increases in levels of extracellular matrix proteins were antagonized by a M3 muscarinic receptor antagonist. Furthermore, function-blocking fibronectin or laminin-1 antibodies antagonized the effect of carbachol on neurite outgrowth. These results indicate that in hippocampal slices stimulation of muscarinic M3 receptors induces neurite outgrowth, which is mediated by fibronectin and laminin-1, two extracellular matrix proteins released by astrocytes. By decreasing fibronectin and laminin levels ethanol prevents carbachol-induced neuritogenesis. These findings highlight the importance of glial-neuronal interactions as important targets in the developmental neurotoxicity of alcohol.

  11. Unexpected antipsychotic-like activity with the muscarinic receptor ligand (5R,6R)6-(3-propylthio-1,2,5-thiadiazol-4-yl)-1-azabicyclo[3.2.1]octane .

    PubMed

    Bymaster, F P; Shannon, H E; Rasmussen, K; Delapp, N W; Mitch, C H; Ward, J S; Calligaro, D O; Ludvigsen, T S; Sheardown, M J; Olesen, P H; Swedberg, M D; Sauerberg, P; Fink-Jensen, A

    1998-09-01

    (5R,6R)6-(3-propylthio-1,2,5-thiadiazol-4-yl)-1-azabicyclo[3 .2.1]octane (PTAC) is a potent muscarinic receptor ligand with high affinity for central muscarinic receptors and no or substantially less affinity for a large number of other receptors or binding sites including dopamine receptors. The ligand exhibits partial agonist effects at muscarinic M2 and M4 receptors and antagonist effects at muscarinic M1, M3 and M5 receptors. PTAC inhibited conditioned avoidance responding, dopamine receptor agonist-induced behavior and D-amphetamine-induced FOS protein M5 expression in the nucleus accumbens without inducing catalepsy, tremor or salivation at pharmacologically relevant doses. The effect of PTAC on conditioned avoidance responding and dopamine receptor agonist-induced behavior was antagonized by the acetylcholine receptor antagonist scopolamine. The compound selectively inhibited dopamine cell firing (acute administration) as well as the number of spontaneously active dopamine cells (chronic administration) in the limbic ventral tegmental area (A10) relative to the non-limbic substantia nigra, pars compacta (A9). The results demonstrate that PTAC exhibits functional dopamine receptor antagonism despite its lack of affinity for the dopamine receptors and indicate that muscarinic receptor partial agonists may be an important new approach in the medical treatment of schizophrenia.

  12. Novel alkoxy-oxazolyl-tetrahydropyridine muscarinic cholinergic receptor antagonists.

    PubMed

    Shannon, H E; Bymaster, F P; Hendrix, J C; Quimby, S J; Mitch, C H

    1995-01-01

    The purpose of the present studies was to compare a novel series of alkoxy-oxazolyl-tetrahydropyridines (A-OXTPs) as muscarinic receptor antagonists. The affinity of these compounds for muscarinic receptors was determined by inhibition of [3H]pirenzepine to M1 receptors in hippocampus, [3H]QNB to M2 receptors in brainstem, and [3H]oxotremorine-M to high affinity muscarinic agonist binding sites in cortex. All of the compounds had higher affinity for [3H]pirenzepine than for [3H]QNB or [3H]oxotremorine-M labeled receptors, consistent with an interpretation that they are relatively selective M1 receptor antagonists, although none were as selective as pirenzepine. In addition, dose-response curves were determined for antagonism of oxotremorine-induced salivation (mediated by M3 receptors) and tremor (mediated by non-M1 receptors) in mice. In general, the A-OXTPs were equipotent and equieffective in antagonizing both salivation and tremor, although there were modest differences for some compounds. Dose-response curves also were determined on behavior maintained under a spatial-alternation schedule of food presentation in rats as a measure of effects on working memory. The A-OXTPs produced dose-related decreases in percent correct responding at doses three- to ten-fold lower than those which decreased rates of responding. However, only one compound, MB-OXTP, produced effects on percent correct responding consistent with a selective effect on memory as opposed to non-memory variables. The present results provide evidence that these alkoxy-oxazolyl-tetrahydropyridines are a novel series of modestly M1-selective muscarinic receptor antagonists, and that one member of the series, MB-OXTP, appears to be more selective in its effects on memory than previously studies muscarinic antagonists. PMID:7753969

  13. The distribution of cerebral muscarinic acetylcholine receptors in vivo in patients with dementia. A controlled study with 123IQNB and single photon emission computed tomography

    SciTech Connect

    Weinberger, D.R.; Gibson, R.; Coppola, R.; Jones, D.W.; Molchan, S.; Sunderland, T.; Berman, K.F.; Reba, R.C. )

    1991-02-01

    A high-affinity muscarinic receptor antagonist, 123IQNB (3-quinuclidinyl-4-iodobenzilate labeled with iodine 123), was used with single photon emission computed tomography to image muscarinic acetylcholine receptors in 14 patients with dementia and in 11 healthy controls. High-resolution single photon emission computed tomographic scanning was performed 21 hours after the intravenous administration of approximately 5 mCi of IQNB. In normal subjects, the images of retained ligand showed a consistent regional pattern that correlated with postmortem studies of the relative distribution of muscarinic receptors in the normal human brain, having high radioactivity counts in the basal ganglia, occipital cortex, and insular cortex, low counts in the thalamus, and virtually no counts in the cerebellum. Eight of 12 patients with a clinical diagnosis of Alzheimer's disease had obvious focal cortical defects in either frontal or posterior temporal cortex. Both patients with a clinical diagnosis of Pick's disease had obvious frontal and anterior temporal defects. A region of interest statistical analysis of relative regional activity revealed a significant reduction bilaterally in the posterior temporal cortex of the patients with Alzheimer's disease compared with controls. This study demonstrates the practicability of acetylcholine receptor imaging with 123IQNB and single photon emission computed tomography. The data suggest that focal abnormalities in muscarinic binding in vivo may characterize some patients with Alzheimer's disease and Pick's disease, but further studies are needed to address questions about partial volume artifacts and receptor quantification.

  14. Is albumin administration in the acutely ill associated with increased mortality? Results of the SOAP study

    PubMed Central

    Vincent, Jean-Louis; Sakr, Yasser; Reinhart, Konrad; Sprung, Charles L; Gerlach, Herwig; Ranieri, V Marco

    2005-01-01

    Introduction Albumin administration in the critically ill has been the subject of some controversy. We investigated the use of albumin solutions in European intensive care units (ICUs) and its relationship to outcome. Methods In a cohort, multicenter, observational study, all patients admitted to one of the participating ICUs between 1 May and 15 May 2002 were followed up until death, hospital discharge, or for 60 days. Patients were classified according to whether or not they received albumin at any time during their ICU stay. Results Of 3,147 admitted patients, 354 (11.2%) received albumin and 2,793 (88.8%) did not. Patients who received albumin were more likely to have cancer or liver cirrhosis, to be surgical admissions, and to have sepsis. They had a longer length of ICU stay and a higher mortality rate, but were also more severely ill, as manifested by higher simplified acute physiology score (SAPS) II and sequential organ failure assessment (SOFA) scores than the other patients. A Cox proportional hazard model indicated that albumin administration was significantly associated with decreased 30-day survival. Moreover, in 339 pairs matched according to a propensity score, ICU and hospital mortality rates were higher in the patients who had received albumin than in those who had not (34.8 versus 20.9% and 41.3 versus 27.7%, respectively, both p < 0.001). Conclusion Albumin administration was associated with decreased survival in this population of acutely ill patients. Further prospective randomized controlled trials are needed to examine the effects of albumin administration in sub-groups of acutely ill patients. PMID:16356223

  15. Characterization and agonist regulation of muscarinic ([3H]N-methyl scopolamine) receptors in isolated ventricular myocytes from rat.

    PubMed

    Horackova, M; Robinson, B; Wilkinson, M

    1990-11-01

    Cell surface muscarinic cholinergic receptors have been characterized and quantified for the first time, in intact, isolated adult rat cardiomyocytes. The cells were previously established as functionally fully compatible with cellular responses in intact cardiac tissue. The specific binding of the hydrophilic radioligand, [3H]-NMS, (N-methyl-[3H]-scopolamine methylchloride) was found to be stereo-specific, saturable, reversible and of high affinity. Binding of [3H]-NMS demonstrated appropriate drug specificity and was positively correlated with increasing cell concentrations. Bmax for [3H]-NMS binding to ventricular myocytes, enzymatically dissociated from adult male rats, was 15.8 +/- 1.03 fmol/25 x 10(3) cells (at 4 degrees C) and KD was 0.27 +/- 0.05 nM (n = 14). Binding assays performed at a higher incubation temperature (30 degrees C) yielded a higher Bmax value (22.1 +/- 1.6 fmol/25 x 10(3) cells; n = 11; P less than 0.005 vs. Bmax at 4 degrees C) but an unchanged KD (0.23 +/- 0.06 nM). Pretreatment of myocytes with the muscarinic agonist carbachol (1 mM) at 37 degrees C resulted in a reduction (down-regulation) in specific binding of the hydrophilic ligand [3H]-NMS. The magnitude of this reduction and its rate of recovery were dependent on the time of the exposure to carbachol. Exposures of 30-60 min elicited down-regulated by 35% (Bmax = 14.29 +/- 1.66 changed to 9.5 +/- 1.79 fmol/25 x 10(3) cells, without change in KD P less than 0.01, n = 4). The down-regulation of the muscarinic receptors by carbachol was insensitive to application of bacitracin - an inhibitor of endocytosis. On the other hand preincubation with 10(-9)M atropine, a muscarinic antagonist, hindered the agonist-induced receptor "loss" from the cell surface confirming the muscarinic nature of these receptors. We conclude that our preparation of intact, isolated ventricular cardiomyocytes is ideally suited for the study of cell surface muscarinic receptor regulation under physiological and

  16. Characterization and agonist regulation of muscarinic ([3H]N-methyl scopolamine) receptors in isolated ventricular myocytes from rat.

    PubMed

    Horackova, M; Robinson, B; Wilkinson, M

    1990-11-01

    Cell surface muscarinic cholinergic receptors have been characterized and quantified for the first time, in intact, isolated adult rat cardiomyocytes. The cells were previously established as functionally fully compatible with cellular responses in intact cardiac tissue. The specific binding of the hydrophilic radioligand, [3H]-NMS, (N-methyl-[3H]-scopolamine methylchloride) was found to be stereo-specific, saturable, reversible and of high affinity. Binding of [3H]-NMS demonstrated appropriate drug specificity and was positively correlated with increasing cell concentrations. Bmax for [3H]-NMS binding to ventricular myocytes, enzymatically dissociated from adult male rats, was 15.8 +/- 1.03 fmol/25 x 10(3) cells (at 4 degrees C) and KD was 0.27 +/- 0.05 nM (n = 14). Binding assays performed at a higher incubation temperature (30 degrees C) yielded a higher Bmax value (22.1 +/- 1.6 fmol/25 x 10(3) cells; n = 11; P less than 0.005 vs. Bmax at 4 degrees C) but an unchanged KD (0.23 +/- 0.06 nM). Pretreatment of myocytes with the muscarinic agonist carbachol (1 mM) at 37 degrees C resulted in a reduction (down-regulation) in specific binding of the hydrophilic ligand [3H]-NMS. The magnitude of this reduction and its rate of recovery were dependent on the time of the exposure to carbachol. Exposures of 30-60 min elicited down-regulated by 35% (Bmax = 14.29 +/- 1.66 changed to 9.5 +/- 1.79 fmol/25 x 10(3) cells, without change in KD P less than 0.01, n = 4). The down-regulation of the muscarinic receptors by carbachol was insensitive to application of bacitracin - an inhibitor of endocytosis. On the other hand preincubation with 10(-9)M atropine, a muscarinic antagonist, hindered the agonist-induced receptor "loss" from the cell surface confirming the muscarinic nature of these receptors. We conclude that our preparation of intact, isolated ventricular cardiomyocytes is ideally suited for the study of cell surface muscarinic receptor regulation under physiological and

  17. Heart ventricles specific stress-induced changes in β-adrenoceptors and muscarinic receptors.

    PubMed

    Tillinger, Andrej; Novakova, Martina; Krizanova, Olga; Kvetnansky, Richard; Myslivecek, Jaromir

    2014-01-01

    The left and right ventricles fulfill different role in heart function. Here we compare chamber specific changes in local catecholamine concentrations; gene expression and the receptor protein amount of all three β-adrenoceptors (β-AR) in rat right heart ventricles exposed to acute (1 session) and repeated (7 sessions) immobilization stress (IMMO) vs. previously observed changes in left ventricles. Density of muscarinic receptors as main cardio-inhibitive receptors was also measured. In the right ventricles, noradrenaline and adrenaline were increased. No β1-AR changes were observed, in spite of the increased sympathetic activity. On the other hand, we have found a decrease of β2-AR gene expression (reduction to 30%) after 7 IMMO and protein (to 59%) after 1 IMMO. β3-AR gene expression was increased after 7 IMMO. Muscarinic receptor density was not changed. When comparing correlation in left and right ventricles, there was strong correlation between adrenaline and β2-AR gene expression, protein and β3-AR gene expression in the left ventricles while only correlation between adrenaline and β2-AR mRNA and protein in the right ventricles was found. Our results show that maintenance of cardiac homeostasis under stress conditions are to a great extent achieved by a balance between different receptors and also by a balanced receptor changes in left vs. right ventricles. Taken together, decrease of cardio-stimulating β2-AR represents a new important mechanism by which β2-AR contributes to the heart physiology.

  18. Endosulfan and cholinergic (muscarinic) transmission: effect on electroencephalograms and (/sup 3/H)quinuclidinyl benzilate in pigeon brain

    SciTech Connect

    Anand, M.; Agrawal, A.K.; Gopal, K.; Sur, R.N.; Seth, P.K.

    1986-08-01

    Single exposure of endosulfan (5 mg/kg) to pigeons (Columbia livia) caused neuronal hyperexcitability as evidence by spike discharges of 200-500 ..mu..V in the electroencephalograms (EEG) from the telencephalon and hyperstriatum, but there was not effect on the ectostriatal area. Cholinergic (muscarinic) receptor binding study using (/sup 3/H)quinuclidinyl benzilate ((/sup 3/H)QNB) as a specific ligand indicated that a single exposure to 5 mg/kg of endosulfan caused a significant increase in (/sup 3/H)QNB binding to the striatal membrane. Behavior study further indicated that a single dose of 200 ..mu..g/kg of oxotremorine produced a significant induction in the tremor in endosulfan-pretreated pigeons. The results of this behavioral and biochemical study indicate the involvement of a cholinergic (muscarinic) transmitter system in endosulfan-induced neurotoxicity.

  19. Increase in cocaine- and amphetamine-regulated transcript (CART) in specific areas of the mouse brain by acute caffeine administration.

    PubMed

    Cho, Jin Hee; Cho, Yun Ha; Kim, Hyo Young; Cha, Seung Ha; Ryu, Hyun; Jang, Wooyoung; Shin, Kyung Ho

    2015-04-01

    Caffeine produces a variety of behavioral effects including increased alertness, reduced food intake, anxiogenic effects, and dependence upon repeated exposure. Although many of the effects of caffeine are mediated by its ability to block adenosine receptors, it is possible that other neural substrates, such as cocaine- and amphetamine-regulated transcript (CART), may be involved in the effects of caffeine. Indeed, a recent study demonstrated that repeated caffeine administration increases CART in the mouse striatum. However, it is not clear whether acute caffeine administration alters CART in other areas of the brain. To explore this possibility, we investigated the dose- and time-dependent changes in CART immunoreactivity (CART-IR) after a single dose of caffeine in mice. We found that a high dose of caffeine (100 mg/kg) significantly increased CART-IR 2 h after administration in the nucleus accumbens shell (AcbSh), dorsal bed nucleus of the stria terminalis (dBNST), central nucleus of the amygdala (CeA), paraventricular hypothalamic nucleus (PVN), arcuate hypothalamic nucleus (Arc), and locus coeruleus (LC), and returned to control levels after 8 h. But this increase was not observed in other brain areas. In addition, caffeine administration at doses of 25 and 50 mg/kg appears to produce dose-dependent increases in CART-IR in these brain areas; however, the magnitude of increase in CART-IR observed at a dose of 50 mg/kg was similar or greater than that observed at a dose of 100 mg/kg. This result suggests that CART-IR in AcbSh, dBNST, CeA, PVN, Arc, and LC is selectively affected by caffeine administration.

  20. Increase in cocaine- and amphetamine-regulated transcript (CART) in specific areas of the mouse brain by acute caffeine administration.

    PubMed

    Cho, Jin Hee; Cho, Yun Ha; Kim, Hyo Young; Cha, Seung Ha; Ryu, Hyun; Jang, Wooyoung; Shin, Kyung Ho

    2015-04-01

    Caffeine produces a variety of behavioral effects including increased alertness, reduced food intake, anxiogenic effects, and dependence upon repeated exposure. Although many of the effects of caffeine are mediated by its ability to block adenosine receptors, it is possible that other neural substrates, such as cocaine- and amphetamine-regulated transcript (CART), may be involved in the effects of caffeine. Indeed, a recent study demonstrated that repeated caffeine administration increases CART in the mouse striatum. However, it is not clear whether acute caffeine administration alters CART in other areas of the brain. To explore this possibility, we investigated the dose- and time-dependent changes in CART immunoreactivity (CART-IR) after a single dose of caffeine in mice. We found that a high dose of caffeine (100 mg/kg) significantly increased CART-IR 2 h after administration in the nucleus accumbens shell (AcbSh), dorsal bed nucleus of the stria terminalis (dBNST), central nucleus of the amygdala (CeA), paraventricular hypothalamic nucleus (PVN), arcuate hypothalamic nucleus (Arc), and locus coeruleus (LC), and returned to control levels after 8 h. But this increase was not observed in other brain areas. In addition, caffeine administration at doses of 25 and 50 mg/kg appears to produce dose-dependent increases in CART-IR in these brain areas; however, the magnitude of increase in CART-IR observed at a dose of 50 mg/kg was similar or greater than that observed at a dose of 100 mg/kg. This result suggests that CART-IR in AcbSh, dBNST, CeA, PVN, Arc, and LC is selectively affected by caffeine administration. PMID:25820086

  1. Abrupt suspension of probiotics administration may increase host pathogen susceptibility by inducing gut dysbiosis

    PubMed Central

    Liu, Zhi; Liu, Wenshu; Ran, Chao; Hu, Jun; Zhou, Zhigang

    2016-01-01

    In this study, we investigated the risk associated with suspension of probiotics administration in tilapia, an animal model that may mimic immune-compromised conditions in humans. Tilapias were fed for 14 days using a probiotics-supplemented diet, followed by a three-day suspension of probiotics treatment and a subsequent challenge by Aeromonas hydrophila. Unexpectedly, the suspension of a probiotic strain Lactobacillus plantarum JCM1149 significantly triggered susceptibility of the host to A. hydrophila. We further observed that suspension of JCM1149 resulted in host gut microbiota dysbiosis and the subsequent disorder in the intestinal metabolites (bile acids, amino acids, and glucose) and damage in the intestinal epithelium, giving rise to a condition similar to antibiotics-induced gut dysbiosis, which collectively impaired tilapia’s gut health and resistance to pathogenic challenges. Additionally, we determined that JCM1149 adhered relatively poorly to tilapia intestinal mucosa and was rapidly released from the gastrointestinal tract (GIT) after suspension, with the rapid loss of probiotic strain probably being the direct cause of gut dysbiosis. Finally, three other probiotic Lactobacillus strains with low intestinal mucosa binding activity showed similar rapid loss phenotype following administration suspension, and induced higher host susceptibility to infection, indicating that the risk is a generic phenomenon in Lactobacillus. PMID:26983596

  2. Effect of perioperative somatostatin administration of sphincteroplasty-induced increase of amylase.

    PubMed

    Roncoroni, L; De Bernardinis, M; Violi, V; Montanari, M; Peracchia, A

    1986-06-01

    Twenty patients undergoing sphincteroplasty for cholelithiasis were randomly divided into two groups of 10. The former (T) were treated with a 4-h somatostatin intravenous drip (250 micrograms/h), started at the beginning of operation, while the latter (C) made up the control group. Serum and urine amylase, amylase creatinine clearance ratio, and liver function tests were assessed for 2 days before surgery, after the operation and for a period of 5 postoperative days. Homogeneity between the two series was verified in experimental conditions. Statistical differences occurred postoperatively in amylase creatinine clearance ratio, which proved higher in C group, and gamma-GT, which was higher in T group. Short-term somatostatin administration proved effective in reducing the postoperative amylase creatinine clearance ratio, although more evident results are reported after long-term administration. Cholestasis or any serious impairment in liver function did not occur, suggesting the suitability of somatostatin use even in patients with jaundice. Since a relationship between postoperative amylase levels and risk of pancreatitis has not yet been proved, the value of somatostatin in the prevention of postoperative pancreatitis after sphincteroplasty needs to be further verified.

  3. Increasing the Use of Group Interventions in a Pediatric Rehabilitation Program: Perceptions of Administrators, Therapists, and Parents

    ERIC Educational Resources Information Center

    Camden, Chantal; Tetreault, Sylvie; Swaine, Bonnie

    2012-01-01

    Objectives: To explore perceptions related to increased utilization of group interventions as a part of the service reorganization within a pediatric rehabilitation program. Methods: Individual interviews with program administrators (n = 13) and focus groups with therapists (n = 19) and parents of children with disabilities (n = 5) were conducted.…

  4. Alterations in alpha-adrenergic and muscarinic cholinergic receptor binding in rat brain following nonionizing radiation

    SciTech Connect

    Gandhi, V.C.; Ross, D.H.

    1987-01-01

    Microwave radiation produces hyperthermia. The mammalian thermoregulatory system defends against changes in temperature by mobilizing diverse control mechanisms. Neurotransmitters play a major role in eliciting thermoregulatory responses. The involvement of adrenergic and muscarinic cholinergic receptors was investigated in radiation-induced hyperthermia. Rats were subjected to radiation at 700 MHz frequency and 15 mW/cm/sup 2/ power density and the body temperature was raised by 2.5 degrees C. Of six brain regions investigated only the hypothalamus showed significant changes in receptor states, confirming its pivotal role in thermoregulation. Adrenergic receptors, studied by (/sup 3/H)clonidine binding, showed a 36% decrease in binding following radiation after a 2.5 degrees C increase in body temperature, suggesting a mechanism to facilitate norepinephrine release. Norepinephrine may be speculated to maintain thermal homeostasis by activating heat dissipation. Muscarinic cholinergic receptors, studied by (3H)quinuclidinyl benzilate binding, showed a 65% increase in binding at the onset of radiation. This may be attributed to the release of acetylcholine in the hypothalamus in response to heat cumulation. The continued elevated binding during the period of cooling after radiation was shut off may suggest the existence of an extra-hypothalamic heat-loss pathway.

  5. Muscarinic receptor-mediated inositol tetrakisphosphate response in bovine adrenal chromaffin cells

    SciTech Connect

    Sanborn, B.B.; Schneider, A.S. )

    1990-01-01

    Inositol trisphosphate (IP{sub 3}), a product of the phosphoinositide cycle, mobilizes intracellular Ca{sup 2+} in many cell types. New evidence suggests that inositol tetrakisphosphate (IP{sub 4}), an IP{sub 3} derivative, may act as another second messenger to further alter calcium homeostasis. However, the function and mechanism of action of IP{sub 4} are presently unresolved. We now report evidence of muscarinic receptor-mediated accumulation of IP{sub 4} in bovine adrenal chromaffin cells, a classic neurosecretory system in which calcium movements have been well studied. Muscarine stimulated an increase in ({sup 3}H)IP{sub 4} and ({sup 3}H)IP{sub 3} accumulation in chromaffin cells and this effect was completely blocked by atropine. ({sup 3}H)IP{sub 4} accumulation was detectable within 15 sec, increased to a maximum by 30 sec and thereafter declined. 2,3-diphosphoglycerate, an inhibitor of IP{sub 3} and IP{sub 4} hydrolysis, enhanced accumulation of these inositol polyphosphates. The results provide the first evidence of a rapid inositol tetrakisphosphate response in adrenal chromaffin cells, which should facilitate the future resolution of the relationship between IP{sub 4} and calcium homeostasis.

  6. Administration's Proposed NSF Budget Includes a 5.5% Increase for Geosciences

    NASA Astrophysics Data System (ADS)

    Showstack, Randy

    2013-04-01

    The fiscal year (FY) 2014 proposed federal budget for the U.S. National Science Foundation (NSF) is $7.63 billion, 7.3% above the FY 2012 actual amount. NSF acting director Cora Marrett said the budget reflects the administration's recognition of NSF and the importance of funding basic research. "We are pleased about where we stand and hope that Congress will be just as pleased with the budget proposal and will help move things forward," she said during a meeting of the NSF Advisory Committee for Geosciences on 11 April. Budget comparisons are to FY 2012 because the 2013 appropriations were enacted at the end of March, less than 2 weeks before President Barack Obama sent the proposed budget to Congress.

  7. The Contingency of Cocaine Administration Accounts for Structural and Functional Medial Prefrontal Deficits and Increased Adrenocortical Activation

    PubMed Central

    Anderson, Rachel M.; Cosme, Caitlin V.; Glanz, Ryan M.; Miller, Mary C.; Romig-Martin, Sara A.; LaLumiere, Ryan T.

    2015-01-01

    The prelimbic region (PL) of the medial prefrontal cortex (mPFC) is implicated in the relapse of drug-seeking behavior. Optimal mPFC functioning relies on synaptic connections involving dendritic spines in pyramidal neurons, whereas prefrontal dysfunction resulting from elevated glucocorticoids, stress, aging, and mental illness are each linked to decreased apical dendritic branching and spine density in pyramidal neurons in these cortical fields. The fact that cocaine use induces activation of the stress-responsive hypothalamo-pituitary-adrenal axis raises the possibility that cocaine-related impairments in mPFC functioning may be manifested by similar changes in neuronal architecture in mPFC. Nevertheless, previous studies have generally identified increases, rather than decreases, in structural plasticity in mPFC after cocaine self-administration. Here, we use 3D imaging and analysis of dendritic spine morphometry to show that chronic cocaine self-administration leads to mild decreases of apical dendritic branching, prominent dendritic spine attrition in PL pyramidal neurons, and working memory deficits. Importantly, these impairments were largely accounted for in groups of rats that self-administered cocaine compared with yoked-cocaine- and saline-matched counterparts. Follow-up experiments failed to demonstrate any effects of either experimenter-administered cocaine or food self-administration on structural alterations in PL neurons. Finally, we verified that the cocaine self-administration group was distinguished by more protracted increases in adrenocortical activity compared with yoked-cocaine- and saline-matched controls. These studies suggest a mechanism whereby increased adrenocortical activity resulting from chronic cocaine self-administration may contribute to regressive prefrontal structural and functional plasticity. SIGNIFICANCE STATEMENT Stress, aging, and mental illness are each linked to decreased prefrontal plasticity. Here, we show that chronic

  8. Iodine-123 N-methyl-4-iododexetimide: a new radioligand for single-photon emission tomographic imaging of myocardial muscarinic receptors.

    PubMed

    Hicks, R J; Kassiou, M; Eu, P; Katsifis, A G; Garra, M; Power, J; Najdovski, L; Lambrecht, R M

    1995-04-01

    Cardiac muscarinic receptor ligands suitable for positron emission tomography have previously been characterised. Attempts to develop radioligands of these receptors suitable for single-photon emission tomographic (SPET) imaging have not been successful due to high lung retention and high non-specific binding of previously investigated potential tracers. The purpose of this study was to evaluate the biodistribution and in vivo imaging characteristics of a new radiopharmaceutical, [123I]N-methyl-4-iododexetimide. Biodistribution studies performed in rats showed high cardiac uptake (2.4% ID/g) 10 min after injection with a heart to lung activity ratio of 5:1. Specificity and stereoselectivity of cardiac binding were demonstrated using blocking experiments in rats. Dynamic imaging studies in anaesthetised greyhounds demonstrated rapid and high myocardial uptake and low lung binding with stable heart to lung activity ratios of > 2.5:1 between 10 and 30 min, making SPECT imaging feasible. Administration of an excess of an unlabelled muscarinic antagonist, methyl-quinuclidinyl benzylate rapidly displaced myocardial activity to background levels and the pharmacologically inactive enantiomer, [123I]N-methyl-4-iodolevetimide, had no detectable cardiac uptake, indicating specific and stereoselective muscarinic receptor binding. SPET revealed higher activity in the inferior than in the anterior wall, this being consistent with previously described regional variation of cardiac parasympathetic innervation. [123I]N-methyl-4-iododexetimide shows promise as an imaging agent for muscarinic receptor distribution in the heart and may be helpful in evaluating diverse cardiac diseases associated with altered muscarinic receptor function, including heart failure and diabetic heart disease. PMID:7607265

  9. M sub 1 muscarinic antagonists interact with. sigma. recognition sites

    SciTech Connect

    Hudkins, R.L. ); DeHaven-Hudkins, D.L. )

    1991-01-01

    The M{sub 1}-selective muscarinic antagonists aprophen, caramiphen, carbetapentane, 2-DAEX, dicyclomine, hexahydrosiladifenidol, iodocaramiphen, nitrocaramiphen, oxybutynin and trihexyphenidyl potently inhibited binding to {sigma} sites in brain. Both basic ester and non-ester structural type compounds which exhibit affinity for the muscarinic receptor also demonstrated affinity for the {sigma} site, while the classical antimuscarinic agents atropine and QNB, and the tricyclic pirenzepine, were ineffective in binding to this site. The authors also observed a significant correlation between the K{sub i} values for {sigma}compounds to inhibit ({sup 3}H)pirenzepine binding and their IC{sub 50} values to inhibit carbachol-stimulated phosphoinositide turnover. These observations may aid in elucidating the relationship of {sigma} binding to inhibition of phosphoinositide turnover stimulated by cholinergic agonists.

  10. Evolution of the toxins muscarine and psilocybin in a family of mushroom-forming fungi.

    PubMed

    Kosentka, Pawel; Sprague, Sarah L; Ryberg, Martin; Gartz, Jochen; May, Amanda L; Campagna, Shawn R; Matheny, P Brandon

    2013-01-01

    Mushroom-forming fungi produce a wide array of toxic alkaloids. However, evolutionary analyses aimed at exploring the evolution of muscarine, a toxin that stimulates the parasympathetic nervous system, and psilocybin, a hallucinogen, have never been performed. The known taxonomic distribution of muscarine within the Inocybaceae is limited, based only on assays of species from temperate regions of the northern hemisphere. Here, we present a review of muscarine and psilocybin assays performed on species of Inocybaceae during the last fifty years. To supplement these results, we used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to determine whether muscarine was present in 30 new samples of Inocybaceae, the majority of which have not been previously assayed or that originated from either the tropics or temperate regions of the southern hemisphere. Our main objective is to test the hypothesis that the presence of muscarine is a shared ancestral feature of the Inocybaceae. In addition, we also test whether species of Inocyabceae that produce psilocybin are monophyletic. Our findings suggest otherwise. Muscarine has evolved independently on several occasions, together with several losses. We also detect at least two independent transitions of muscarine-free lineages to psilocybin-producing states. Although not ancestral for the family as a whole, muscarine is a shared derived trait for an inclusive clade containing three of the seven major lineages of Inocybaceae (the Inocybe, Nothocybe, and Pseudosperma clades), the common ancestor of which may have evolved ca. 60 million years ago. Thus, muscarine represents a conserved trait followed by several recent losses. Transitions to psilocybin from muscarine-producing ancestors occurred more recently between 10-20 million years ago after muscarine loss in two separate lineages. Statistical analyses firmly reject a single origin of muscarine-producing taxa. PMID:23717644

  11. Evolution of the Toxins Muscarine and Psilocybin in a Family of Mushroom-Forming Fungi

    PubMed Central

    Kosentka, Pawel; Sprague, Sarah L.; Ryberg, Martin; Gartz, Jochen; May, Amanda L.; Campagna, Shawn R.; Matheny, P. Brandon

    2013-01-01

    Mushroom-forming fungi produce a wide array of toxic alkaloids. However, evolutionary analyses aimed at exploring the evolution of muscarine, a toxin that stimulates the parasympathetic nervous system, and psilocybin, a hallucinogen, have never been performed. The known taxonomic distribution of muscarine within the Inocybaceae is limited, based only on assays of species from temperate regions of the northern hemisphere. Here, we present a review of muscarine and psilocybin assays performed on species of Inocybaceae during the last fifty years. To supplement these results, we used liquid chromatography–tandem mass spectrometry (LC–MS/MS) to determine whether muscarine was present in 30 new samples of Inocybaceae, the majority of which have not been previously assayed or that originated from either the tropics or temperate regions of the southern hemisphere. Our main objective is to test the hypothesis that the presence of muscarine is a shared ancestral feature of the Inocybaceae. In addition, we also test whether species of Inocyabceae that produce psilocybin are monophyletic. Our findings suggest otherwise. Muscarine has evolved independently on several occasions, together with several losses. We also detect at least two independent transitions of muscarine-free lineages to psilocybin-producing states. Although not ancestral for the family as a whole, muscarine is a shared derived trait for an inclusive clade containing three of the seven major lineages of Inocybaceae (the Inocybe, Nothocybe, and Pseudosperma clades), the common ancestor of which may have evolved ca. 60 million years ago. Thus, muscarine represents a conserved trait followed by several recent losses. Transitions to psilocybin from muscarine-producing ancestors occurred more recently between 10–20 million years ago after muscarine loss in two separate lineages. Statistical analyses firmly reject a single origin of muscarine-producing taxa. PMID:23717644

  12. Evolution of the toxins muscarine and psilocybin in a family of mushroom-forming fungi.

    PubMed

    Kosentka, Pawel; Sprague, Sarah L; Ryberg, Martin; Gartz, Jochen; May, Amanda L; Campagna, Shawn R; Matheny, P Brandon

    2013-01-01

    Mushroom-forming fungi produce a wide array of toxic alkaloids. However, evolutionary analyses aimed at exploring the evolution of muscarine, a toxin that stimulates the parasympathetic nervous system, and psilocybin, a hallucinogen, have never been performed. The known taxonomic distribution of muscarine within the Inocybaceae is limited, based only on assays of species from temperate regions of the northern hemisphere. Here, we present a review of muscarine and psilocybin assays performed on species of Inocybaceae during the last fifty years. To supplement these results, we used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to determine whether muscarine was present in 30 new samples of Inocybaceae, the majority of which have not been previously assayed or that originated from either the tropics or temperate regions of the southern hemisphere. Our main objective is to test the hypothesis that the presence of muscarine is a shared ancestral feature of the Inocybaceae. In addition, we also test whether species of Inocyabceae that produce psilocybin are monophyletic. Our findings suggest otherwise. Muscarine has evolved independently on several occasions, together with several losses. We also detect at least two independent transitions of muscarine-free lineages to psilocybin-producing states. Although not ancestral for the family as a whole, muscarine is a shared derived trait for an inclusive clade containing three of the seven major lineages of Inocybaceae (the Inocybe, Nothocybe, and Pseudosperma clades), the common ancestor of which may have evolved ca. 60 million years ago. Thus, muscarine represents a conserved trait followed by several recent losses. Transitions to psilocybin from muscarine-producing ancestors occurred more recently between 10-20 million years ago after muscarine loss in two separate lineages. Statistical analyses firmly reject a single origin of muscarine-producing taxa.

  13. Increased seizure latency and decreased severity of pentylenetetrazol-induced seizures in mice after essential oil administration.

    PubMed

    Koutroumanidou, Eleni; Kimbaris, Athanasios; Kortsaris, Alexandros; Bezirtzoglou, Eugenia; Polissiou, Moschos; Charalabopoulos, Konstantinos; Pagonopoulou, Olga

    2013-01-01

    The effect of pretreatment with essential oils (EOs) from eight aromatic plants on the seizure latency and severity of pentylenetetrazol- (PTZ-) induced seizures in mice was evaluated. Weight-dependent doses of Rosmarinus officinalis, Ocimum basilicum, Mentha spicata, Mentha pulegium, Lavandula angustifolia, Mentha piperita, Origanum dictamnus, and Origanum vulgare, isolated from the respective aromatic plants from NE Greece, were administered 60 minutes prior to intraperitoneal (i.p.) injection of a lethal dose of PTZ to eight respective groups of Balb-c mice. Control group received only one i.p. PTZ injection. Motor and behavioral activity of the animals after EOs administration, development of tonic-clonic seizures, seizure latency and severity, and percentage of survival after PTZ administration were determined for each group. All groups of mice treated with the EOs showed reduced activity and stability after the administration of the oil, except for those treated with O. vulgare (100% mortality after the administration of the oil). After PTZ administration, mice from the different groups showed increased latency and reduced severity of seizures (ranging from simple twitches to complete seizures). Mice who had received M. piperita demonstrated no seizures and 100% survival. The different drastic component and its concentration could account for the diversity of anticonvulsant effects. PMID:23819045

  14. Activation of T84 cell chloride channels by carbachol involves a phosphoinositide-coupled muscarinic M3 receptor.

    PubMed

    Dickinson, K E; Frizzell, R A; Sekar, M C

    1992-04-10

    Muscarinic agonists stimulate Cl- secretion across monolayers of the colon tumor epithelial cell line, T84. The muscarinic receptor has been characterized in T84 cell homogenates by radioligand binding using [3H]N-methylscopolamine ([3H]NMS). [3H]NMS bound to a single population of sites at 25 degrees C in 100 mM NaCl, 20 mM HEPES, 10 mM MgCl2, pH 7.4 buffer, with calculated Kd = 278 (+/- 44) pM and Bmax = 40 (+/- 6) fmol/mg protein (n = 4). Binding was reversible (diss. t1/2 = 18 +/- 3 min) and stereoselective (dexetimide Ki = 0.3 nM) much greater than levetimide (Ki = 8300 nM). Antagonists exhibited the following rank order of potencies and Ki values (nM): atropine (0.54) greater than 4-diphenylacetoxy-N-methylpiperidine methobromide (4-DAMP) (0.84) greater than dicyclomine (14) = hexahydrosiladifenidol (18) greater than pirenzepine (136) greater than AF-DX 116 (3610). The same sequence was observed for inhibition of carbachol-induced 125I efflux from T84 monolayers. This is indicative of an M3 'glandular' muscarinic receptor. Coupling to second messenger systems was examined by labelling monolayers with [14C]arachidonic acid (AA) or [3H]inositol. Carbachol (0.3 mM) did not release [14C]AA from labelled lipids, but ionomycin produced a dose-dependent increase in media [14C]AA. Carbachol (0.3 mM) elevated inositol monophosphate 14-fold. The results suggest that muscarinic agonists stimulate Cl- secretion by interacting with an M3 receptor coupled to inositide lipid hydrolysis. PMID:1379932

  15. Ligand binding properties of muscarinic acetylcholine receptor subtypes (m1-m5) expressed in baculovirus-infected insect cells.

    PubMed

    Dong, G Z; Kameyama, K; Rinken, A; Haga, T

    1995-07-01

    Five subtypes of muscarinic acetylcholine receptors (m1-m5) have been expressed in insect cells (Spodoptera frugiperda, Sf9) using the baculovirus system. Up to 6 nmol of muscarinic acetylcholine receptors were produced by 1 liter culture; 0.3 to 0.6 (human m1), 3 to 6 (human m2), 2 to 4 (rat m3), 1 to 2 (rat m4) and 0.5 to 1 (human m5) nmol. Pirenzepine, AF-DX116 and hexahidrosiladifenidol showed the highest affinity for the m1, m2 and m3 subtype, respectively, indicating that these receptors expressed in Sf9 cells retain the same substrate specificity as those in mammalian tissues or cultured cells. Among 32 kinds of muscarinic ligands examined in the present studies, prifinium was found to have the highest affinity for the m4 subtype, and pilocarpine, oxotremorine, McN-A343 and promethazine the highest affinity for the m5 subtype, although the differences in the affinities among the five subtypes were less than 10-fold. Alcuronium increased the binding of [3H]N-methylscopalamine to the m2 subtype, but not the m1, m4 and m5 subtypes and only slightly to the m3 subtype. Similar but smaller effects of fangchinoline and tetrandrine were found for [3H]N-methylscopalamine binding to only the m3 subtype. These effects may also be useful for the discrimination of individual subtypes. PMID:7616422

  16. Permanent alterations in muscarinic receptors and pupil size produced by chronic atropinization in kittens

    SciTech Connect

    Smith, E.L.; Redburn, D.A.; Harwerth, R.S.; Maguire, G.W.

    1984-02-01

    Chronic mydriasis was induced in six kittens (four monocular, two binocular) and two adult cats (both monocular) by the daily topical application of atropine. Both the kittens and the adult cats were atropinized for a 13-week period with the treatment regimen beginning at the time of eye opening for the kittens. Pupil size measurements, obtained 1 year after the atropinization were discontinued, revealed that, although the pupils of the adult cats were normal, the pupils of the kittens' treated eyes were consistently smaller than pupils in control eyes. The status of the muscarinic receptors in the kittens' irides was investigated using /sup 3/H-QNB binding assays. In comparison with iris muscle homogenates from the control eyes, those from the treated eyes demonstrated an eightfold increase in the number of receptor binding sites. The results indicate that pupil size can be altered permanently by chronic mydriasis initiated early in the life of a kitten and that the permanent change in pupil size may result, in part, from a type of permanent supersensitivity response in the muscle following chronic blockade of muscarinic transmission by atropine.

  17. Local heating of human skin causes hyperemia without mediation by muscarinic cholinergic receptors or prostanoids.

    PubMed

    Golay, Sandrine; Haeberli, Christian; Delachaux, Anne; Liaudet, Lucas; Kucera, Paul; Waeber, Bernard; Feihl, François

    2004-11-01

    Local changes in surface temperature have a powerful influence on the perfusion of human skin. Heating increases local skin blood flow, but the mechanisms and mediators of this response (thermal hyperemia response) are incompletely elucidated. In the present study, we examined the possible dependence of the thermal hyperemia response on stimulation of muscarinic cholinergic receptors and on production of vasodilator prostanoids. In 13 male healthy subjects aged 20-30 yr, a temperature-controlled chamber was positioned on the volar face of one forearm and used to raise surface temperature from 34 to 41 degrees C. The time course of the resulting thermal hyperemia response was recorded with a laser-Doppler imager. In one experiment, each of eight subjects received an intravenous bolus of the antimuscarinic agent glycopyrrolate (4 microg/kg) on one visit and saline on the other. The thermal hyperemia response was determined within the hour after the injections. Glycopyrrolate effectively inhibited the skin vasodilation induced by iontophoresis of acetylcholine but did not influence the thermal hyperemia response. In a second experiment, conducted in five other subjects, 1 g of the cyclooxygenase inhibitor aspirin administered orally totally abolished the vasodilation induced in the skin by anodal current but also failed to modify the thermal hyperemia response. The present study excludes the stimulation of muscarinic receptors and the production of vasodilator prostaglandins as essential and nonredundant mechanisms for the vasodilation induced by local heating in human forearm skin. PMID:15247159

  18. Expression of the rat muscarinic receptor gene m3 in Dictyostelium discoideum.

    PubMed

    Voith, G; Kramm, H; Zündorf, I; Winkler, T; Dingermann, T

    1998-10-01

    We functionally expressed the rat muscarinic m3 receptor (rm3) in the cellular slime mold Dictyostelium discoideum under the control of the homologous discoidin I gamma promoter. Cells transfected with the authentic rm3 receptor gene expressed about 100 functional receptor molecules per cell, corresponding to a Bmax for [3H]-NMS of 36 +/- 9 fmol/mg of protein in isolated membranes. Genetic fusion of the Dictyostelium contact site A (csA) leader peptide to the amino terminus of rm3 increased the receptor expression by about 17-fold. Remarkable, in [3H]-NMS ligand binding experiments performed with whole cells no characteristic saturable binding was observed and there was no significant difference in [3H]-NMS binding to whole cells of rm3 and csA/rm3 transformants. The recombinant rm3 receptor showed an about 10-fold higher affinity to the M3-selective antagonist p-F-HHSiD compared to the M2-selective antagonist AQ-RA 741, suggesting that membranes derived from transgenic D. discoideum cells may be useful for the search of new subtype-specific muscarinic receptor ligands. PMID:9812338

  19. Short-Term Desensitization of Muscarinic K+ Current in the Heart

    PubMed Central

    Murakami, Shingo; Inanobe, Atsushi; Kurachi, Yoshihisa

    2013-01-01

    Acetylcholine (ACh) rapidly increases cardiac K+ currents (IKACh) by activating muscarinic K+ (KACh) channels followed by a gradual amplitude decrease within seconds. This phenomenon is called short-term desensitization and its precise mechanism and physiological role are still unclear. We constructed a mathematical model for IKACh to examine the conditions required to reconstitute short-term desensitization. Two conditions were crucial: two distinct muscarinic receptors (m2Rs) with different affinities for ACh, which conferred an IKACh response over a wide range of ACh concentrations, and two distinct KACh channels with different affinities for the G-protein βγ subunits, which contributed to reconstitution of the temporal behavior of IKACh. Under these conditions, the model quantitatively reproduced several unique properties of short-term desensitization observed in myocytes: 1), the peak and quasi-steady states with 0.01–100 μM [ACh]; 2), effects of ACh preperfusion; and 3), recovery from short-term desensitization. In the presence of 10 μM ACh, the IKACh model conferred recurring spontaneous firing after asystole of 8.9 s and 10.7 s for the Demir and Kurata sinoatrial node models, respectively. Therefore, two different populations of KACh channels and m2Rs may participate in short-term desensitization of IKACh in native myocytes, and may be responsible for vagal escape at nodal cells. PMID:24048003

  20. The Mine Safety and Health Administration's criterion threshold value policy increases miners' risk of pneumoconiosis

    SciTech Connect

    Weeks, J.L.

    2006-06-15

    Background The Mine Safety and Health Administration (MSHA) proposes to issue citations for non-compliance with the exposure limit for respirable coal mine dust when measured exposure exceeds the exposure limit with a 'high degree of confidence.' This criterion threshold value (CTV) is derived from the sampling and analytical error of the measurement method. This policy is based on a combination of statistical and legal reasoning: the one-tailed 95% confidence limit of the sampling method, the apparent principle of due process and a standard of proof analogous to 'beyond a reasonable doubt.' This policy raises the effective exposure limit, it is contrary to the precautionary principle, it is not a fair sharing of the burden of uncertainty, and it employs an inappropriate standard of proof. Its own advisory committee and NIOSH have advised against this policy. For longwall mining sections, it results in a failure to issue citations for approximately 36% of the measured values that exceed the statutory exposure limit. Citations for non-compliance with the respirable dust standard should be issued for any measure exposure that exceeds the exposure limit.

  1. Dietary resveratrol administration increases MnSOD expression and activity in mouse brain

    SciTech Connect

    Robb, Ellen L.; Winkelmolen, Lieke; Visanji, Naomi; Brotchie, Jonathan; Stuart, Jeffrey A.

    2008-07-18

    trans-Resveratrol (3,4',5-trihydroxystilbene; RES) is of interest for its reported protective effects in a variety of pathologies, including neurodegeneration. Many of these protective properties have been attributed to the ability of RES to reduce oxidative stress. In vitro studies have shown an increase in antioxidant enzyme activities following exposure to RES, including upregulation of mitochondrial superoxide dismutase, an enzyme that is capable of reducing both oxidative stress and cell death. We sought to determine if a similar increase in endogenous antioxidant enzymes is observed with RES treatment in vivo. Three separate modes of RES delivery were utilized; in a standard diet, a high fat diet and through a subcutaneous osmotic minipump. RES given in a high fat diet proved to be effective in elevating antioxidant capacity in brain resulting in an increase in both MnSOD protein level (140%) and activity (75%). The increase in MnSOD was not due to a substantial proliferation of mitochondria, as RES treatment induced a 10% increase in mitochondrial abundance (Citrate Synthase activity). The potential neuroprotective properties of MnSOD have been well established, and we demonstrate that a dietary delivery of RES is able to increase the expression and activity of this enzyme in vivo.

  2. Stimulation of brain muscarinic acetylcholine receptors acutely reverses radiogenic hypodipsia

    SciTech Connect

    Mickley, G.A.; Stevens, K.E.

    1986-03-01

    A sufficiently large dose of ionizing radiation produces changes in water consumption. However, the direction, durations, and physiological substrates of these alterations remain in question. Here we report a 5-d hypodipsia in rats exposed to 600 rads /sup 60/Co but a more transient, albeit larger, reduction in drinking after 1000 /sup 60/Co. Brain cholinergic neurons have been implicated as mediators of thirst. Therefore, we explored the role of hypothalamic muscarinic receptors in the production of radiation-induced hypodipsia. This was accomplished through the intrahypothalamic injection of carbachol (a muscarinic agonist) or atropine (a muscarinic antagonist) in irradiated rats. Intracranial carbachol produced acute reversal of radiogenic hypodipsia while atropine potentiated the hypodipsia. These post-irradiation drug-induced behaviors were similar to those observed after the same drug treatments before irradiation. Since cholinergic neuronal functions persist and are labile (can be pharmacologically stimulated and blocked) after irradiation, this suggests that other neuronal systems and/or neurochemicals may be more prominently involved in radiogenic hypodipsia.

  3. Purification of muscarinic acetylcholine receptors by affinity chromatography.

    PubMed Central

    André, C; De Backer, J P; Guillet, J C; Vanderheyden, P; Vauquelin, G; Strosberg, A D

    1983-01-01

    Calf forebrain homogenates contain 2.8 pM muscarinic acetylcholine receptors per mg of protein. [3H]Antagonist saturation binding experiments under equilibrium conditions revealed a single class of sites with equilibrium dissociation constants of 0.82 nM for [3H]dexetimide and 0.095 nM for [3H]quinuclidinyl benzilate. Displacement binding studies with agonists revealed the presence of low and high affinity sites. Here we describe the solubilization of muscarinic acetylcholine receptors with digitonin and their purification by affinity chromatography using an affinity gel which consisted of dexetimide coupled to Affi-Gel 10 (i.e., carboxy N-hydroxysuccinimide esters linked via a 1 nm spacer arm to agarose beads). Purified proteins were obtained by specific elution with muscarinic drugs, i.e., the antagonist atropine and the irreversible ligand propylbenzilylcholine mustard. SDS-polyacrylamide gel electrophoresis of the radioiodinated purified preparations revealed a major 70-K protein. Images Fig. 3. PMID:6605245

  4. Acute administration of diosgenin or dioscorea improves hyperglycemia with increases muscular steroidogenesis in STZ-induced type 1 diabetic rats.

    PubMed

    Sato, K; Fujita, S; Iemitsu, M

    2014-09-01

    Acute dehydroepiandrosterone (DHEA) administration improves hyperglycemia in rats with streptozotocin (STZ)-induced type 1 diabetes mellitus. Diosgenin, a steroid structurally similar to DHEA (dehydroepiandrosterone), is contained highly levels in dioscorea; however, it is still unclear whether this natural product improves hyperglycemia in the type 1 diabetes model rats through an increase muscular GLUT4 signaling. After 1 week of STZ injection, fasting glucose level was measured in blood taken from the tail vein every 30 min for 150 min after injection of diosgenin or dioscorea (3mg/kg). On another day, muscle was resected 150 min after diosgenin or dioscorea injections. Serum DHEA level increased significantly 120 min after diosgenin or dioscorea injections; concomitantly, blood glucose level decreased significantly. Moreover, GLUT4 translocation, as well as phosphorylation of Akt and PKC ζ/λ, increased significantly by diosgenin or dioscorea administration. However, these effects of diosgenin and dioscorea were blocked by a 5α-reductase inhibitor that inhibits synthesizing dehydrotestosterone (DHT) from testosterone. Additionally, significant correlations were observed between blood glucose level, GLUT4 translocation level, and muscular sex steroid hormone level 150 min after the administrations. These results suggest that the diosgenin-induced increase in the DHEA level may contribute to the improvement of hyperglycemia by activating the muscular GLUT4 signaling pathway in type 1 diabetes model rats.

  5. Increase in liver nuclear tri-iodothyronine receptors associated with increased deoxyribonucleic acid by long-term administration of tri-iodothyronine in thyroidectomized rats

    PubMed Central

    Nakamura, Hirotoshi; Hamada, Satoshi; Imura, Hiroo

    1979-01-01

    The effect of long-term administration of small amounts of tri-iodothyronine was examined on the nuclear tri-iodothyronine receptors in rat liver. The maximal binding capacity (Cmax.) and association constant (Ka) of the receptors were determined in thyroidectomized rats given vehicle alone (group A), 2μg of tri-iodothyronine/100g body wt. (group B) or 7μg of tri-iodothyronine/100g body wt. (group C) for 2 weeks. Scatchard analyses with correction for the amount of endogenous tri-iodothyronine revealed that Cmax. values per g of liver were increased to 1.5 and 2.7 times the control value in groups B and C respectively. Since concentrations of DNA per g of liver were significantly increased in the two groups of hormone-treated rats, Cmax. values per mg of DNA were nearly the same in group B, but still increased significantly in group C compared with group A. Ka values remained unchanged in all three groups of animals. Mitochondrial α-glycerophosphate dehydrogenase activity was 9.6 and 28.7 times as high in groups B and C, respectively, as in group A. Concentrations of endogenous tri-iodothyronine bound to non-histone protein were substantially increased in groups B and C, although concentrations of serum tri-iodothyronine remained rather low. The results obtained indicate that the long-term administration of tri-iodothyronine in small doses induces an increase in the nuclear receptors associated with increased DNA with and without accompanying a relative increase in the receptor concentration in thyroidectomized rats. Also the hormonal effect is closely related to the total number of the nuclear receptors and the concentrations of nuclear tri-iodothyronine bound to the receptors rather than the serum tri-iodothyronine concentrations. PMID:230825

  6. Transient increase in obese gene expression after food intake or insulin administration.

    PubMed

    Saladin, R; De Vos, P; Guerre-Millo, M; Leturque, A; Girard, J; Staels, B; Auwerx, J

    1995-10-12

    Obesity is a disorder of energy balance, indicating a chronic disequilibrium between energy intake and expenditure. Recently, the mouse ob gene, and subsequently its human and rat homologues, have been cloned. The ob gene product, leptin, is expressed exclusively in adipose tissue, and appears to be a signalling factor regulating body-weight homeostasis and energy balance. Because the level of ob gene expression might indicate the size of the adipose depot, we suggest that it is regulated by factors modulating adipose tissue size. Here we show that ob gene exhibits diurnal variation, increasing during the night, after rats start eating. This variation was linked to changes in food intake, as fasting prevented the cyclic variation and decreased ob messenger RNA. Furthermore, refeeding fasted rats restored ob mRNA within 4 hours to levels of fed animals. A single insulin injection in fasted animals increased ob mRNA to levels of fed controls. Experiments to control glucose and insulin independently in animals, and studies in primary adipocytes, showed that insulin regulates ob gene expression directly in rats, regardless of its glucose-lowering effects. Whereas the ob gene product, leptin, has been shown to reduce food intake and increase energy expenditure, our data demonstrate that ob gene expression is increased after food ingestion in rats, perhaps through a direct action of insulin on the adipocyte.

  7. Roux-en-Y gastric bypass increases intravenous ethanol self-administration in dietary obese rats.

    PubMed

    Polston, James E; Pritchett, Carolyn E; Tomasko, Jonathan M; Rogers, Ann M; Leggio, Lorenzo; Thanos, Panayotis K; Volkow, Nora D; Hajnal, Andras

    2013-01-01

    Roux-en-Y gastric bypass surgery (RYGB) is an effective treatment for severe obesity. Clinical studies however have reported susceptibility to increased alcohol use after RYGB, and preclinical studies have shown increased alcohol intake in obese rats after RYGB. This could reflect a direct enhancement of alcohol's rewarding effects in the brain or an indirect effect due to increased alcohol absorption after RGYB. To rule out the contribution that changes in alcohol absorption have on its rewarding effects, here we assessed the effects of RYGB on intravenously (IV) administered ethanol (1%). For this purpose, high fat (60% kcal from fat) diet-induced obese male Sprague Dawley rats were tested ~2 months after RYGB or sham surgery (SHAM) using both fixed and progressive ratio schedules of reinforcement to evaluate if RGYB modified the reinforcing effects of IV ethanol. Compared to SHAM, RYGB rats made significantly more active spout responses to earn IV ethanol during the fixed ratio schedule, and achieved higher breakpoints during the progressive ratio schedule. Although additional studies are needed, our results provide preliminary evidence that RYGB increases the rewarding effects of alcohol independent of its effects on alcohol absorption.

  8. Gabapentin potentiates sensitivity to the interoceptive effects of alcohol and increases alcohol self-administration in rats.

    PubMed

    Besheer, Joyce; Frisbee, Suzanne; Randall, Patrick A; Jaramillo, Anel A; Masciello, Maria

    2016-02-01

    Gabapentin, a drug used in the treatment of epileptic seizures and neuropathic pain, has shown efficacy in the treatment of alcohol dependence. Moreover, given that gabapentin is used in the general population (e.g., non-dependent individuals, social drinkers), we sought to utilize preclinical assessments to examine the effects of gabapentin on sensitivity to moderate alcohol doses and alcohol self-administration in rats with a history of moderate drinking. To this end, we assessed whether gabapentin (0, 10, 30, 120 mg/kg, IG) pretreatment alters sensitivity to experimenter- and self-administered alcohol, and whether gabapentin alone has alcohol-like discriminative stimulus effects in rats trained to discriminate alcohol dose (1 g/kg, IG) vs. water. Second, we assessed whether gabapentin (0, 10, 30, 60 mg/kg, IG) would alter alcohol self-administration. Gabapentin pretreatment potentiated the interoceptive effects of both experimenter-administered and self-administered alcohol in discrimination-trained rats. Additionally, the highest gabapentin doses tested (30 and 120 mg/kg) were found to have partial alcohol-like discriminative stimulus effects when administered alone (e.g., without alcohol). In the self-administration trained rats, gabapentin pretreatment (60 mg/kg) resulted in an escalation in alcohol self-administration. Given the importance of interoceptive drug cues in priming and maintaining self-administration, these data define a specific behavioral mechanism (i.e., potentiation of alcohol effects) by which gabapentin may increase alcohol self-administration in non-dependent populations.

  9. Chronic administration of caderofloxacin, a new fluoroquinolone, increases hepatic CYP2E1 expression and activity in rats

    PubMed Central

    Liu, Li; Miao, Ming-xing; Zhong, Ze-yu; Xu, Ping; Chen, Yang; Liu, Xiao-dong

    2016-01-01

    Aim: Caderofloxacin is a new fluoroquinolone that is under phase III clinical trials in China. Here we examined the effects of caderofloxacin on rat hepatic cytochrome P450 (CYP450) isoforms as well as the potential of caderofloxacin interacting with co-administered drugs. Methods: Male rats were treated with caderofloxacin (9 mg/kg, ig) once or twice daily for 14 consecutive days. The effects of caderofloxacin on CYP3A, 2D6, 2C19, 1A2, 2E1 and 2C9 were evaluated using a “cocktail” of 6 probes (midazolam, dextromethorphan, omeprazole, theophylline, chlorzoxazone and diclofenac) injected on d 0 (prior to caderofloxacin exposure) and d 15 (after caderofloxacin exposure). Hepatic microsomes from the caderofloxacin-treated rats were used to assess CYP2E1 activity and chlorzoxazone metabolism. The expression of CYP2E1 mRNA and protein in hepatic microsomes was analyzed with RT-PCR and Western blotting, respectively. Results: Fourteen-day administration of caderofloxacin significantly increased the activity of hepatic CYP2E1, leading to enhanced metabolism of chlorzoxazone. In vitro microsomal study confirmed that CYP2E1 was a major metabolic enzyme involved in chlorzoxazone metabolism, and the 14-d administration of caderofloxacin significantly increased the activity of CYP2E1 in hepatic microsomes, resulting in increased formation of 6-hydroxychlorzoxazone. Furthermore, the 14-d administration of caderofloxacin significantly increased the expression of CYP2E1 mRNA and protein in liver microsomes, which was consistent with the pharmacokinetic results. Conclusion: Fourteen-day administration of caderofloxacin can induce the expression and activity of hepatic CYP2E1 in rats. When caderofloxacin is administered, a potential drug-drug interaction mediated by CYP2E1 induction should be considered. PMID:26838075

  10. Acute and chronic caffeine administration increases physical activity in sedentary adults.

    PubMed

    Schrader, Patrick; Panek, Leah M; Temple, Jennifer L

    2013-06-01

    Caffeine is a commonly used stimulant thought to have ergogenic properties. Most studies on the ergogenic effects of caffeine have been conducted in athletes. The purpose of this study was to test the hypothesis that caffeine reduces ratings of perceived exertion and increases liking of physical activity in sedentary adults. Participants completed treadmill walking at 60% to 70% of their maximal heart rate at baseline and for 6 subsequent visits, during which half of the participants were given caffeine (3 mg/kg) and half given placebo in a sports drink vehicle. To investigate the potential synergistic effects of acute and chronic caffeine on self-determined exercise duration, participants were rerandomized to either the same or different condition for the last visit, creating 4 chronic/acute treatment groups (placebo/placebo, placebo/caffeine, caffeine/placebo, caffeine/caffeine). Participants rated how much they liked the activity and perceived exertion at each visit. There was a main effect of time on liking of physical activity, with liking increasing over time and an interaction of sex and caffeine treatment on liking, with liking of activity increasing in female participants treated with caffeine, but not with placebo. There was no effect of caffeine on ratings of perceived exertion. Individuals who received caffeine on the final test day exercised for significantly longer than those who received placebo. These data suggest that repeated exposure to physical activity significantly increases liking of exercise and reduces ratings of perceived exertion and that caffeine does little to further modify these effects.

  11. Oral administration of lithium increases tissue magnesium contents but not plasma magnesium level in rats.

    PubMed

    Kiełczykowska, Małgorzata; Musik, Irena; Hordyjewska, Anna; Boguszewska, Anna; Lewandowska, Anna; Pasternak, Kazimierz

    2007-01-01

    The aim of this work was to determine the influence of different doses of lithium on magnesium concentration in plasma and tissues of rats. For a period of eight weeks rats had been provided with aqueous solutions of Li(2)CO(3) whose concentrations were established as follows: 0.7; 1.4; 2.6; 3.6; 7.1; 10.7 mmol Li(+)/l. Magnesium concentration was determined in plasma and tissue supernatants. Lithium caused no changes in magnesium concentration in plasma, whereas Mg concentration in tissues was found to be enhanced, although the degree of the increment depended on the studied tissue. In the liver, brain and heart muscle, the increase was statistically insignificant vs. control. In the kidney, the higher Li doses were required to result in the significant Mg enhancement, whereas in femoral muscle all the used doses caused well-marked Mg increase vs. control. Positive correlations between average daily Li intake and tissue Mg concentration in the kidney (r = 0.650) and femoral muscle (r = 0.696) were found. In conclusion, the present study indicates that the different Li doses disturbed tissue homeostasis of magnesium. The increase in Mg tissue concentration, observed in groups receiving higher Li doses can influence nervous-muscular excitability.

  12. Sugar administration to newly emerged Aedes albopictus males increases their survival probability and mating performance.

    PubMed

    Bellini, Romeo; Puggioli, Arianna; Balestrino, Fabrizio; Brunelli, Paolo; Medici, Anna; Urbanelli, Sandra; Carrieri, Marco

    2014-04-01

    Aedes albopictus male survival in laboratory cages is no more than 4-5 days when kept without any access to sugar indicating their need to feed on a sugar source soon after emergence. We therefore developed a device to administer energetic substances to newly emerged males when released as pupae as part of a sterile insect technique (SIT) programme, made with a polyurethane sponge 4 cm thick and perforated with holes 2 cm in diameter. The sponge was imbibed with the required sugar solution and due to its high retention capacity the sugar solution was available for males to feed for at least 48 h. When evaluated in lab cages, comparing adults emerged from the device with sugar solution vs the device with water only (as negative control), about half of the males tested positive for fructose using the Van Handel anthrone test, compared to none of males in the control cage. We then tested the tool in semi-field and in field conditions with different sugar concentrations (10%, 15%, and 20%) and compared results to the controls fed with water only. Males were recaptured by a battery operated manual aspirator at 24 and 48 h after pupae release. Rather high share 10-25% of captured males tested positive for fructose in recollections in the vicinity of the control stations, while in the vicinity of the sugar stations around 40-55% of males were positive, though variability between replicates was large. The sugar positive males in the control test may have been released males that had access to natural sugar sources found close to the release station and/or wild males present in the environment. Only a slight increase in the proportion of positive males was obtained by increasing the sugar concentration in the feeding device from 10% to 20%. Surprisingly, modification of the device to add a black plastic inverted funnel above the container reduced rather than increased the proportion of fructose positive males collected around the station. No evidence of difference in the

  13. Sugar administration to newly emerged Aedes albopictus males increases their survival probability and mating performance.

    PubMed

    Bellini, Romeo; Puggioli, Arianna; Balestrino, Fabrizio; Brunelli, Paolo; Medici, Anna; Urbanelli, Sandra; Carrieri, Marco

    2014-04-01

    Aedes albopictus male survival in laboratory cages is no more than 4-5 days when kept without any access to sugar indicating their need to feed on a sugar source soon after emergence. We therefore developed a device to administer energetic substances to newly emerged males when released as pupae as part of a sterile insect technique (SIT) programme, made with a polyurethane sponge 4 cm thick and perforated with holes 2 cm in diameter. The sponge was imbibed with the required sugar solution and due to its high retention capacity the sugar solution was available for males to feed for at least 48 h. When evaluated in lab cages, comparing adults emerged from the device with sugar solution vs the device with water only (as negative control), about half of the males tested positive for fructose using the Van Handel anthrone test, compared to none of males in the control cage. We then tested the tool in semi-field and in field conditions with different sugar concentrations (10%, 15%, and 20%) and compared results to the controls fed with water only. Males were recaptured by a battery operated manual aspirator at 24 and 48 h after pupae release. Rather high share 10-25% of captured males tested positive for fructose in recollections in the vicinity of the control stations, while in the vicinity of the sugar stations around 40-55% of males were positive, though variability between replicates was large. The sugar positive males in the control test may have been released males that had access to natural sugar sources found close to the release station and/or wild males present in the environment. Only a slight increase in the proportion of positive males was obtained by increasing the sugar concentration in the feeding device from 10% to 20%. Surprisingly, modification of the device to add a black plastic inverted funnel above the container reduced rather than increased the proportion of fructose positive males collected around the station. No evidence of difference in the

  14. Increased 5-HT Levels Following Repeated Administration of Nigella sativa L. (Black Seed) Oil Produce Antidepressant Effects in Rats.

    PubMed

    Perveen, Tahira; Haider, Saida; Zuberi, Nudrat Anwar; Saleem, Sadia; Sadaf, Sana; Batool, Zehra

    2014-03-01

    The seeds of Nigella sativa L., commonly known as black seed or black cumin, and its extracts are used in folk medicine in the Middle East and in Asian countries for the promotion of good health and as a remedy for many ailments. These seeds have many acclaimed medicinal properties such as broncho-dilatory, immunopotentiating, analgesic, anti-inflammatory, and hypotensive. In the present study, the antidepressant activity following the repeated administration of Nigella sativa L. oil has been monitored using the forced swim test. Rats treated with Nigella sativa L. oil exhibited a significant increase in struggling time after oral administration of Nigella sativa L. oil (0.1 ml/kg/day) for four weeks. Nigella sativa L. oil increased brain 5-HT levels and decreased 5-HT turnover (5-HT/5-HIAA ratio). Levels of tryptophan increased significantly in the brain and plasma following the repeated administration of Nigella sativa L. oil. Nigella sativa L. oil showed a potential antidepressant-like effect.

  15. Long-term nitric oxide deficiency causes muscarinic supersensitivity and reduces β3-adrenoceptor-mediated relaxation, causing rat detrusor overactivity

    PubMed Central

    Mónica, F Z T; Bricola, A A O; Báu, F R; Freitas, L L Lopes; Teixeira, S A; Muscará, M N; Abdalla, F M F; Porto, C S; De Nucci, G; Zanesco, A; Antunes, E

    2008-01-01

    Background and purpose: Overactive bladder is a complex and widely prevalent condition, but little is known about its physiopathology. We have carried out morphological, biochemical and functional assays to investigate the effects of long-term nitric oxide (NO) deficiency on muscarinic receptor and β-adrenoceptor modulation leading to overactivity of rat detrusor muscle. Experimental approach: Male Wistar rats received Nω-nitro-L-arginine methyl ester (L-NAME) in drinking water for 7–30 days. Functional responses to muscarinic and β-adrenoceptor agonists were measured in detrusor smooth muscle (DSM) strips in Krebs–Henseleit solution. Measurements of [3H]inositol phosphate, NO synthase (NOS) activity, [3H]quinuclidinyl benzilate ([3H]QNB) binding and bladder morphology were also performed. Key results: Long-term L-NAME treatment significantly increased carbachol-induced DSM contractile responses after 15 and 30 days; relaxing responses to the β3-adrenoceptor agonist BRL 37-344 were significantly reduced at 30 days. Constitutive NOS activity in bladder was reduced by 86% after 7 days and maintained up to 30 days of L-NAME treatment. Carbachol increased sixfold the [3H]inositol phosphate in bladder tissue from rats treated with L-NAME. [3H]QNB was bound with an apparent KD twofold higher in bladder membranes after L-NAME treatment compared with that in control. No morphological alterations in DSM were found. Conclusions and implications: Long-term NO deficiency increased rat DSM contractile responses to a muscarinic agonist, accompanied by significantly enhanced KD values for muscarinic receptors and [3H]inositol phosphate accumulation in bladder. This supersensitivity for muscarinic agonists along with reductions of β3-adrenoceptor-mediated relaxations indicated that overactive DSM resulted from chronic NO deficiency. PMID:18297104

  16. Cortisol administration increases hippocampal activation to infant crying in males depending on childhood neglect.

    PubMed

    Bos, Peter A; Montoya, Estrella R; Terburg, David; van Honk, Jack

    2014-10-01

    Animal studies show that exposure to parental neglect alters stress regulation and can lead to neural hyposensitivity or hypersensitivity in response to cortisol, most pronounced in the hippocampus. Cortisol, the end product of the hypothalamic-pituitary-adrenal (HPA) axis, has also been related to parenting more directly, for example, in both sexes, cortisol levels increase when listening to infants crying, possibly to activate and facilitate effective care behavior. Severe trauma is known to negatively affect the HPA-axis in humans; however, it is unknown whether normal variation in parental care in the healthy population can alter sensitivity of the hippocampus to cortisol. Here, we investigate whether variation in experienced neglect changes neural sensitivity to cortisol when humans listen to infant crying, which is an unequivocal signal relevant for care behavior. In a placebo-controlled, within-subject neuroimaging study, we administered 40 mg cortisol to 21 healthy young males without children and used a validated task for measuring neural responses to infant crying. The Dutch version of the Childhood Trauma Questionnaire was used to index participants' early exposure to abuse and neglect. The data show that cortisol markedly increased hippocampal activation toward crying infants, and this effect varied significantly with parental neglect, even in our nonclinical subject sample. Without exposure to severe trauma or neglect, reduced self-experienced quality of parental care in the normal range already substantially increased hippocampal responsivity to cortisol. Altered hippocampal sensitivity to cortisol might be a cross-species marker for the risk of developing later life psychopathology. PMID:24757127

  17. Cortisol administration increases hippocampal activation to infant crying in males depending on childhood neglect.

    PubMed

    Bos, Peter A; Montoya, Estrella R; Terburg, David; van Honk, Jack

    2014-10-01

    Animal studies show that exposure to parental neglect alters stress regulation and can lead to neural hyposensitivity or hypersensitivity in response to cortisol, most pronounced in the hippocampus. Cortisol, the end product of the hypothalamic-pituitary-adrenal (HPA) axis, has also been related to parenting more directly, for example, in both sexes, cortisol levels increase when listening to infants crying, possibly to activate and facilitate effective care behavior. Severe trauma is known to negatively affect the HPA-axis in humans; however, it is unknown whether normal variation in parental care in the healthy population can alter sensitivity of the hippocampus to cortisol. Here, we investigate whether variation in experienced neglect changes neural sensitivity to cortisol when humans listen to infant crying, which is an unequivocal signal relevant for care behavior. In a placebo-controlled, within-subject neuroimaging study, we administered 40 mg cortisol to 21 healthy young males without children and used a validated task for measuring neural responses to infant crying. The Dutch version of the Childhood Trauma Questionnaire was used to index participants' early exposure to abuse and neglect. The data show that cortisol markedly increased hippocampal activation toward crying infants, and this effect varied significantly with parental neglect, even in our nonclinical subject sample. Without exposure to severe trauma or neglect, reduced self-experienced quality of parental care in the normal range already substantially increased hippocampal responsivity to cortisol. Altered hippocampal sensitivity to cortisol might be a cross-species marker for the risk of developing later life psychopathology.

  18. Corticosteroid administration modifies ozone-induced increases in sheep airway blood flow

    SciTech Connect

    Gunther, R.A.; Yousef, M.A.; Schelegle, E.S.; Cross, C.E. )

    1992-09-01

    Recently, we have shown that exposure of intubated conscious sheep to 3 to 4 ppm ozone (O3) for 3 h increases bronchial blood flow (Qbr). The purpose of the present study was to assess the potential role of corticosteroids in modulating this increase. Six nasally intubated sheep were exposed to filtered room air, 3.5 ppm O3 on two separate occasions, and 3.5 ppm O3 plus methyl-prednisone, for 3 h. Qbr was measured using a chronically implanted 20 MHz pulsed Doppler flow probe. Qbr, mean aortic pressure, cardiac output, pulmonary artery pressure, arterial blood gases, and core temperature were monitored. After 3 h of 3.5 ppm O3, Qbr increased from 3.2 +/- 0.5 (mean +/- SEM) to 8.5 +/- 1.6 KHz, whereas bronchial vascular resistance (BVR) decreased from the baseline value of 43.6 +/- 8.0 to 15.0 +/- 3 mm Hg/KHz. With corticosteroids, baseline Qbr was 3.2 +/- 0.6 and BVR was 44.2 +/- 9.7; after 3 h of 3.5 ppm O3, Qbr was 3.3 +/- 0.5 KHz and BVR was 39.0 +/- 8.0 mm Hg/KHz. The two 3.5-ppm O3 exposures without corticosteroids were impressively reproducible. Except for Qbr and BVR, no other measured cardiovascular parameters were affected by O3. The results indicate that corticosteroids are capable of interfering with mediator, neurohumoral, or inflammatory cell mechanisms responsible for vasodilation of the airway microcirculation after O3 exposure, but do not specifically address the specific processes whereby this attenuation occurs.

  19. The Cu-Zn superoxide dismutase (SOD1) inhibits ERK phosphorylation by muscarinic receptor modulation in rat pituitary GH3 cells

    SciTech Connect

    Secondo, Agnese; De Mizio, Mariarosaria; Zirpoli, Laura; Santillo, Mariarosaria; Mondola, Paolo

    2008-11-07

    The Cu-Zn superoxide dismutase (SOD1) belongs to a family of isoenzymes that are able to dismutate the oxygen superoxide in hydrogen peroxide and molecular oxygen. This enzyme is secreted by many cellular lines and it is also released trough a calcium-dependent depolarization mechanism involving SNARE protein SNAP 25. Using rat pituitary GH3 cells that express muscarinic receptors we found that SOD1 inhibits P-ERK1/2 pathway trough an interaction with muscarinic M1 receptor. This effect is strengthened by oxotremorine, a muscarinic M agonist and partially reverted by pyrenzepine, an antagonist of M1 receptor; moreover this effect is independent from increased intracellular calcium concentration induced by SOD1. Finally, P-ERK1/2 inhibition was accompanied by the reduction of GH3 cell proliferation. These data indicate that SOD1 beside the well studied antioxidant properties can be considered as a neuromodulator able to affect mitogen-activated protein kinase in rat pituitary cells trough a M1 muscarinic receptor.

  20. AKAP79, PKC, PKA and PDE4 participate in a Gq-linked muscarinic receptor and adenylate cyclase 2 cAMP signalling complex

    PubMed Central

    Shen, Jia X.; Cooper, Dermot M. F.

    2014-01-01

    AC2 (adenylate cyclase 2) is stimulated by activation of Gq-coupled muscarinic receptors through PKC (protein kinase C) to generate localized cAMP in HEK (human embryonic kidney)-293 cells. In the present study, we utilized a sensitive live-cell imaging technique to unravel the proteins that play essential roles in a Gq-coupled muscarinic receptor-mediated cAMP signalling complex. We reveal that, upon agonist binding to the Gq-coupled muscarinic receptor, AKAP79 (A-kinase-anchoring protein 79) recruits PKC to activate AC2 to produce cAMP. The cAMP formed is degraded by PDE4 (phosphodiesterase 4) activated by an AKAP-anchored PKA (protein kinase A). Calcineurin, a phosphatase bound to AKAP79, is not involved in this regulation. Overall, a transient cAMP increase is generated from AC2 by Gq-coupled muscarinic receptor activation, subject to sophisticated regulation through AKAP79, PKC, PDE4 and PKA, which significantly enhances acetylcholine-mediated signalling. PMID:23889134

  1. [Iron substitution in outpatients in Switzerland: Increase of costs associated with intravenous administration].

    PubMed

    Giger, Max; Achermann, Rita

    2013-01-01

    Iron anaemia and iron-deficient erythropoiesis are treated with oral iron supplements. For chronic haemodialysis or in the case of therapy failure or intolerance to oral iron therapy, intravenous supplements are administered. The costs of iron supplements borne by statutory health care insurance had strongly increased during the observation period from 2006 to 2010. Based on the invoice data of a large health insurance company with a market share of around 18 %, prescription data of iron preparations and laboratory tests were analysed and extrapolated to the Swiss population. During the 5-year observation period, costs of intravenous iron substitution increased by 16.5 m EUR (340.3 %) and the number of individuals treated by 243.5 %. A sharp rise was observed in women of menstruating age, which was mainly due to prescriptions issued by primary care physicians. More than 8 % of intravenous iron substitutions were administered without prior laboratory analysis,and must therefore be regarded as off-label use. A cost-benefit analysis is needed to demonstrate the additional value of intravenous over oral iron supplementation, and intravenous iron supplementation should be administered only to patients with proven iron deficiency.

  2. [Iron substitution in outpatients in Switzerland: Increase of costs associated with intravenous administration].

    PubMed

    Giger, Max; Achermann, Rita

    2013-01-01

    Iron anaemia and iron-deficient erythropoiesis are treated with oral iron supplements. For chronic haemodialysis or in the case of therapy failure or intolerance to oral iron therapy, intravenous supplements are administered. The costs of iron supplements borne by statutory health care insurance had strongly increased during the observation period from 2006 to 2010. Based on the invoice data of a large health insurance company with a market share of around 18 %, prescription data of iron preparations and laboratory tests were analysed and extrapolated to the Swiss population. During the 5-year observation period, costs of intravenous iron substitution increased by 16.5 m EUR (340.3 %) and the number of individuals treated by 243.5 %. A sharp rise was observed in women of menstruating age, which was mainly due to prescriptions issued by primary care physicians. More than 8 % of intravenous iron substitutions were administered without prior laboratory analysis,and must therefore be regarded as off-label use. A cost-benefit analysis is needed to demonstrate the additional value of intravenous over oral iron supplementation, and intravenous iron supplementation should be administered only to patients with proven iron deficiency. PMID:23916272

  3. Exercise increases insulin signaling in the hippocampus: physiological effects and pharmacological impact of intracerebroventricular insulin administration in mice.

    PubMed

    Muller, Alexandre P; Gnoatto, Jussânia; Moreira, Julia D; Zimmer, Eduardo R; Haas, Clarissa B; Lulhier, Francisco; Perry, Marcos L S; Souza, Diogo O; Torres-Aleman, Ignácio; Portela, Luis V

    2011-10-01

    Increasing evidence indicates that physical exercise induces adaptations at the cellular, molecular, and systemic levels that positively affect the brain. Insulin plays important functional roles within the brain that are mediated by insulin-receptor (IR) signaling. In the hippocampus, insulin improves synaptic plasticity, memory formation, and learning via direct modulation of GABAergic and glutamatergic receptors. Separately, physical exercise and central insulin administration exert relevant roles in cognitive function. We here use CF1 mice to investigate (i) the effects of voluntary exercise on hippocampal insulin signaling and memory performance and (ii) whether central insulin administration alters the effects of exercise on hippocampal insulin signaling and memory performance. Adult mice performed 30 days of voluntary exercise on running wheel and afterward both, sedentary and exercised groups, received intracerebroventricular (icv) injection of saline or insulin (0.5-5 mU). Memory performance was assessed using the inhibitory avoidance and water maze tasks. Hippocampal tissue was measured for [U-(14)C] glucose oxidation and the immunocontent of insulin receptor/signaling (IR, pTyr, pAktser473). Additionally, the phosphorylation of the glutamate NMDA receptor NR2B subunit and the capacity of glutamate uptake were measured, and immunohistochemistry was used to determine glial reactivity. Exercise significantly increased insulin peripheral sensitivity, spatial learning, and hippocampal IR/pTyrIR/pAktser473 immunocontent. Glucose oxidation, glutamate uptake, and astrocyte number also increased relative to the sedentary group. In both memory tasks, 5 mU icv insulin produced amnesia but only in exercised animals. This amnesia was associated a rapid (15 min) and persistent (24 h) increase in hippocampal pNR2B immunocontent that paralleled the increase in glial reactivity. In conclusion, physical exercise thus increased hippocampal insulin signaling and improved

  4. Inflammatory Pain Promotes Increased Opioid Self-Administration: Role of Dysregulated Ventral Tegmental Area μ Opioid Receptors

    PubMed Central

    Hipólito, Lucia; Wilson-Poe, Adrianne; Campos-Jurado, Yolanda; Zhong, Elaine; Gonzalez-Romero, Jose; Virag, Laszlo; Whittington, Robert; Comer, Sandra D.; Carlton, Susan M.; Walker, Brendan M.; Bruchas, Michael R.

    2015-01-01

    Pain management in opioid abusers engenders ethical and practical difficulties for clinicians, often resulting in pain mismanagement. Although chronic opioid administration may alter pain states, the presence of pain itself may alter the propensity to self-administer opioids, and previous history of drug abuse comorbid with chronic pain promotes higher rates of opioid misuse. Here, we tested the hypothesis that inflammatory pain leads to increased heroin self-administration resulting from altered mu opioid receptor (MOR) regulation of mesolimbic dopamine (DA) transmission. To this end, the complete Freund's adjuvant (CFA) model of inflammation was used to assess the neurochemical and functional changes induced by inflammatory pain on MOR-mediated mesolimbic DA transmission and on rat intravenous heroin self-administration under fixed ratio (FR) and progressive ratio (PR) schedules of reinforcement. In the presence of inflammatory pain, heroin intake under an FR schedule was increased for high, but attenuated for low, heroin doses with concomitant alterations in mesolimbic MOR function suggested by DA microdialysis. Consistent with the reduction in low dose FR heroin self-administration, inflammatory pain reduced motivation for a low dose of heroin, as measured by responding under a PR schedule of reinforcement, an effect dissociable from high heroin dose PR responding. Together, these results identify a connection between inflammatory pain and loss of MOR function in the mesolimbic dopaminergic pathway that increases intake of high doses of heroin. These findings suggest that pain-induced loss of MOR function in the mesolimbic pathway may promote opioid dose escalation and contribute to opioid abuse-associated phenotypes. SIGNIFICANCE STATEMENT This study provides critical new insights that show that inflammatory pain alters heroin intake through a desensitization of MORs located within the VTA. These findings expand our knowledge of the interactions between

  5. Oral administration of lactulose: a novel therapy for acute carbon monoxide poisoning via increasing intestinal hydrogen production.

    PubMed

    Fan, Dan-Feng; Hu, Hui-Jun; Sun, Xue-Jun; Meng, Xiang-En; Zhang, Yu; Pan, Shu-Yi

    2016-01-01

    It has been known that the pathophysiology of carbon monoxide (CO) poisoning is related to hypoxia, the increased production of reactive oxygen species (ROS) and oxidative stress. Studies have shown that the novel, safe and effective free radical scavenger, hydrogen, has neuroprotective effects in both acute CO poisoning and delayed neuropsychological sequelae in CO poisoning. Orally administered lactulose, which may be used by some intestinal bacteria as a food source to produce endogenous hydrogen, can ameliorate oxidative stress. Based on the available findings, we hypothesize that oral administration of lactulose may be a novel therapy for acute CO poisoning via increasing intestinal hydrogen production.

  6. Acute Carnosine Administration Increases Respiratory Chain Complexes and Citric Acid Cycle Enzyme Activities in Cerebral Cortex of Young Rats.

    PubMed

    Macedo, Levy W; Cararo, José H; Maravai, Soliany G; Gonçalves, Cinara L; Oliveira, Giovanna M T; Kist, Luiza W; Guerra Martinez, Camila; Kurtenbach, Eleonora; Bogo, Maurício R; Hipkiss, Alan R; Streck, Emilio L; Schuck, Patrícia F; Ferreira, Gustavo C

    2016-10-01

    Carnosine (β-alanyl-L-histidine) is an imidazole dipeptide synthesized in excitable tissues of many animals, whose biochemical properties include carbonyl scavenger, anti-oxidant, bivalent metal ion chelator, proton buffer, and immunomodulating agent, although its precise physiological role(s) in skeletal muscle and brain tissues in vivo remain unclear. The aim of the present study was to investigate the in vivo effects of acute carnosine administration on various aspects of brain bioenergetics of young Wistar rats. The activity of mitochondrial enzymes in cerebral cortex was assessed using a spectrophotometer, and it was found that there was an increase in the activities of complexes I-III and II-III and succinate dehydrogenase in carnosine-treated rats, as compared to vehicle-treated animals. However, quantitative real-time RT-PCR (RT-qPCR) data on mRNA levels of mitochondrial biogenesis-related proteins (nuclear respiratory factor 1 (Nrf1), peroxisome proliferator-activated receptor-γ coactivator 1-α (Ppargc1α), and mitochondrial transcription factor A (Tfam)) were not altered significantly and therefore suggest that short-term carnosine administration does not affect mitochondrial biogenesis. It was in agreement with the finding that immunocontent of respiratory chain complexes was not altered in animals receiving carnosine. These observations indicate that acute carnosine administration increases the respiratory chain and citric acid cycle enzyme activities in cerebral cortex of young rats, substantiating, at least in part, a neuroprotector effect assigned to carnosine against oxidative-driven disorders. PMID:26476839

  7. Acute Carnosine Administration Increases Respiratory Chain Complexes and Citric Acid Cycle Enzyme Activities in Cerebral Cortex of Young Rats.

    PubMed

    Macedo, Levy W; Cararo, José H; Maravai, Soliany G; Gonçalves, Cinara L; Oliveira, Giovanna M T; Kist, Luiza W; Guerra Martinez, Camila; Kurtenbach, Eleonora; Bogo, Maurício R; Hipkiss, Alan R; Streck, Emilio L; Schuck, Patrícia F; Ferreira, Gustavo C

    2016-10-01

    Carnosine (β-alanyl-L-histidine) is an imidazole dipeptide synthesized in excitable tissues of many animals, whose biochemical properties include carbonyl scavenger, anti-oxidant, bivalent metal ion chelator, proton buffer, and immunomodulating agent, although its precise physiological role(s) in skeletal muscle and brain tissues in vivo remain unclear. The aim of the present study was to investigate the in vivo effects of acute carnosine administration on various aspects of brain bioenergetics of young Wistar rats. The activity of mitochondrial enzymes in cerebral cortex was assessed using a spectrophotometer, and it was found that there was an increase in the activities of complexes I-III and II-III and succinate dehydrogenase in carnosine-treated rats, as compared to vehicle-treated animals. However, quantitative real-time RT-PCR (RT-qPCR) data on mRNA levels of mitochondrial biogenesis-related proteins (nuclear respiratory factor 1 (Nrf1), peroxisome proliferator-activated receptor-γ coactivator 1-α (Ppargc1α), and mitochondrial transcription factor A (Tfam)) were not altered significantly and therefore suggest that short-term carnosine administration does not affect mitochondrial biogenesis. It was in agreement with the finding that immunocontent of respiratory chain complexes was not altered in animals receiving carnosine. These observations indicate that acute carnosine administration increases the respiratory chain and citric acid cycle enzyme activities in cerebral cortex of young rats, substantiating, at least in part, a neuroprotector effect assigned to carnosine against oxidative-driven disorders.

  8. Effectiveness of increasing the frequency of posaconazole syrup administration to achieve optimal plasma concentrations in patients with haematological malignancy.

    PubMed

    Park, Wan Beom; Cho, Joo-Youn; Park, Sang-In; Kim, Eun Jung; Yoon, Seonghae; Yoon, Seo Hyun; Lee, Jeong-Ok; Koh, Youngil; Song, Kyoung-Ho; Choe, Pyoeng Gyun; Yu, Kyung-Sang; Kim, Eu Suk; Bang, Su Mi; Kim, Nam Joong; Kim, Inho; Oh, Myoung-Don; Kim, Hong Bin; Song, Sang Hoon

    2016-07-01

    Few data are available on whether adjusting the dose of posaconazole syrup is effective in patients receiving anti-cancer chemotherapy. The aim of this prospective study was to analyse the impact of increasing the frequency of posaconazole administration on optimal plasma concentrations in adult patients with haematological malignancy. A total of 133 adult patients receiving chemotherapy for acute myeloid leukaemia or myelodysplastic syndrome who received posaconazole syrup 200 mg three times daily for fungal prophylaxis were enrolled in this study. Drug trough levels were measured by liquid chromatography-tandem mass spectrometry. In 20.2% of patients (23/114) the steady-state concentration of posaconazole was suboptimal (<500 ng/mL) on Day 8. In these patients, the frequency of posaconazole administration was increased to 200 mg four times daily. On Day 15, the median posaconazole concentration was significantly increased from 368 ng/mL [interquartile range (IQR), 247-403 ng/mL] to 548 ng/mL (IQR, 424-887 ng/mL) (P = 0.0003). The median increase in posaconazole concentration was 251 ng/mL (IQR, 93-517 ng/mL). Among the patients with initially suboptimal levels, 79% achieved the optimal level unless the steady-state level was <200 ng/mL. This study shows that increasing the administration frequency of posaconazole syrup is effective for achieving optimal levels in patients with haematological malignancy undergoing chemotherapy. PMID:27234674

  9. The modulatory role of M2 muscarinic receptor on apomorphine-induced yawning and genital grooming.

    PubMed

    Gamberini, Maria Thereza; Bolognesi, Maria Laura; Nasello, Antonia Gladys

    2012-12-01

    The interaction between dopaminergic and cholinergic pathways in the induction of behavioral responses has been previously established. In the brain, M2 receptors are found predominantly in presynaptic cholinergic neurons as autoreceptors, and in dopaminergic neurons as heteroceptors, suggesting a control role of acetylcholine and dopamine release, respectively. Our aim was to investigate the role of M2 receptors on the yawning and genital grooming of rats induced by apomorphine, a dopaminergic receptor agonist, focusing on the interaction between cholinergic and dopaminergic pathways. Initially, the effect of atropine, a non-selective muscarinic antagonist, on yawning and genital grooming induced by apomorphine (100 μg/kg s.c.) was analyzed. Atropine doses of 0.5, 1 and 2 mg/kg i.p. were administered to Wistar rats 30 min before induction of the behavioral responses by apomorphine. Number of yawns and time spent genital grooming were quantified over a 60 min period. Apomorphine-induced yawning was increased by low dose (0.5 mg/kg i.p.) but not by high doses (1 and 2 mg/kg, i.p.) of atropine. Genital grooming was antagonized by 2 mg/kg i.p. of atropine and showed no changes at the other doses tested. Tripitramine, a selective M2 cholinergic antagonist, was used as a tool for distinguishing between M2 and all other muscarinic receptor subtypes in yawning and genital grooming. Tripitramine doses of 0.01, 0.02 and 0.04 μmol/kg i.p. were administered to Wistar rats 30 min before apomorphine (100 μg/kg s.c.). Number of yawns and time spent genital grooming were also quantified over a 60 min period. Tripitramine 0.01 μmol/kg increased all parameters. Higher doses, which possibly block all subtypes of muscarinic receptor, did not modify the response of apomorphine, suggesting a non-selective effect of tripitramine at these doses. Given that low doses of tripitramine increased the behavioral responses induced by apomorphine and that the main distribution of the M2

  10. Cholinergic stimulation of pancreatic amylase release and muscarinic receptors: effect of ionophore A23187

    SciTech Connect

    Larose, L.; Morisset, J.

    1985-07-22

    Dispersed rat pancreatic acini were incubated in 0.5 mM calcium medium with increasing concentrations of carbamylcholine, with or without the ionophore A23187 (10/sup -6/M). Addition of the ionophore reduced maximal amylase release, increased the maximal effective concentration of carbamylcholine and dramatically impaired the agonist's capacity to induce enzyme secretion at low concentration. The ionophore also abolished the inhibition of secretion observed at high carbamylcholine concentrations. These effects of the ionophore on the cholinergic secretory response cannot be explained by interaction at the muscarinic receptor since neither the Bmax, the affinity of the receptor for the (/sup 3/H)QNB nor the binding of carbamylcholine were affected by the ionophore. It is suggested that for the conditions studied, the ionophore can interact with the secretory process at one or several points ulterior to the initial recognition site of carbamylcholine on its receptor. 30 references, 3 figures.

  11. Allosteric interactions of three muscarine antagonists at bovine tracheal smooth muscle and cardiac M2 receptors.

    PubMed

    Roffel, A F; Elzinga, C R; Meurs, H; Zaagsma, J

    1989-03-01

    The kinetics of [3H]dexetimide dissociation from muscarine receptors in bovine cardiac left ventricular and tracheal smooth muscle membranes were studied in the absence and presence of three muscarine antagonists. It was found that [3H]dexetimide dissociation from cardiac muscarine receptors was monophasic and very fast (half life less than 1 min) and was slowed by the cardioselective muscarine antagonists, gallamine, methoctramine and AF-DX 116, concentration dependently. [3H]Dexetimide dissociation from tracheal muscarine receptors was biphasic, with a fast phase (half-life less than 1 min) followed after 4-5 min by a slow phase (half-life = 38.5 min). The fast component, but not the slow component, was slowed by the muscarine antagonists with concentration dependencies very similar to those found in the heart. We conclude from these data that the major population of tracheal smooth muscle muscarine receptors resembles the cardiac M2 type not only with respect to equilibrium binding affinities but also with respect to the secondary, allosteric binding site on the muscarine receptor. The results also imply that the cardiac receptor subtype is much more sensitive to allosteric modulation than the glandular/smooth muscle receptor subtype. PMID:2714370

  12. Distribution and effects of the muscarinic receptor subtypes in the primary visual cortex

    PubMed Central

    Groleau, Marianne; Kang, Jun Il; Huppé-Gourgues, Frédéric; Vaucher, Elvire

    2015-01-01

    Muscarinic cholinergic receptors modulate the activity and plasticity of the visual cortex. Muscarinic receptors are divided into five subtypes that are not homogeneously distributed throughout the cortical layers and cells types. This distribution results in complex action of the muscarinic receptors in the integration of visual stimuli. Selective activation of the different subtypes can either strengthen or weaken cortical connectivity (e.g., thalamocortical vs. corticocortical), i.e., it can influence the processing of certain stimuli over others. Moreover, muscarinic receptors differentially modulate some functional properties of neurons during experience-dependent activity and cognitive processes and they contribute to the fine-tuning of visual processing. These functions are involved in the mechanisms of attention, maturation and learning in the visual cortex. This minireview describes the anatomo-functional aspects of muscarinic modulation of the primary visual cortex’s (V1) microcircuitry. PMID:26150786

  13. Muscarinic M2 receptors in bovine tracheal smooth muscle: discrepancies between binding and function.

    PubMed

    Roffel, A F; Elzinga, C R; Van Amsterdam, R G; De Zeeuw, R A; Zaagsma, J

    1988-08-01

    Previous work showing that AF-DX 116, a cardioselective muscarinic antagonist in functional experiments, does not discriminate between muscarinic receptors in bovine cardiac and tracheal membranes has been extended. In addition to AF-DX 116 we used the muscarinic antagonists, atropine, pirenzepine, 4-DAMP methobromide, gallamine, hexahydrosiladifenidol and methoctramine, in radioligand binding experiments on bovine cardiac left ventricular and tracheal smooth muscle membranes. The functional antagonism of the methacholine-induced contraction of bovine tracheal smooth muscle strips was also evaluated. An excellent correlation was found for all compounds between the binding affinities for muscarinic receptors in cardiac and tracheal smooth muscle membranes; moreover, the affinities found in cardiac membranes correspond with the pA2 values reported for atrial preparations of rat and guinea pig. However, significant and occasionally marked discrepancies were found between binding and functional affinities of these muscarinic antagonists on bovine tracheal smooth muscle. PMID:3215279

  14. The use of a psychological intervention to increase adherence during factor administration in a child with haemophilia.

    PubMed

    Penica, S; Williams, K E

    2008-09-01

    While factor replacement treatments allow children with haemophilia to lead near normal lives, these treatments can be difficult to administer, especially to younger children. The intervenous infusions required by these treatments can be painful and result in children attempting to avoid treatment by exhibiting a range of inappropriate behaviours. Their children's uncooperative behaviour during prophylaxis was cited by parents as a significant barrier to treatment adherence. This study provides a case illustration of the use of psychological interventions to increase adherence during factor administration. Single-case methodology was used to demonstrate the effectiveness of the psychological interventions including counterconditioning, distraction, and positive differential reinforcement. The intervention resulted in increased adherence across several months of intervention. Psychological interventions can be effectively used by caregivers and care providers to increase adherence in the treatment of haemophilia.

  15. Laboratory alcohol self-administration experiments do not increase subsequent real-life drinking in young adult social drinkers

    PubMed Central

    Sommer, Christian; Seipt, Christian; Spreer, Maik; Blümke, Toni; Markovic, Alexandra; Jünger, Elisabeth; Plawecki, Martin H.; Zimmermann, Ulrich S.

    2015-01-01

    Background While the utility of experimental free-access alcohol self-administration paradigms is well-established, little data exist addressing the question of whether study participation influences subsequent natural alcohol consumption. We here present drinking reports of young adults before and after participation in intravenous alcohol self-administration studies. Methods Timeline Follow-back (TLFB) drinking reports for the 6 weeks immediately preceding the first, and the 6 weeks after the last experimental alcohol challenge were examined from subjects completing one of two similar alcohol self-administration paradigms. In study 1, eighteen social drinkers (9 females, mean age 24.1 years) participated in 3 alcohol self-infusion sessions up to a maximum blood alcohol concentration (BAC) of 160 mg%. Study 2 involved 60 participants (30 females, mean age 18.3 years) of the Dresden Longitudinal Study on Alcohol Use in Young Adults (D-LAYA), who participated in 2 sessions of alcohol self-infusion up to a maximum BAC of 120 mg%, and a non-exposed age- matched control group of 42 (28 females, mean age 18.4 years) subjects. Results In study 1, participants reported (3.7%) fewer heavy drinking days as well as a decrease of 2.5 drinks per drinking day after study participation compared to pre-study levels (p<.05 respectively).. In study 2, alcohol-exposed participants reported 7.1% and non- alcohol-exposed controls 6.5% fewer drinking days at post-study measurement (p<.001), while percent heavy drinking days and drinks per drinking day did not differ. Conclusion These data suggest that participation in intravenous alcohol self-administration experiments does not increase subsequent real-life drinking of young adults. PMID:25903217

  16. Physiological and biochemical studies of newly synthesized muscarinic acetylcholine receptors in embryonic chicken heart

    SciTech Connect

    Hunter, D.D.

    1986-01-01

    Exposure of either chicken embryos in ovo or cultured embryonic chicken cardiac cells in vitro to the muscarinic agonist carbachol results in a 70-90% decrease in the number of muscarinic acetylcholine receptors (mAChR) expressed in cardiac cells. Block of agonist-receptor interactions in ovo with the antagonist atropine or removal of the agonist in vitro results in a gradual increase in mAChR number, reaching the control level in 14 hr. Measurements of physiological sensitivity of atria or cultured cells show that, even after the complete recovery of receptor number, the sensitivity to agonist is reduced. The sensitivity of the mAChR-mediated inhibition of adenylate cyclase is also decreased at this time. Newly synthesized mAChR which appear following affinity alkylation in cultured cells are also poorly coupled to the stimulation of /sup 86/Rb/sup +/ efflux, indicating that decreased physiological sensitivity is not due to an unknown effect of long-term agonist exposure on general cellular function, but rather reflects an intrinsic property of newly synthesized mAChR. This increase in sensitivity is also not blocked by cycloheximide. The increase in sensitivity of the mAChR-mediated responses is due neither to a lack of expression of newly synthesized mAChR on the surface nor to reduced agonist affinity of the mAChR. The diminished sensitivity and subsequent maturation observed in cells containing newly synthesized receptors is due either to a small change in mAChR, or to a change in an as-yet-undefined component of the mAChR transduction system; this alteration represents a novel locus for modulation of cholinergic signals in the heart.

  17. Programmed administration of parathyroid hormone increases bone formation and reduces bone loss in hindlimb-unloaded ovariectomized rats

    NASA Technical Reports Server (NTRS)

    Turner, R. T.; Evans, G. L.; Cavolina, J. M.; Halloran, B.; Morey-Holton, E.

    1998-01-01

    Gonadal insufficiency and reduced mechanical usage are two important risk factors for osteoporosis. The beneficial effects of PTH therapy to reverse the estrogen deficiency-induced bone loss in the laboratory rat are well known, but the influence of mechanical usage in this response has not been established. In this study, the effects of programed administration of PTH on cancellous bone volume and turnover at the proximal tibial metaphysis were determined in hindlimb-unloaded, ovariectomized (OVX), 3-month-old Sprague-Dawley rats. PTH was administered to weight-bearing and hindlimb-unloaded OVX rats with osmotic pumps programed to deliver 20 microg human PTH (approximately 80 microg/kg x day) during a daily 1-h infusion for 7 days. Compared with sham-operated rats, OVX increased longitudinal and radial bone growth, increased indexes of cancellous bone turnover, and resulted in net resorption of cancellous bone. Hindlimb unloading of OVX rats decreased longitudinal and radial bone growth, decreased osteoblast number, increased osteoclast number, and resulted in a further decrease in cancellous bone volume compared with those in weight-bearing OVX rats. Programed administration of PTH had no effect on either radial or longitudinal bone growth in weight-bearing and hindlimb-unloaded OVX rats. PTH treatment had dramatic effects on selected cancellous bone measurements; PTH maintained cancellous bone volume in OVX weight-bearing rats and greatly reduced cancellous bone loss in OVX hindlimb-unloaded rats. In the latter animals, PTH treatment prevented the hindlimb unloading-induced reduction in trabecular thickness, but the hormone was ineffective in preventing either the increase in osteoclast number or the loss of trabecular plates. Importantly, PTH treatment increased the retention of a baseline flurochrome label, osteoblast number, and bone formation in the proximal tibial metaphysis regardless of the level of mechanical usage. These findings demonstrate that

  18. Contextual cues associated with nicotine administration increase arc mRNA expression in corticolimbic areas of the rat brain.

    PubMed

    Schiltz, Craig A; Kelley, Ann E; Landry, Charles F

    2005-03-01

    Conditioned responses to cues associated with the administration of drugs of misuse are an impediment to continued abstinence for drug-free addicted individuals. In order to study the neuroanatomical and cellular response of the brain to cues associated with nicotine administration, we conditioned Sprague-Dawley rats to receive an ascending dose regimen of nicotine over 14 days in two distinct non-home cage environments and assessed expression of the early response gene arc in corticolimbic areas in response to the nicotine-associated context. All of the rats received the same dose regimen of nicotine. Three days after the last training day, the rats were exposed to the test environment. The rats that had previously received nicotine exhibited increased motor activity compared with the rats that had received saline in the test environment. After 45 min in the test environment, brains were taken for Northern blotting and in situ hybridization analysis, which revealed an increase in levels of activity-regulated, dendritically localized mRNA for arc in a variety of brain regions (medial and lateral prefrontal cortices, cingulate cortex, primary sensory cortex, sensorimotor cortex, ventral striatum and amygdala). Plasma corticosterone levels were not different between the groups, suggesting that exposure to nicotine cues is insufficient to activate the hypothalamo-pituitary-adrenal axis. Given that Arc plays a direct role in neuronal plasticity and memory consolidation, its induction by nicotine-associated cues in brain regions critical for cognitive and emotional processing suggests that rats may be learning that these cues are no longer necessarily predictive of nicotine administration. Further work will be needed in order to assess the role of arc expression in the extinction of conditioned responses to drug-paired cues.

  19. Trans-stilbene oxide administration increased hepatic glucuronidation of morphine but decreased biliary excretion of morphine glucuronide in rats

    SciTech Connect

    Fuhrman-Lane, C.; Fujimoto, J.M.

    1982-09-01

    The effect of the inducing agent trans-stilbene oxide (TSO) on the metabolism and biliary excretion of (/sup 14/C)morphine was studied in the isolated in situ perfused rat liver. After administration of morphine by intraportal injection or by the segmented retrograde intrabiliary injection technique, the TSO-treated group showed a marked decrease in the biliary recovery of morphine as its glucuronide conjugate (morphine-3-glucuronide (MG)). However, recovery of MG in the venous outflow of the single pass perfusate was greatly increased. These findings suggested that TSO treatment enhanced the formation of MG from morphine and changed the primary route of hepatic elimination of MG. TSO treatment also decreased the excretion of morphine (as MG) in the bile of anesthetized renal-ligated rats. This decreased biliary function required several days to develop and appeared closely associated with the inductive effect of TSO. After i.v. administration of (/sup 14/C)MG itself, biliary recovery was also markedly decreased in TSO-treated rats. It is postulated that the effect of the TSO treatment led to either a decrease in canalicular transport of MG into bile or an increase in the efficiency of transfer of MG to the blood at the sinusoidal side of the hepatocyte. Regardless of the mechanism, the results indicate the need to study compartmentalization of drug transport and metabolism functions.

  20. Ventilatory effects of low-dose paraoxon result from central muscarinic effects

    SciTech Connect

    Houze, Pascal; Pronzola, Laetita; Kayouka, Maya; Villa, Antoine; Debray, Marcel; Baud, Frederic J.

    2008-12-01

    Paraoxon induces respiratory toxicity. Atropine completely reversed parathion- and paraoxon-induced respiratory toxicity. The aim of this study was to assess the peripheral or central origin of ventilatory effects of low-dose paraoxon. Male Sprague-Dawley rats were given paraoxon 0.215 mg/kg subcutaneously and treated with either atropine (10 mg/kg sc) or ascending doses of methylatropine of 5.42 (equimolar to that of atropine), 54.2, and 542 mg/kg administered subcutaneously 30 min after paraoxon. Ventilation at rest was assessed using whole-body plethysmography and rat temperature using infra-red telemetry. Results are expressed as mean {+-} SE. Statistical analysis used two-way ANOVA for repeated measurements. Paraoxon induced a significant decrease in temperature 30 min after injection lasting the 90 min of the study period. This effect was partially corrected by atropine, but not by methylatropine whatever the dose. Paraoxon induced a decrease in respiratory rate resulting from an increase in expiratory time associated with an increase in tidal volume. Atropine completely reversed the ventilatory effects of low-dose paraoxon while the equimolar dose of methylatropine had no significant effects. The 54.2 and 542 mg/kg doses of methylatropine had no significant effects. Atropine crosses the blood-brain barrier and reverses peripheral and central muscarinic effects. In contrast, methylatropine does not cross the blood-brain barrier. Atropine completely reversed the ventilatory effects of low-dose paraoxon, while methylatropine had no significant effects at doses up to 100-fold the equimolar dose of atropine. We conclude that the ventilatory effects of low-dose paraoxon are mediated by disrupted muscarinic signaling in the central nervous system.

  1. CENTRAL ADMINISTRATION OF THE RF-AMIDE PEPTIDES, QRFP-26 AND QRFP-43, INCREASES HIGH FAT FOOD INTAKE IN RATS

    PubMed Central

    Primeaux, Stefany D.; Blackmon, Christine; Barnes, Maria J.; Braymer, H. Douglas; Bray, George A.

    2008-01-01

    Pyrogultamylated arginine-phenylalanineamide peptide (QRFP) is strongly conserved across species and is a member of the family of RFamide-related peptides, with the motif Arg-Phe-NH2 at the C-terminal end. The precursor peptide for QRFP generates a 26-amino acid peptide (QRFP-26) and a 43-amino acid peptide (QRFP-43), both of which bind to the G protein-coupled receptor, GPR103. Recently, QRFP has been characterized in rats, mice and humans and has been reported to have orexigenic properties. In rodents, prepro-QRFP mRNA is expressed in localized regions of the mediobasal hypothalamus, a region implicated in feeding behavior. Increased intake of a high fat diet contributes to increased weight gain and obesity. Therefore, the current experiments investigated the effects of QRFP administration in rats and the effects of a high fat diet on prepro-QRFP mRNA and GPR103 receptor mRNA levels. Intracerebroventricular administration of QRFP-26 (3.0nM, 5.0nM) and QRFP-43 (1.0nM, 3.0nM) dose-dependently increased 1h, 2h, and 4h cumulative intake of high fat (55% fat), but not low fat (10% fat) diet. In Experiment 2, hypothalamic prepro-QRFP mRNA levels and GPR103 receptor mRNA levels were measured in rats fed a high fat or a low fat diet for 21 days. Prepro-QRFP mRNA was significantly increased in the ventromedial nucleus/arcuate nucleus of the hypothalamus of rats fed a high fat diet compared to those fed a low fat diet, while GPR103 mRNA levels were unchanged. These findings suggest that QRFP is a regulator of dietary fat intake and is influenced by the intake of a high fat diet. PMID:18765262

  2. Activation of muscarinic receptors by non-neuronal acetylcholine.

    PubMed

    Wessler, Ignaz Karl; Kirkpatrick, Charles James

    2012-01-01

    The biological role of acetylcholine and the cholinergic system is revisited based particularly on scientific research early and late in the last century. On the one hand, acetylcholine represents the classical neurotransmitter, whereas on the other hand, acetylcholine and the pivotal components of the cholinergic system (high-affinity choline uptake, choline acetyltransferase and its end product acetylcholine, muscarinic and nicotinic receptors and esterase) are expressed by more or less all mammalian cells, i.e. by the majority of cells not innervated by neurons at all. Moreover, it has been demonstrated that acetylcholine and "cholinergic receptors" are expressed in non-neuronal organisms such as plants and protists. Acetylcholine is even synthesized by bacteria and algae representing an extremely old signalling molecule on the evolutionary timescale. The following article summarizes examples, in which non-neuronal acetylcholine is released from primitive organisms as well as from mammalian non-neuronal cells and binds to muscarinic receptors to modulate/regulate phenotypic cell functions via auto-/paracrine pathways. The examples demonstrate that non-neuronal acetylcholine and the non-neuronal cholinergic system are vital for various types of cells such as epithelial, endothelial and immune cells.

  3. Acute Administration of Branched-Chain Amino Acids Increases the Pro-BDNF/Total-BDNF Ratio in the Rat Brain.

    PubMed

    Scaini, Giselli; Morais, Meline O S; Furlanetto, Camila B; Kist, Luiza W; Pereira, Talita C B; Schuck, Patrícia F; Ferreira, Gustavo C; Pasquali, Matheus A B; Gelain, Daniel P; Moreira, José Cláudio F; Bogo, Maurício R; Streck, Emilio L

    2015-05-01

    Maple syrup urine disease (MSUD) is caused by an inborn error in metabolism resulting from a deficiency in the branched-chain α-keto acid dehydrogenase complex activity. This blockage leads to accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine and valine, as well as their corresponding α-keto acids and α-hydroxy acids. High levels of BCAAs are associated with neurological dysfunction and the role of pro- and mature brain-derived neurotrophic factor (BDNF) in the neurological dysfunction of MSUD is still unclear. Thus, in the present study we investigated the effect of an acute BCAA pool administration on BDNF levels and on the pro-BDNF cleavage-related proteins S100A10 and tissue plasminogen activator (tPA) in rat brains. Our results demonstrated that acute Hyper-BCAA (H-BCAA) exposure during the early postnatal period increases pro-BDNF and total-BDNF levels in the hippocampus and striatum. Moreover, tPA levels were significantly decreased, without modifications in the tPA transcript levels in the hippocampus and striatum. On the other hand, the S100A10 mRNA and S100A10 protein levels were not changed in the hippocampus and striatum. In the 30-day-old rats, we observed increased pro-BDNF, total-BDNF and tPA levels only in the striatum, whereas the tPA and S100A10 mRNA expression and the immunocontent of S100A10 were not altered. In conclusion, we demonstrated that acute H-BCAA administration increases the pro-BDNF/total-BDNF ratio and decreases the tPA levels in animals, suggesting that the BCAA effect may depend, at least in part, on changes in BDNF post-translational processing. PMID:25681161

  4. Oral adenosine-5’-triphosphate (ATP) administration increases blood flow following exercise in animals and humans

    PubMed Central

    2014-01-01

    Introduction Extracellular adenosine triphosphate (ATP) stimulates vasodilation by binding to endothelial ATP-selective P2Y2 receptors; a phenomenon, which is posited to be accelerated during exercise. Herein, we used a rat model to examine how different dosages of acute oral ATP administration affected the femoral blood flow response prior to, during, and after an exercise bout. In addition, we performed a single dose chronic administration pilot study in resistance trained athletes. Methods Animal study: Male Wistar rats were gavage-fed the body surface area, species adjusted human equivalent dose (HED) of either 100 mg (n=4), 400 mg (n=4), 1,000 mg (n=5) or 1,600 mg (n=5) of oral ATP as a disodium salt (Peak ATP®, TSI, Missoula, MT). Rats that were not gavage-fed were used as controls (CTL, n=5). Blood flow was monitored continuously: a) 60 min prior to, b) during and c) 90 min following an electrically-evoked leg-kicking exercise. Human Study: In a pilot study, 12 college-aged resistance-trained subjects were given 400 mg of ATP (Peak ATP®, TSI, Missoula, MT) daily for 12 weeks, and prior to an acute arm exercise bout at weeks 1, 4, 8, and 12. Ultrasonography-determined volumetric blood flow and vessel dilation in the brachial artery was measured at rest, at rest 30 minutes after supplementation, and then at 0, 3, and 6 minutes after the exercise. Results Animal Study: Rats fed 1,000 mg HED demonstrated significantly greater recovery blood flow (p < 0.01) and total blood flow AUC values (p < 0.05) compared to CTL rats. Specifically, blood flow was elevated in rats fed 1,000 mg HED versus CTL rats at 20 to 90 min post exercise when examining 10-min blood flow intervals (p < 0.05). When examining within-group differences relative to baseline values, rats fed the 1,000 mg and 1,600 mg HED exhibited the most robust increases in blood flow during exercise and into the recovery period. Human study: At weeks 1, 8, and 12, ATP supplementation significantly increased

  5. Muscarine-like compounds derived from a pyrolysis product of cellulose.

    PubMed

    Defant, Andrea; Mancini, Ines; Matucci, Rosanna; Bellucci, Cristina; Dosi, Federico; Malferrari, Danilo; Fabbri, Daniele

    2015-06-14

    Cellulose represents a key component of a renewable biomass source, from which chiral compounds with a high added value in the application for the synthesis of potentially bioactive molecules can be obtained. The anhydrosugar (1R,5S)-1-hydroxy-3,6-dioxa-bicyclo[3.2.1]octan-2-one (LAC), produced on the gram-scale by catalytic pyrolysis of cellulose, was used as a building block in the synthesis of five new enantiomerically pure muscarine-like products. The structures of the target compounds 4-8 showed different substituents at the C-2 and C-4 positions, but each of them had the same (2S,4R) configuration as the natural (+)-muscarine. A renewed interest in new muscarinic analogues is due to the design and synthesis of molecules exhibiting a higher selectivity for a specific muscarinic receptor and due to the development of effective agents in the treatment of Alzheimer's disease and other cognitive disorders. In this context, products 4-8 were investigated with respect to their binding affinity to human M1-M5 muscarinic acetylcholine receptors. The data indicated that compound 8, emerging as the most active in the series with values comparable to natural (+)-muscarine and a moderate selectivity in favor of the hM2 subtype receptor, also exhibited the highest stability during the interaction with the hM2 (3UON) subtype muscarinic receptor by using a docking calculation.

  6. Muscarinic receptor immunoreactivity in the superior salivatory nucleus neurons innervating the salivary glands of the rat.

    PubMed

    Ueda, Hirotaka; Mitoh, Yoshihiro; Fujita, Masako; Kobashi, Motoi; Yamashiro, Takashi; Sugimoto, Tomosada; Ichikawa, Hiroyuki; Matsuo, Ryuji

    2011-07-15

    The superior salivatory nucleus (SSN) contains preganglionic parasympathetic neurons to the submandibular and sublingual salivary glands. Cevimeline, a muscarinic acetylcholine receptor agonist, stimulates the salivary glands and is presently used as sialogogue in the treatment of dry mouth. Since cevimeline passes through the blood-brain barrier, it is also able to act on muscarinic acetylcholine receptors in the central nervous system. Our preliminary experiment using the whole-cell patch-clamp technique has shown that cevimeline excites SSN neurons in rat brain slices, suggesting that SSN neurons have muscarinic acetylcholine receptors; however, it is unclear which subtypes of muscarinic acetylcholine receptors exist in SSN neurons. In the present study, we investigated immunohistochemically muscarinic acetylcholine receptor subtypes, M1 receptor (M1R), M2R, M3R, M4R, and M5R in SSN neurons. SSN neurons innervating the salivary glands, retrogradely labeled with a fluorescent tracer from the chorda-lingual nerve, mostly expressed M3R immunoreactivity (-ir) (92.3%) but not M1R-ir. About half of such SSN neurons also showed M2R- (40.1%), M4R- (54.0%) and M5R-ir (46.0%); therefore, it is probable that SSN neurons co-express M3R-ir with at least two of the other muscarinic receptor subtypes. This is the first report to show that SSN neurons contain muscarinic acetylcholine receptors.

  7. Coupling of G Proteins to Reconstituted Monomers and Tetramers of the M2 Muscarinic Receptor*

    PubMed Central

    Redka, Dar'ya S.; Morizumi, Takefumi; Elmslie, Gwendolynne; Paranthaman, Pranavan; Shivnaraine, Rabindra V.; Ellis, John; Ernst, Oliver P.; Wells, James W.

    2014-01-01

    G protein-coupled receptors can be reconstituted as monomers in nanodiscs and as tetramers in liposomes. When reconstituted with G proteins, both forms enable an allosteric interaction between agonists and guanylyl nucleotides. Both forms, therefore, are candidates for the complex that controls signaling at the level of the receptor. To identify the biologically relevant form, reconstituted monomers and tetramers of the purified M2 muscarinic receptor were compared with muscarinic receptors in sarcolemmal membranes for the effect of guanosine 5′-[β,γ-imido]triphosphate (GMP-PNP) on the inhibition of N-[3H]methylscopolamine by the agonist oxotremorine-M. With monomers, a stepwise increase in the concentration of GMP-PNP effected a lateral, rightward shift in the semilogarithmic binding profile (i.e. a progressive decrease in the apparent affinity of oxotremorine-M). With tetramers and receptors in sarcolemmal membranes, GMP-PNP effected a vertical, upward shift (i.e. an apparent redistribution of sites from a state of high affinity to one of low affinity with no change in affinity per se). The data were analyzed in terms of a mechanistic scheme based on a ligand-regulated equilibrium between uncoupled and G protein-coupled receptors (the “ternary complex model”). The model predicts a rightward shift in the presence of GMP-PNP and could not account for the effects at tetramers in vesicles or receptors in sarcolemmal membranes. Monomers present a special case of the model in which agonists and guanylyl nucleotides interact within a complex that is both constitutive and stable. The results favor oligomers of the M2 receptor over monomers as the biologically relevant state for coupling to G proteins. PMID:25023280

  8. Repeated oral administration of capsaicin increases anxiety-like behaviours with prolonged stress-response in rats.

    PubMed

    Choi, Y-J; Kim, J Y; Yoo, S B; Lee, J-H; Jahng, J W

    2013-09-01

    This study was conducted to examine the psycho-emotional effects of repeated oral exposure to capsaicin, the principal active component of chili peppers. Each rat received 1 mL of 0.02 percent capsaicin into its oral cavity daily, and was subjected to behavioural tests following 10 daily administrations of capsaicin. Stereotypy counts and rostral grooming were significantly increased, and caudal grooming decreased, in capsaicin-treated rats during the ambulatory activity test. In elevated plus maze test, not only the time spent in open arms but also the percent arm entry into open arms was reduced in capsaicin-treated rats compared with control rats. In forced swim test, although swimming duration was decreased, struggling increased in the capsaicin group, immobility duration did not differ between the groups. Repeated oral capsaicin did not affect the basal levels of plasma corticosterone; however, the stress-induced elevation of plasma corticosterone was prolonged in capsaicin treated rats. Oral capsaicin exposure significantly increased c-Fos expression not only in the nucleus tractus of solitarius but also in the paraventricular nucleus. Results suggest that repeated oral exposure to capsaicin increases anxiety-like behaviours in rats, and dysfunction of the hypothalamic-pituitary-adrenal axis may play a role in its pathophysiology. PMID:23938388

  9. Brief intermittent cocaine self-administration and abstinence sensitizes cocaine effects on the dopamine transporter and increases drug seeking.

    PubMed

    Calipari, Erin S; Siciliano, Cody A; Zimmer, Benjamin A; Jones, Sara R

    2015-02-01

    Although traditional sensitization paradigms, which result in an augmentation of cocaine-induced locomotor behavior and dopamine (DA) overflow following repeated experimenter-delivered cocaine injections, are often used as a model to study drug addiction, similar effects have been difficult to demonstrate following cocaine self-administration. We have recently shown that intermittent access (IntA) to cocaine can result in increased cocaine potency at the DA transporter (DAT); however, traditional sensitization paradigms often show enhanced effects following withdrawal/abstinence periods. Therefore, we determined a time course of IntA-induced sensitization by examining the effects of 1 or 3 days of IntA, as well as a 7-day abstinence period on DA function, cocaine potency, and reinforcement. Here we show that cocaine potency is increased following as little as 3 days of IntA and further augmented following an abstinence period. In addition, IntA plus abstinence produced greater evoked DA release in the presence of cocaine as compared with all other groups, demonstrating that following abstinence, both cocaine's ability to increase DA release and inhibit uptake at the DAT, two separate mechanisms for increasing DA levels, are enhanced. Finally, we found that IntA-induced sensitization of the DA system resulted in an increased reinforcing efficacy of cocaine, an effect that was augmented after the 7-day abstinence period. These results suggest that sensitization of the DA system may have an important role in the early stages of drug abuse and may drive the increased drug seeking and taking that characterize the transition to uncontrolled drug use. Human data suggest that intermittency, sensitization, and periods of abstinence have an integral role in the process of addiction, highlighting the importance of utilizing pre-clinical models that integrate these phenomena, and suggesting that IntA paradigms may serve as novel models of human addiction. PMID:25212486

  10. Brief Intermittent Cocaine Self-Administration and Abstinence Sensitizes Cocaine Effects on the Dopamine Transporter and Increases Drug Seeking

    PubMed Central

    Calipari, Erin S; Siciliano, Cody A; Zimmer, Benjamin A; Jones, Sara R

    2015-01-01

    Although traditional sensitization paradigms, which result in an augmentation of cocaine-induced locomotor behavior and dopamine (DA) overflow following repeated experimenter-delivered cocaine injections, are often used as a model to study drug addiction, similar effects have been difficult to demonstrate following cocaine self-administration. We have recently shown that intermittent access (IntA) to cocaine can result in increased cocaine potency at the DA transporter (DAT); however, traditional sensitization paradigms often show enhanced effects following withdrawal/abstinence periods. Therefore, we determined a time course of IntA-induced sensitization by examining the effects of 1 or 3 days of IntA, as well as a 7-day abstinence period on DA function, cocaine potency, and reinforcement. Here we show that cocaine potency is increased following as little as 3 days of IntA and further augmented following an abstinence period. In addition, IntA plus abstinence produced greater evoked DA release in the presence of cocaine as compared with all other groups, demonstrating that following abstinence, both cocaine's ability to increase DA release and inhibit uptake at the DAT, two separate mechanisms for increasing DA levels, are enhanced. Finally, we found that IntA-induced sensitization of the DA system resulted in an increased reinforcing efficacy of cocaine, an effect that was augmented after the 7-day abstinence period. These results suggest that sensitization of the DA system may have an important role in the early stages of drug abuse and may drive the increased drug seeking and taking that characterize the transition to uncontrolled drug use. Human data suggest that intermittency, sensitization, and periods of abstinence have an integral role in the process of addiction, highlighting the importance of utilizing pre-clinical models that integrate these phenomena, and suggesting that IntA paradigms may serve as novel models of human addiction. PMID:25212486

  11. Down-regulation of phospholipase C-beta1 following chronic muscarinic receptor activation.

    PubMed

    Sorensen, S D; Linseman, D A; Fisher, S K

    1998-04-01

    To determine whether prolonged activation of a phospholipase C-coupled receptor can lead to a down-regulation of its effector enzyme, SH-SY5Y neuroblastoma cells were incubated for 24 h with the muscarinic receptor agonist, oxotremorine-M. Under these conditions, significant reductions (46-53%) in muscarinic cholinergic receptor density, G(alphaq/11) and phospholipase C-beta1 (but not the beta3-or gamma1 isoforms) were observed. These results suggest that a selective down-regulation of phospholipase C-beta1 may play a role in adaptation to chronic muscarinic receptor activation. PMID:9617763

  12. Neuropeptide Y administration acutely increases hypothalamic corticotropin-releasing factor immunoreactivity: lack of effect in other rat brain regions

    SciTech Connect

    Haas, D.A.; George, S.R.

    1987-12-21

    The effect of acute central administration of Neuropeptide Y (NPY) to adult male rats on the brain content of corticotropin-releasing factor immunoreactivity (CRF-ir) was investigated. The brain regions studied included frontal cortex, hippocampus, medulla-pons, midbrain-thalamus, cerebellum, neurointermediate lobe of pituitary, median eminence and the remaining hypothalamus. CRF-ir was determined in each of these regions using radioimmunoassay specific for rat CRF. CRF-ir was found to be significantly increased in the major site of CRF localization in the brain, the hypothalamus, in NPY-treated rats as compared to vehicle-treated controls either 15 minutes (p<0.025) or 45 minutes (p<0.005) post-injection. This increase was localized to the median eminence (p<0.05 after 15 minutes, p<0.01 after 45 minutes). No statistically significant differences were noted in any of the other brain regions assessed. Plasma adrenocorticotropin levels were also found to increase following NPY treatment, an effect which became significant after 45 minutes (p<0.05). These data show that NPY can alter the content of hypothalamic CRF and may play a role in its regulation. 33 references, 4 figures.

  13. The administration of atomoxetine during alcohol deprivation induces a time-limited increase in alcohol consumption after relapse.

    PubMed

    Alén, Francisco; Serrano, Antonia; Gorriti, Miguel Ángel; Pavón, Francisco Javier; Orio, Laura; de Heras, Raquel Gómez; Ramírez-López, María Teresa; Antón, María; Pozo, Miguel Ángel; Rodríguez de Fonseca, Fernando

    2014-11-01

    The administration of selective serotonin reuptake inhibitors (SSRIs) typically used as antidepressants increases alcohol consumption after an alcohol deprivation period in rats. However, the appearance of this effect after the treatment with selective noradrenaline reuptake inhibitors (SNRIs) has not been studied. In the present work we examined the effects of a 15-d treatment with the SNRI atomoxetine (1, 3 and 10 mg/kg, i.p.) in male rats trained to drink alcohol solutions in a 4-bottle choice test. The treatment with atomoxetine (10 mg/kg, i.p.) during an alcohol deprivation period increased alcohol consumption after relapse. This effect only lasted one week, disappearing thereafter. Treatment with atomoxetine did not cause a behavioral sensitized response to a challenge dose of amphetamine (1.5 mg/kg, i.p.), indicating the absence of a supersensitive dopaminergic transmission. This effect is markedly different from that of SSRI antidepressants that produced both long-lasting increases in alcohol consumption and behavioral sensitization. Clinical implications are discussed. PMID:25025529

  14. Prolonged administration of antidepressant drugs leads to increased binding of [(3)H]MPEP to mGlu5 receptors.

    PubMed

    Nowak, Gabriel; Pomierny-Chamioło, Lucyna; Siwek, Agata; Niedzielska, Ewa; Pomierny, Bartosz; Pałucha-Poniewiera, Agnieszka; Pilc, Andrzej

    2014-09-01

    Metabotropic glutamate 5 (mGlu5) receptors are functionally connected with NMDA receptors. The antidepressant activity of the NMDA receptor antagonist ketamine in both preclinical and clinical studies, along with the antidepressant-like activities of negative allosteric modulators (NAMs) of mGlu5, led us to investigate if prolonged administration of various antidepressant drugs or the mGlu5 NAM, MTEP, causes changes in mGlu5 receptor availability or protein expression or in expression of Homer proteins in the rat brain. Our results clearly show that prolonged treatment with antidepressants with various mechanisms of action (such as escitalopram, reboxetine, milnacipran, moclobemide and imipramine) or with MTEP led to significant increases in [(3)H]MPEP binding in homogenates of the hippocampus and/or cerebral cortex. Increases in mGlu5 expression were also observed, though they did not always parallel the increase in binding. The results indicate that adaptive up-regulation of mGlu5 receptors may be a common change induced by antidepressant drugs. PMID:24796254

  15. Perceptual distortions and delusional thinking following ketamine administration are related to increased pharmacological MRI signal changes in the parietal lobe.

    PubMed

    Stone, James; Kotoula, Vasileia; Dietrich, Craige; De Simoni, Sara; Krystal, John H; Mehta, Mitul A

    2015-09-01

    Ketamine produces effects in healthy humans that resemble the positive, negative and cognitive symptoms of schizophrenia. We investigated the effect of ketamine administration on brain activity as indexed by blood-oxygen-level-dependent (BOLD) signal change response, and its relationship to ketamine-induced subjective changes, including perceptual distortion. Thirteen healthy participants volunteered for the study. All underwent a 15-min functional MRI acquisition with a ketamine infusion commencing after 5 min (approx 0.26 mg/kg over 20s followed by an infusion of approx. 0.42 mg/kg/h). Following the scan, participants self-rated ketamine-induced effects using the Psychotomimetic States Inventory. Ketamine led to widespread cortical and subcortical increases in BOLD response (FWE-corrected p < 0.01). Self-rated perceptual distortions and delusional thoughts correlated with increased BOLD response in the paracentral lobule (FWE-corrected p < 0.01). The findings suggest that BOLD increases in parietal cortices reflect ketamine effects on circuits that contribute to its capacity to produce perceptual alterations and delusional interpretations.

  16. Chronic oxytocin administration inhibits food intake, increases energy expenditure, and produces weight loss in fructose-fed obese rhesus monkeys.

    PubMed

    Blevins, James E; Graham, James L; Morton, Gregory J; Bales, Karen L; Schwartz, Michael W; Baskin, Denis G; Havel, Peter J

    2015-03-01

    Despite compelling evidence that oxytocin (OT) is effective in reducing body weight (BW) in diet-induced obese (DIO) rodents, studies of the effects of OT in humans and rhesus monkeys have primarily focused on noningestive behaviors. The goal of this study was to translate findings in DIO rodents to a preclinical translational model of DIO. We tested the hypothesis that increased OT signaling would reduce BW in DIO rhesus monkeys by inhibiting food intake and increasing energy expenditure (EE). Male DIO rhesus monkeys from the California National Primate Research Center were adapted to a 12-h fast and maintained on chow and a daily 15% fructose-sweetened beverage. Monkeys received 2× daily subcutaneous vehicle injections over 1 wk. We subsequently identified doses of OT (0.2 and 0.4 mg/kg) that reduced food intake and BW in the absence of nausea or diarrhea. Chronic administration of OT for 4 wk (0.2 mg/kg for 2 wk; 0.4 mg/kg for 2 wk) reduced BW relative to vehicle by 3.3 ± 0.4% (≈0.6 kg; P < 0.05). Moreover, the low dose of OT suppressed 12-h chow intake by 26 ± 7% (P < 0.05). The higher dose of OT reduced 12-h chow intake by 27 ± 5% (P < 0.05) and 8-h fructose-sweetened beverage intake by 18 ± 8% (P < 0.05). OT increased EE during the dark cycle by 14 ± 3% (P < 0.05) and was associated with elevations of free fatty acids and glycerol and reductions in triglycerides suggesting increased lipolysis. Together, these data suggest that OT reduces BW in DIO rhesus monkeys through decreased food intake as well as increased EE and lipolysis.

  17. Radioligand binding to muscarinic receptors of bovine aortic endothelial cells.

    PubMed

    Brunner, F; Kukovetz, W R

    1991-02-01

    1. Muscarinic receptors on endothelial cells of bovine thoracic aorta were characterized by binding assays in which (-)-[3H]-N-methyl quinuclidinyl benzilate ([3H]-NMeQNB) was used as radioligand. 2. Binding of [3H]-NMeQNB to crude membranes of freshly isolated endothelial cells was atropine-displaceable and of high affinity (KD = 0.48 nM) to a single class of sites (maximum binding capacity: 14 +/- 3 fmol mg-1 protein). Stereospecificity of the binding sites was demonstrated in experiments in which [3H]-NMeQNB binding was inhibited by dexetimide in the nanomolar range (KI = 0.63 nM) and by levetimide, its stereoisomer in the micromolar range (KI = 3.2 microM) (selectivity factor: approximately 5000). 3. Drug competition curves indicated a single class of binding sites for antagonists and the following apparent affinities (KI, nM): methyl atropine: 1.1: 4-diphenylacetoxy N-methyl piperidine methyl bromide (4-DAMP): 3.4; pirenzepine: 16; 11-[2-diethylamino-methyl)-1-piperidinyl- acetyl]-5,11-dihydro-6H-pyrido(2,3-b)1,4-benzodiazepine-6-one (AF-DX 116); 2.500. Competition of acetylcholine with [3H]-NMeQNB was best described by two affinity sites (or states) (KH = 0.82 microM, KL = 1.6 microM). In the presence of guanylimido diphosphate [Gpp(NH)p] (100 microM), acetylcholine affinity (IC50) was slightly, but significantly reduced (factor approximately 4). 4. Binding of [3H]-NMeQNB to freshly harvested intact cells was also atropine-displaceable, stereospecific (selectivity factor: approximately 3500) and of high affinity (KD = 0.35 nM). The maximum binding capacity (9 +/- 2 fmol mg-1 total cell protein) was comparable to that of membranes and corresponded to approximately 900 binding sites per endothelial cell. Binding to enzymatically harvested and cultured endothelial cells, or membranes derived therefrom, showed no atropine-displaceable binding. 5. The results suggest that (1) bovine aortic endothelial cells contain muscarinic binding sites with all necessary

  18. Muscarinic receptor subtypes differentially control synaptic input and excitability of cerebellum-projecting medial vestibular nucleus neurons.

    PubMed

    Zhu, Yun; Chen, Shao-Rui; Pan, Hui-Lin

    2016-04-01

    Neurons in the vestibular nuclei have a vital function in balance maintenance, gaze stabilization, and posture. Although muscarinic acetylcholine receptors (mAChRs) are expressed and involved in regulating vestibular function, it remains unclear how individual mAChR subtypes regulate vestibular neuronal activity. In this study, we determined which specific subtypes of mAChRs control synaptic input and excitability of medial vestibular nucleus (MVN) neurons that project to the cerebellum. Cerebellum-projecting MVN neurons were labeled by a fluorescent retrograde tracer and then identified in rat brainstem slices. Quantitative PCR analysis suggested that M2 and M3 were the possible major mAChR subtypes expressed in the MVN. The mAChR agonist oxotremorine-M significantly reduced the amplitude of glutamatergic excitatory post-synaptic currents evoked by stimulation of vestibular primary afferents, and this effect was abolished by the M2-preferring antagonist AF-DX 116. However, oxotremorine-M had no effect on GABA-mediated spontaneous inhibitory post-synaptic currents of labeled MVN neurons. Furthermore, oxotremorine-M significantly increased the firing activity of labeled MVN neurons, and this effect was blocked by the M3-preferring antagonist J104129 in most neurons tested. In addition, AF-DX 116 reduced the onset latency and prolonged the excitatory effect of oxotremorine-M on the firing activity of labeled MVN neurons. Our findings suggest that M3 is the predominant post-synaptic mAChR involved in muscarinic excitation of cerebellum-projecting MVN neurons. Pre-synaptic M2 mAChR regulates excitatory glutamatergic input from vestibular primary afferents, which in turn influences the excitability of cerebellum-projecting MVN neurons. This new information has important therapeutic implications for treating vestibular disorders with mAChR subtype-selective agents. Medial vestibular nucleus (MVN) neurons projecting to the cerebellum are involved in balance control. We

  19. Muscarinic receptor subtypes differentially control synaptic input and excitability of cerebellum-projecting medial vestibular nucleus neurons.

    PubMed

    Zhu, Yun; Chen, Shao-Rui; Pan, Hui-Lin

    2016-04-01

    Neurons in the vestibular nuclei have a vital function in balance maintenance, gaze stabilization, and posture. Although muscarinic acetylcholine receptors (mAChRs) are expressed and involved in regulating vestibular function, it remains unclear how individual mAChR subtypes regulate vestibular neuronal activity. In this study, we determined which specific subtypes of mAChRs control synaptic input and excitability of medial vestibular nucleus (MVN) neurons that project to the cerebellum. Cerebellum-projecting MVN neurons were labeled by a fluorescent retrograde tracer and then identified in rat brainstem slices. Quantitative PCR analysis suggested that M2 and M3 were the possible major mAChR subtypes expressed in the MVN. The mAChR agonist oxotremorine-M significantly reduced the amplitude of glutamatergic excitatory post-synaptic currents evoked by stimulation of vestibular primary afferents, and this effect was abolished by the M2-preferring antagonist AF-DX 116. However, oxotremorine-M had no effect on GABA-mediated spontaneous inhibitory post-synaptic currents of labeled MVN neurons. Furthermore, oxotremorine-M significantly increased the firing activity of labeled MVN neurons, and this effect was blocked by the M3-preferring antagonist J104129 in most neurons tested. In addition, AF-DX 116 reduced the onset latency and prolonged the excitatory effect of oxotremorine-M on the firing activity of labeled MVN neurons. Our findings suggest that M3 is the predominant post-synaptic mAChR involved in muscarinic excitation of cerebellum-projecting MVN neurons. Pre-synaptic M2 mAChR regulates excitatory glutamatergic input from vestibular primary afferents, which in turn influences the excitability of cerebellum-projecting MVN neurons. This new information has important therapeutic implications for treating vestibular disorders with mAChR subtype-selective agents. Medial vestibular nucleus (MVN) neurons projecting to the cerebellum are involved in balance control. We

  20. Pharmacological characterization of muscarinic receptors in neonatal rat cardiomyocytes.

    PubMed

    Yang, C M; Chen, F F; Sung, T C; Hsu, H F; Wu, D

    1993-09-01

    [N-methyl-3H]scopolamine methylchloride ([3H]NMS) was used to characterize the muscarinic receptors (mAChRs) in the intact cardiomyocytes. The specific binding of [3H]NMS was proportional to cell concentration, saturable with respect to [3H]NMS concentration, and time dependent. Scatchard analysis of binding isotherms showed that [3H]NMS bound to the freshly isolated and cultured cardiomyocytes with dissociation constants of 275 +/- 64 and 207 +/- 20 pM as well as maximum receptor densities of 0.13 +/- 0.09 and 5.36 +/- 0.20 fmol/10(5) cells, respectively. Heterogeneity of mAChRs was demonstrated by competitive binding experiments against [3H]NMS with M2 and M3 antagonists. These receptors (80%) exhibited high affinities for 11-([2-[(diethylamino)methyl]-1-piperidinyl]-acetyl)-5,11-dihydro- 6H-pyrido[2,3-b][1,4]benzodiazepine-6-one (AF-DX-116) and methoctramine similar to those of M2 subtype. The low-affinity M2 antagonist binding constants were close to those reported for M3 receptors and possessed high affinity for 4-diphenylacetoxyl-N-methylpiperidine (4-DAMP) and hexahydrosiladifenidol. On the basis of biochemical studies, AF-DX-116 blocked adenosine 3',5'-cyclic monophosphate (cAMP) inhibition with high affinity (pKB 7.4), while it antagonized inositol phosphate formation with low affinity (pKB 6.5). 4-DAMP possessed high affinity in blocking inositol phosphate formation (pKB 9.0) and low affinity for antagonism of cAMP inhibition (pKB 7.7). Although no other muscarinic receptor mRNA has been detected in these cells, these data suggest the presence of a second population of mAChRs, which may not be identical to the classical cardiac "M2" receptors.

  1. Pharmacological characterization of muscarinic receptors in neonatal rat cardiomyocytes.

    PubMed

    Yang, C M; Chen, F F; Sung, T C; Hsu, H F; Wu, D

    1993-09-01

    [N-methyl-3H]scopolamine methylchloride ([3H]NMS) was used to characterize the muscarinic receptors (mAChRs) in the intact cardiomyocytes. The specific binding of [3H]NMS was proportional to cell concentration, saturable with respect to [3H]NMS concentration, and time dependent. Scatchard analysis of binding isotherms showed that [3H]NMS bound to the freshly isolated and cultured cardiomyocytes with dissociation constants of 275 +/- 64 and 207 +/- 20 pM as well as maximum receptor densities of 0.13 +/- 0.09 and 5.36 +/- 0.20 fmol/10(5) cells, respectively. Heterogeneity of mAChRs was demonstrated by competitive binding experiments against [3H]NMS with M2 and M3 antagonists. These receptors (80%) exhibited high affinities for 11-([2-[(diethylamino)methyl]-1-piperidinyl]-acetyl)-5,11-dihydro- 6H-pyrido[2,3-b][1,4]benzodiazepine-6-one (AF-DX-116) and methoctramine similar to those of M2 subtype. The low-affinity M2 antagonist binding constants were close to those reported for M3 receptors and possessed high affinity for 4-diphenylacetoxyl-N-methylpiperidine (4-DAMP) and hexahydrosiladifenidol. On the basis of biochemical studies, AF-DX-116 blocked adenosine 3',5'-cyclic monophosphate (cAMP) inhibition with high affinity (pKB 7.4), while it antagonized inositol phosphate formation with low affinity (pKB 6.5). 4-DAMP possessed high affinity in blocking inositol phosphate formation (pKB 9.0) and low affinity for antagonism of cAMP inhibition (pKB 7.7). Although no other muscarinic receptor mRNA has been detected in these cells, these data suggest the presence of a second population of mAChRs, which may not be identical to the classical cardiac "M2" receptors. PMID:8214023

  2. In vitro pharmacological properties of 4-bromodexetimide for muscarinic receptors.

    PubMed

    Strijckmans, V; Coulon, C; Kassiou, M; Loc'h, C; Mazière, B

    1996-01-01

    The decrease of m-AChR density observed in neurodegenerative disorders has generated considerable interest in non-invasive mapping of muscarinic acetylcholine receptors (m-AChR) in the central nervous system. The aim of our study was to evaluate the selectivity of 4-bromodexetimide for the M1, M2, M3 and M4 m-AChR subtypes using in vitro binding analysis to determine the potential use of the bromine-76 labelled 4-bromodexetimide in the investigation of m-AChR subtypes in human brain with Positron Emission Tomography. Subtype selectivity of 4-bromodexetimide was determined in competition studies against tritiated subtype selective ligands using various rat or rabbit structure homogenates reflecting a single binding site and in optimal saturation and low non specific binding conditions. These conditions were reached for every subtype studied by analyzing the data from the saturation experiments of the tritiated ligands. 4-bromodexetimide displayed nanomolar affinities for the four m-AChR subtypes and a preferential selectivity for the M1 and M4 subtypes. The saturation analysis of [76Br]4-bromodexetimide, performed with rat cortex membranes showed high affinity for m-AChR receptors (Kd = 1.8 nM). As in vivo studies of [76Br]4-bromodexetimide showed preferential localization in the cortex and the striatum which are M1 and M4 rich structures and since it binds preferentially to the M1 and M4 subtypes, this radiotracer can still allow a combined subtype specific measurement of these muscarinic receptors. PMID:8649190

  3. A human embryonic lung fibroblast with a high density of muscarinic acetylcholine receptors.

    PubMed

    André, C; Marullo, S; Convents, A; Lü, B Z; Guillet, J G; Hoebeke, J; Strosberg, D A

    1988-01-15

    Binding studies with the radiolabeled muscarinic antagonists dexetimide, quinuclidinyl benzilate and N-methylscopolamine showed that the human embryonic lung fibroblast CCL137 possesses approximately 2 X 10(5) muscarinic receptors/cell, i.e. 2.1 pmol/mg membrane protein. These receptors showed a marked stereoselectivity towards dexetimide and levetimide and only low affinity for another antagonist, pirenzepine. The muscarinic agonist carbamylcholine inhibited forskolin-stimulated adenylate cyclase and induced phosphatidylinositide turnover in the intact cells. Both effects were inhibited by the muscarinic antagonist atropine. Affinity labeling with tritiated propylbenzylcholine mustard revealed a protein of 72 kDa. Finally, down-regulation of the membrane receptors following prolonged treatment with the agonist carbamylcholine was assessed by means of the hydrophilic antagonist N-methylscopolamine. PMID:2828056

  4. Molecular regulation of receptors: interaction of beta-estradiol and progesterone with the muscarinic system.

    PubMed Central

    Sokolovsky, M; Egozi, Y; Avissar, S

    1981-01-01

    The effects of various substrates on the binding of agonists to muscarinic receptors were studied in the rat hypothalamus and adenohypophysis by competition experiments using the highly specific tritiated muscarinic antagonist N-methyl-4-piperidyl benzilate. It was found that agonist binding properties were affected only by the steroid sex hormones (beta-estradiol and progesterone), both of which resulted in a decrease in the proportion of high-affinity binding sites and a decrease in the dissociation constant. This suggests a link between the muscarinic system and the mechanism by which these steroids exert their gonadotropin-releasing effect on the adenohypophysis. We propose a model to depict the putative relationship between the muscarinic system and other receptor systems, including that which controls the steroid sex hormones. PMID:6946492

  5. Administration of a non-NMDA antagonist, GYKI 52466, increases excitotoxic Purkinje cell degeneration caused by ibogaine.

    PubMed

    O'Hearn, E; Molliver, M E

    2004-01-01

    Ibogaine is a tremorigenic hallucinogen that has been proposed for clinical use in treating addiction. We previously reported that ibogaine, administered systemically, produces degeneration of a subset of Purkinje cells in the cerebellum, primarily within the vermis. Ablation of the inferior olive affords protection against ibogaine-induced neurotoxicity leading to the interpretation that ibogaine itself is not directly toxic to Purkinje cells. We postulated that ibogaine produces sustained excitation of inferior olivary neurons that leads to excessive glutamate release at climbing fiber terminals, causing subsequent excitotoxic injury to Purkinje cells. The neuronal degeneration induced by ibogaine provides an animal model for studying excitotoxic injury in order to analyze the contribution of glutamate receptors to this injury and to evaluate neuroprotective strategies. Since non-N-methyl-D-aspartate (NMDA) receptors mediate Purkinje cell excitation by climbing fibers, we hypothesized that 1-4-aminophenyl-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine (GYKI-52466), which antagonizes non-NMDA receptors, may have a neuroprotective effect by blocking glutamatergic excitation at climbing fiber synapses. To test this hypothesis, rats were administered systemic ibogaine plus GYKI-52466 and the degree of neuronal injury was analyzed in cerebellar sections. The results indicate that the AMPA antagonist GYKI-52466 (10 mg/kg i.p. x 3) does not protect against Purkinje cell injury at the doses used. Rather, co-administration of GYKI-52466 with ibogaine produces increased toxicity evidenced by more extensive Purkinje cell degeneration. Several hypotheses that may underlie this result are discussed. Although the reason for the increased toxicity found in this study is not fully explained, the present results show that a non-NMDA antagonist can produce increased excitotoxic injury under some conditions. Therefore, caution should be exercised before employing glutamate

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

    SciTech Connect

    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.

  7. Ozone-induced changes in muscarinic bronchial reactivity by different testing methods.

    PubMed

    Roum, J H; Murlas, C

    1984-12-01

    We examined the effect of ozone (O3) on muscarinic bronchial reactivity in the guinea pig and compared reactivity determined by two different routes of agonist delivery. Reactivity before and from 4 h to 2 days after O3 exposure (3.0 ppm, 2 h) was determined by measuring specific airway resistance upon administration of intravenous acetylcholine and/or aerosolized methacholine challenge in 34 unanesthetized, spontaneously breathing animals. Before exposure, we observed more gradual and reproducible results to intravenous agonist. After exposure, hyperreactivity to parenteral agonist occurred consistently, but not to inhaled agonist. Hyperreactivity demonstrable by either route was similar in magnitude and time course within 14 h of exposure. Two days later, hyperreactivity to inhaled agonist had remitted; that to intravenous drug persisted. Our results indicate that variability in the occurrence and time course of O3-induced hyperreactivity to inhaled agonist may be a consequence of the technique employed. The consistent occurrence of hyperreactivity after O3 to parenteral agonist suggests mechanisms other than airway mucosal hyperpermeability are responsible for this hyperreactivity.

  8. Assay of muscarinic acetylcholine receptor function in cultured cardiac cells by stimulation of /sup 86/Rb+ efflux

    SciTech Connect

    Hunter, D.D.; Nathanson, N.M.

    1985-09-01

    An assay for the increase in potassium permeability mediated by muscarinic acetylcholine receptors (mAChR) in cultured cardiac cells is described, using the K+ ion substitute /sup 86/Rb+ as the tracer ion. Cardiac cells accumulate /sup 86/Rb+ from the extracellular medium in a Na+/K+ ATPase-dependent manner. Subsequent efflux of /sup 86/Rb+ in the absence and presence of muscarinic agonists follows kinetics similar to those previously reported for /sup 42/K+. The mAChR agonist carbamylcholine (carbachol) stimulated /sup 86/Rb+ efflux with an EC50 of 50 nM. The half-time for efflux is reduced by greater than 40% at maximally effective concentrations of agonist. Stimulation of /sup 86/Rb+ efflux by carbachol is blocked by the mAChR antagonist atropine with an IC50 of 15 nM. The stimulation of 86Rb+ efflux by carbachol is not affected by the presence of the Na+/K+ ATPase inhibitor ouabain. This assay provides a method for quantitating the mAChR-mediated increase in K+ permeability in cardiac cells without the use of /sup 42/K+.

  9. Activation of alpha adrenergic and muscarinic receptors modifies early glucose suppression of cytoplasmic Ca(2+) in pancreatic β-cells.

    PubMed

    Hellman, Bo; Dansk, Heléne; Grapengiesser, Eva

    2014-03-14

    Elevation of glucose induces transient inhibition of insulin release by lowering cytoplasmic Ca(2+) ([Ca(2+)]i) below baseline in pancreatic β-cells. The period of [Ca(2+)]i decrease (phase 0) coincides with increased glucagon release and is therefore the starting point for antisynchronous pulses of insulin and glucagon. We now examine if activation of adrenergic α2A and muscarinic M3 receptors affects the initial [Ca(2+)]i response to increase of glucose from 3 to 20mM in β-cells situated in mouse islets. In the absence of receptor stimulation the elevation of glucose lowered [Ca(2+)]i during 90-120 s followed by rise due to opening of voltage-dependent Ca(2+) channels. The period of [Ca(2+)]i decrease was prolonged by activation of the α2A adrenergic receptors (1 μM epinephrine or 100 nM clonidine) and shortened by stimulation of the muscarinic M3 receptors (0.1 μM acetylcholine). The latter effect was mimicked by the Na/K pump inhibitor ouabain (10-100 μM). The results indicate that prolonged initial decrease (phase 0) is followed by slow [Ca(2+)]i rise and shorter decrease followed by fast rise. It is concluded that the period of initial decrease of [Ca(2+)]i regulates the subsequent β-cell response to glucose.

  10. Subchronic administration of riparin III induces antidepressive-like effects and increases BDNF levels in the mouse hippocampus.

    PubMed

    Vasconcelos, Auriana S; Oliveira, Iris C M; Vidal, Laura T M; Rodrigues, Gabriel C; Gutierrez, Stanley J C; Barbosa-Filho, José M; Vasconcelos, Silvânia M M; de França Fonteles, Marta M; Gaspar, Danielle M; de Sousa, Francisca C F

    2015-08-01

    Riparin III (Rip III) is an alcamide isolated from Aniba riparia that has presented effects of antidepressant and anxiolytic activities in acute stress behavioral models. The trial's goal was to investigate the activity of Rip III in mice exposed to corticosterone-induced chronic depression model. Swiss female mice, 22-25 g, were distributed in following experimental groups: control group (vehicle1: saline containing 0.1% dimethyl sulfoxide and 0.1% Tween-80, SC+ vehicle 2: distilled water emulsified with 2% Tween-80, PO); stressed group (corticosterone, 20 mg/kg, SC, + vehicle 2, orally); Rip III group (50 mg/kg, orally); and fluvoxamine (Flu) group (50 mg/kg, orally). The mice were exposed to the behavioral tests, and posteriorly, Brain-derived neurotrophic factor protein levels were assessed in hippocampal samples. Statistical analysis of the data was performed by one-way anova, followed by Newman-Keuls test. Both administrations of Rip III and Flu significantly reduced the immobility time in tail suspension and forced swimming tests after 21 days without affecting locomotor function. There was also an increase in BDNF protein levels in the mice hippocampus. These findings further support the hypothesis that Rip III could be a new pharmacological target for the treatment of mood disorders.

  11. Amyloid beta-peptide disrupts carbachol-induced muscarinic cholinergic signal transduction in cortical neurons.

    PubMed

    Kelly, J F; Furukawa, K; Barger, S W; Rengen, M R; Mark, R J; Blanc, E M; Roth, G S; Mattson, M P

    1996-06-25

    Cholinergic pathways serve important functions in learning and memory processes, and deficits in cholinergic transmission occur in Alzheimer disease (AD). A subset of muscarinic cholinergic receptors are linked to G-proteins that activate phospholipase C, resulting in the liberation of inositol trisphosphate and Ca2+ release from intracellular stores. We now report that amyloid beta-peptide (Abeta), which forms plaques in the brain in AD, impairs muscarinic receptor activation of G proteins in cultured rat cortical neurons. Exposure of rodent fetal cortical neurons to Abeta25-35 and Abeta1-40 resulted in a concentration and time-dependent attenuation of carbachol-induced GTPase activity without affecting muscarinic receptor ligand binding parameters. Downstream events in the signal transduction cascade were similarly attenuated by Abeta. Carbachol-induced accumulation of inositol phosphates (IP, IP2, IP3, and IP4) was decreased and calcium imaging studies revealed that carbachol-induced release of calcium was severely impaired in neurons pretreated with Abeta. Muscarinic cholinergic signal transduction was disrupted with subtoxic levels of exposure to AP. The effects of Abeta on carbachol-induced GTPase activity and calcium release were attenuated by antioxidants, implicating free radicals in the mechanism whereby Abeta induced uncoupling of muscarinic receptors. These data demonstrate that Abeta disrupts muscarinic receptor coupling to G proteins that mediate induction of phosphoinositide accumulation and calcium release, findings that implicate Abeta in the impairment of cholinergic transmission that occurs in AD. PMID:8692890

  12. Alterations of muscarinic receptor subtypes in pathways relating to memory: Effects of lesions and transplants

    SciTech Connect

    Dawson, V.L.

    1989-01-01

    Muscarinic cholinergic receptors have been classified pharmacologically into two distinct populations designated muscarinic type-one (M-1) and mscarinic type-two (M-2). The semiquantitative technique of receptor autoradiography was used to examine the anatomical and cellular distribution, and densities of M-1 and M-2 receptors in the rate brain. Muscarinic receptors were labeled with the classical antagonist ({sup 3}H)quinuclidinyl benzilate (QNB). Differentiation of the muscarinic subtypes was accomplished by competition studies of ({sup 3}H)QNB against the relatively selective M-1 antagonist pirenzepine (PZ), and the relatively selective M-2 antagonist, AFDX-116. In addition, M-1 and M-2 receptors were directly labeled with ({sup 3}H)PZ and ({sup 3}H)AFDX-116, respectively. Cholinergic pathways from the large cholinergic neurons in the nucleus basalis magnocellularis (NBM) to the cortex and from the medial septum (MS) to the hippocampus were examined by lesioning with the selective cholinergic neurotoxin, AF64A. Bilateral cerebral cortical infarction was performed in order to analyze potential changes in muscarinic receptor populations in subcortical structures that are sensitive to cortical infarction. Finally, the response of muscarinic receptors to fetal septodiagonal band transplants in the deafferentated hippocampus was examined.

  13. Muscarinic antagonists in the treatment of acquired pendular and downbeat nystagmus: a double-blind, randomized trial of three intravenous drugs.

    PubMed

    Barton, J J; Huaman, A G; Sharpe, J A

    1994-03-01

    We performed a double-blind, randomized trial of intravenous scopolamine, benztropine, and glycopyrrolate in 7 patients with acquired nystagmus and oscillopsia. Five patients had pendular nystagmus and 2, downbeat nystagmus. We recorded eye movements with a magnetic search coil technique and tested visual acuity and motion perception before and after administration of each drug. Scopolamine reduced nystagmus in all patients. Benztropine was moderately effective and glycopyrrolate had a negligible impact. Visual acuity improved only with scopolamine; motion discrimination and oscillopsia improved significantly with scopolamine and benztropine. Pendular and downbeat nystagmus respond to intravenous antagonists of central muscarinic receptors.

  14. Chronic administration of branched-chain amino acids impairs spatial memory and increases brain-derived neurotrophic factor in a rat model.

    PubMed

    Scaini, Giselli; Comim, Clarissa M; Oliveira, Giovanna M T; Pasquali, Matheus A B; Quevedo, João; Gelain, Daniel P; Moreira, José Cláudio F; Schuck, Patrícia F; Ferreira, Gustavo C; Bogo, Maurício R; Streck, Emilio L

    2013-09-01

    Maple syrup urine disease (MSUD) is a neurometabolic disorder that leads to the accumulation of branched-chain amino acids (BCAAs) and their α-keto branched-chain by-products. Because the neurotoxic mechanisms of MSUD are poorly understood, this study aimed to evaluate the effects of chronic administration of a BCAA pool (leucine, isoleucine and valine). This study examined the effects of BCAA administration on spatial memory and the levels of brain-derived neurotrophic factor (BNDF). We examined both pro-BDNF and bdnf mRNA expression levels after administration of BCAAs. Furthermore, this study examined whether antioxidant treatment prevented the alterations induced by BCAA administration. Our results demonstrated an increase in BDNF in the hippocampus and cerebral cortex, accompanied by memory impairment in spatial memory tasks. Additionally, chronic administration of BCAAs did not induce a detectable change in pro-BDNF levels. Treatment with N-acetylcysteine and deferoxamine prevented both the memory deficit and the increase in the BDNF levels induced by BCAA administration. In conclusion, these results suggest that when the brain is chronically exposed to high concentrations of BCAA (at millimolar concentrations) an increase in BDNF levels occurs. This increase in BDNF may be related to the impairment of spatial memory. In addition, we demonstrated that antioxidant treatment prevented the negative consequences related to BCAA administration, suggesting that oxidative stress might be involved in the pathophysiological mechanism(s) underlying the brain damage observed in MSUD. PMID:23109061

  15. Prostaglandins and muscarinic agonists induce cyclic AMP attenuation by two distinct mechanisms in the pregnant-rat myometrium. Interaction between cyclic AMP and Ca2+ signals.

    PubMed Central

    Goureau, O; Tanfin, Z; Harbon, S

    1990-01-01

    In pregnant-rat myometrium (day 21 of gestation), isoprenaline-induced cyclic AMP accumulation, resulting from receptor-mediated activation of adenylate cyclase, was negatively regulated by prostaglandins [PGE2, PGF2 alpha; EC50 (concn. giving 50% of maximal response) = 2 nM] and by the muscarinic agonist carbachol (EC50 = 2 microM). PG-induced inhibition was prevented by pertussis-toxin treatment, supporting the idea that it was mediated by the inhibitory G-protein Gi through the inhibitory pathway of the adenylate cyclase. Both isoprenaline-induced stimulation and PG-evoked inhibition of cyclic AMP were insensitive to Ca2+ depletion. By contrast, carbachol-evoked attenuation of cyclic AMP accumulation was dependent on Ca2+ and was insensitive to pertussis toxin. The inhibitory effect of carbachol was mimicked by ionomycin. Indirect evidence was thus provided for the enhancement of cyclic AMP degradation by a Ca2(+)-dependent phosphodiesterase activity in the muscarinic-mediated effect. The attenuation of cyclic AMP elicited by carbachol coincided with carbachol-stimulated inositol phosphate (InsP3, InsP2 and InsP) generation, which displayed an almost identical EC50 (3 microM) and was similarly unaffected by pertussis toxin. Both carbachol effects were reproduced by oxotremorine, whereas pilocarpine (a partial muscarinic agonist) failed to induce any decrease in cyclic AMP accumulation and concurrently was unable to stimulate the generation of inositol phosphates. These data support our proposal for a carbachol-mediated enhancement of a Ca2(+)-dependent phosphodiesterase activity, compatible with the rises in Ca2+ associated with muscarinic-induced increased generation of inositol phosphates. They further illustrate that a cross-talk between the two major transmembrane signalling systems contributed to an ultimate decrease in cyclic AMP in the pregnant-rat myometrium near term. PMID:1700899

  16. Increasing Coverage in Mass Drug Administration for Lymphatic Filariasis Elimination in an Urban Setting: a Study of Malindi Town, Kenya

    PubMed Central

    Njomo, Doris W.; Mukoko, Dunstan A.; Nyamongo, Nipher K.; Karanja, Joan

    2014-01-01

    Introduction Implementation of Mass Drug Administration (MDA) in urban settings is an obstacle to Lymphatic Filariasis (LF) elimination. No urban-specific guidelines on MDA in urban areas exist. Malindi district urban area had received 4 MDA rounds by the time the current study was implemented. Programme data showed average treatment coverage of 28.4% (2011 MDA), far below recommended minimum of 65–80%. Methods To identify, design and test strategies for increased treatment coverage in urban areas, a quasi-experimental study was conducted in Malindi urban area. Three sub-locations with lowest treatment coverage in 2011 MDA were purposively selected. In the pre-test phase, 947 household heads sampled using systematic random method were interviewed for quantitative data. For qualitative data, 12 Focus Group Discussions (FGDs) with single sex adult and youth male and female groups and 3 with community drug distributors (CDDs) were conducted. Forty in-depth interviews with opinion leaders and self-administered questionnaires with District Public Health officers purposively selected were carried out. The quantitative data were analyzed using SPSS version 16 and statistical significance assessed by χ2 test.The qualitative data were analyzed manually according to study's themes. Results and Discussion The identified strategies were implemented prior to and during 2012 MDA in two sub-locations (experimental) while in the third (control), usual MDA strategies were applied. In the post-test phase, 2012 MDA coverage in experimental and control sub-locations was comparatively assessed for effect of the newly designed strategies on urban MDA. Results indicated improved treatment coverage in experimental sub-locations, 77.1% in Shella and 66.0% in Barani. Central (control) sub-location also attained high coverage, 70.4% indicating average treatment coverage of 71%. Conclusion The identified strategies contributed to increased treatment coverage in experimental sites and

  17. Involvement of Ca2+ Signaling in the Synergistic Effects between Muscarinic Receptor Antagonists and β2-Adrenoceptor Agonists in Airway Smooth Muscle

    PubMed Central

    Fukunaga, Kentaro; Kume, Hiroaki; Oguma, Tetsuya; Shigemori, Wataru; Tohda, Yuji; Ogawa, Emiko; Nakano, Yasutaka

    2016-01-01

    Long-acting muscarinic antagonists (LAMAs) and short-acting β2-adrenoceptor agonists (SABAs) play important roles in remedy for COPD. To propel a translational research for development of bronchodilator therapy, synergistic effects between SABAs with LAMAs were examined focused on Ca2+ signaling using simultaneous records of isometric tension and F340/F380 in fura-2-loaded tracheal smooth muscle. Glycopyrronium (3 nM), a LAMA, modestly reduced methacholine (1 μM)-induced contraction. When procaterol, salbutamol and SABAs were applied in the presence of glycopyrronium, relaxant effects of these SABAs are markedly enhanced, and percent inhibition of tension was much greater than the sum of those for each agent and those expected from the BI theory. In contrast, percent inhibition of F340/F380 was not greater than those values. Bisindolylmaleimide, an inhibitor of protein kinase C (PKC), significantly increased the relaxant effect of LAMA without reducing F340/F380. Iberiotoxin, an inhibitor of large-conductance Ca2+-activated K+ (KCa) channels, significantly suppressed the effects of these combined agents with reducing F340/F380. In conclusion, combination of SABAs with LAMAs synergistically enhances inhibition of muscarinic contraction via decreasing both Ca2+ sensitization mediated by PKC and Ca2+ dynamics mediated by KCa channels. PKC and KCa channels may be molecular targets for cross talk between β2-adrenoceptors and muscarinic receptors. PMID:27657061

  18. Dorsal raphe nucleus acetylcholine-mediated neurotransmission modulates post-ictal antinociception: The role of muscarinic and nicotinic cholinergic receptors.

    PubMed

    de Oliveira, Rithiele Cristina; de Oliveira, Ricardo; Biagioni, Audrey Francisco; Falconi-Sobrinho, Luiz Luciano; Coimbra, Norberto Cysne

    2016-01-15

    The dorsal raphe nucleus (DRN) is a key structure of the endogenous pain inhibitory system. Although the DRN is rich in serotoninergic neurons, cholinergic neurons are also found in that nucleus. Both ictal and inter-ictal states are followed by post-ictal analgesia. The present study investigated the role of cholinergic mechanisms in postictal antinociceptive processes using microinjections of atropine and mecamylamine, muscarinic and nicotinic cholinergic receptor antagonists, respectively, in the DRN of rats. Intraperitoneal injection of pentylenetetrazole (PTZ) (at 64mg/kg) caused tonic and tonic-clonic seizures. The convulsive motor reactions were followed by an increase in pain thresholds, a phenomenon known as post-ictal analgesia. Pre-treatment of the DRN with atropine or mecamylamine at 1µg, 3µg and 5µg/0.2µL decreased the post-ictal antinociceptive phenomenon. The present results showed that the post-ictal analgesia was mediated by muscarinic and nicotinic cholinergic receptors in the DRN, a structure crucially involved in the neural network that organises post-ictal hypoalgesia. PMID:26620541

  19. Bilateral injection of fasciculin into the amygdala of rats: effects on two avoidance tasks, acetylcholinesterase activity, and cholinergic muscarinic receptors.

    PubMed

    Quillfeldt, J; Raskovsky, S; Dalmaz, C; Dias, M; Huang, C; Netto, C A; Schneider, F; Izquierdo, I; Medina, J H; Silveira, R

    1990-11-01

    These experiments examined the effects of the bilateral injection of fasciculin-2 (FAS), a natural acetylcholinesterase (AChE) inhibitory peptide, into the amygdala of rats on acquisition and retention of two avoidance behaviors. Intraamygdala injection of FAS (150 ng/amygdala) produced a pronounced and long-lasting inhibition of AChE activity: 85% and 74% on day 2 and day 5, respectively. After 48 hr, FAS-treated animals showed no changes in training or test session performance in a step-down inhibitory avoidance task (training-test interval was 24 hr). In a 2-way shuttle avoidance task, intraamygdala FAS slightly reduced retention test performance without modifying training session scores. Two and five days after FAS injections into the amygdala, the density of muscarinic receptor decreased about 50% as measured by the specific bindings of 3H-quinuclidinyl benzilate and 3H-oxotremorine. No alterations were observed in the apparent dissociation constants. On the other hand, the central-type benzodiazepine receptor population of the amygdala remained unchanged, suggesting that FAS microinjection did not produce damage to neuronal components of these nuclei. In conclusion, the results presented have indicated that a clear-cut and long-lasting inhibition of AChE activity in the amygdala is not accompanied by a facilitation of learning and memory of two different avoidance tasks. Compensation of the increased cholinergic activity by a down-regulation of muscarinic receptors could account for these findings.

  20. Increased Orbitofrontal Brain Activation after Administration of a Selective Adenosine A2A Antagonist in Cocaine Dependent Subjects

    PubMed Central

    Moeller, F. Gerard; Steinberg, Joel L.; Lane, Scott D.; Kjome, Kimberly L.; Ma, Liangsuo; Ferre, Sergi; Schmitz, Joy M.; Green, Charles E.; Bandak, Stephen I.; Renshaw, Perry F.; Kramer, Larry A.; Narayana, Ponnada A.

    2012-01-01

    Background: Positron Emission Tomography imaging studies provide evidence of reduced dopamine function in cocaine dependent subjects in the striatum, which is correlated with prefrontal cortical glucose metabolism, particularly in the orbitofrontal cortex. However, whether enhancement of dopamine in the striatum in cocaine dependent subjects would be associated with changes in prefrontal cortical brain activation is unknown. One novel class of medications that enhance dopamine function via heteromer formation with dopamine receptors in the striatum is the selective adenosine A2A receptor antagonists. This study sought to determine the effects administration of the selective adenosine A2A receptor antagonist SYN115 on brain function in cocaine dependent subjects. Methodology/Principle Findings: Twelve cocaine dependent subjects underwent two fMRI scans (one after a dose of placebo and one after a dose of 100 mg of SYN115) while performing a working memory task with three levels of difficulty (3, 5, and 7 digits). fMRI results showed that for 7-digit working memory activation there was significantly greater activation from SYN115 compared to placebo in portions of left (L) lateral orbitofrontal cortex, L insula, and L superior and middle temporal pole. Conclusion/Significance: These findings are consistent with enhanced dopamine function in the striatum in cocaine dependent subjects via blockade of adenosine A2A receptors producing increased brain activation in the orbitofrontal cortex and other cortical regions. This suggests that at least some of the changes in brain activation in prefrontal cortical regions in cocaine dependent subjects may be related to altered striatal dopamine function, and that enhancement of dopamine function via adenosine A2A receptor blockade could be explored further for amelioration of neurobehavioral deficits associated with chronic cocaine use. PMID:22654774

  1. Blockade of muscarinic receptors impairs the retrieval of well-trained memory

    PubMed Central

    Soma, Shogo; Suematsu, Naofumi; Shimegi, Satoshi

    2014-01-01

    Acetylcholine (ACh) is known to play an important role in memory functions, and its deficit has been proposed to cause the cognitive decline associated with advanced age and Alzheimer's disease (the cholinergic hypothesis). Although many studies have tested the cholinergic hypothesis for recently acquired memory, only a few have investigated the role of ACh in the retrieval process of well-trained cognitive memory, which describes the memory established from repetition and daily routine. To examine this point, we trained rats to perform a two-alternative forced-choice visual detection task. Each trial was started by having the rats pull upward a central-lever, which triggered the presentation of a visual stimulus to the right or left side of the display monitor, and then pulling upward a stimulus-relevant choice-lever located on both sides. Rats learned the task within 10 days, and the task training was continued for a month. Task performance was measured with or without systemic administration of a muscarinic ACh receptor (mAChR) antagonist, scopolamine (SCOP), prior to the test. After 30 min of SCOP administration, rats stopped manipulating any lever even though they explored the lever and surrounding environment, suggesting a loss of the task-related associative memory. Three hours later, rats were recovered to complete the trial, but the rats selected the levers irrespective of the visual stimulus, suggesting they remembered a series of lever-manipulations in association with a reward, but not association between the reward and visual stimulation. Furthermore, an m1-AChR, but not nicotinic AChR antagonist caused a similar deficit in the task execution. SCOP neither interfered with locomotor activity nor drinking behavior, while it influenced anxiety. These results suggest that the activation of mAChRs at basal ACh levels is essential for the recall of well-trained cognitive memory. PMID:24782760

  2. Activation of Muscarinic M1 Acetylcholine Receptors Induces Long-Term Potentiation in the Hippocampus.

    PubMed

    Dennis, Siobhan H; Pasqui, Francesca; Colvin, Ellen M; Sanger, Helen; Mogg, Adrian J; Felder, Christian C; Broad, Lisa M; Fitzjohn, Steve M; Isaac, John T R; Mellor, Jack R

    2016-01-01

    Muscarinic M1 acetylcholine receptors (M1Rs) are highly expressed in the hippocampus, and their inhibition or ablation disrupts the encoding of spatial memory. It has been hypothesized that the principal mechanism by which M1Rs influence spatial memory is by the regulation of hippocampal synaptic plasticity. Here, we use a combination of recently developed, well characterized, selective M1R agonists and M1R knock-out mice to define the roles of M1Rs in the regulation of hippocampal neuronal and synaptic function. We confirm that M1R activation increases input resistance and depolarizes hippocampal CA1 pyramidal neurons and show that this profoundly increases excitatory postsynaptic potential-spike coupling. Consistent with a critical role for M1Rs in synaptic plasticity, we now show that M1R activation produces a robust potentiation of glutamatergic synaptic transmission onto CA1 pyramidal neurons that has all the hallmarks of long-term potentiation (LTP): The potentiation requires NMDA receptor activity and bi-directionally occludes with synaptically induced LTP. Thus, we describe synergistic mechanisms by which acetylcholine acting through M1Rs excites CA1 pyramidal neurons and induces LTP, to profoundly increase activation of CA1 pyramidal neurons. These features are predicted to make a major contribution to the pro-cognitive effects of cholinergic transmission in rodents and humans. PMID:26472558

  3. Identification of drugs competing with d-tubocurarine for an allosteric site on cardiac muscarinic receptors.

    PubMed

    Waelbroeck, M

    1994-10-01

    d-Tubocurarine behaved as a weak allosteric inhibitor of N-[3H] methylscopolamine binding to cardiac M2 muscarinic receptors. In a low ionic strength buffer devoid of bivalent ions, d-tubocurarine recognized cardiac M2 receptors in the micromolar concentration range and decreased their affinity for N-[3H]methylscopolamine by at most 4-fold. To identify the compounds that preferentially recognize this accessory site (as opposed to the classical muscarinic binding site), we measured the inhibition by different drugs of N-[3H]methylscopolamine binding, in the absence or presence of d-tubocurarine. The effect of gallamine was competitively inhibited by d-tubocurarine; both drugs compete for the same accessory site on muscarinic receptors. The effects of dexetimide, levetimide, 4-diphenylacetoxy-N-ethylpiperidine ethobromide, AF-DX 116, and telenzepine on N-[3H]methylscopolamine binding were not affected or were barely affected by d-tubocurarine; these compounds preferentially recognize another binding site (probably the muscarinic binding site). The dose-effect curves for pentamethylene-bis(4-diphenylacetoxymethylpiperidine) bromide and methoctramine were shifted, but at most 10-fold, by d-tubocurarine. It is likely that (in this low ionic strength incubation buffer) methoctramine and pentamethylene-bis(4-diphenylacetoxymethylpiperidine)bromide had comparable affinities for the muscarinic site and the accessory site. d-Tubocurarine competitively inhibited their binding to the accessory site and allosterically inhibited their binding to the muscarinic site. This resulted in a large decrease (40-60-fold) of their overall affinity for muscarinic receptors. PMID:7969047

  4. Autoradiographic visualization of muscarinic receptor subtypes in human and guinea pig lung

    SciTech Connect

    Mak, J.C.; Barnes, P.J. )

    1990-06-01

    Muscarinic receptor subtypes have been localized in human and guinea pig lung sections by an autoradiographic technique, using (3H)(-)quinuclidinyl benzilate (( 3H)QNB) and selective muscarinic antagonists. (3H)QNB was incubated with tissue sections for 90 min at 25 degrees C, and nonspecific binding was determined by incubating adjacent serial sections in the presence of 1 microM atropine. Binding to lung sections had the characterization expected for muscarinic receptors. Autoradiography revealed that muscarinic receptors were widely distributed in human lung, with dense labeling over submucosal glands and airway ganglia, and moderate labeling over nerves in intrapulmonary bronchi and of airway smooth muscle of large and small airways. In addition, alveolar walls were uniformly labeled. In guinea pig lung, labeling of airway smooth muscle was similar, but in contrast to human airways, epithelium was labeled but alveolar walls were not. The muscarinic receptors of human airway smooth muscle from large to small airways were entirely of the M3-subtype, whereas in guinea pig airway smooth muscle, the majority were the M3-subtype with a very small population of the M2-subtype present. In human bronchial submucosal glands, M1- and M3-subtypes appeared to coexist in the proportions of 36 and 64%, respectively. In human alveolar walls the muscarinic receptors were entirely of the M1-subtype, which is absent from the guinea pig lung. No M2-receptors were demonstrated in human lung. The localization of M1-receptors was confirmed by direct labeling with (3H)pirenzepine. With the exception of the alveolar walls in human lung, the localization of muscarinic receptor subtypes on structures in the lung is consistent with known functional studies.

  5. Increased locomotor response to novelty and propensity to intravenous amphetamine self-administration in adult offspring of stressed mothers.

    PubMed

    Deminière, J M; Piazza, P V; Guegan, G; Abrous, N; Maccari, S; Le Moal, M; Simon, H

    1992-07-17

    It is suggested that drug addiction is more likely to develop in individuals who are particularly sensitive to the reinforcing effects of drugs. Animal studies of intravenous drug self-administration (SA) have shown that rats display a large range of individual differences in the propensity to develop drug-seeking. Predisposed animals are characterized by a higher locomotor reactivity to both novelty and psychostimulants. In this report, we show that prenatal stress (restraint of the mother during the last week of pregnancy) may contribute to an individual's vulnerability to develop amphetamine self-administration. The adult offspring of stressed mothers exhibited: (i) a higher locomotor response to novelty and to an injection of amphetamine (0.3 mg/kg, i.v.); (ii) a higher level of amphetamine self-administration. The data indicate that individual predisposition to drug-seeking in the adult may be induced by prenatal events. PMID:1511342

  6. Oral administration of MSG increases expression of glutamate receptors and transporters in the gastrointestinal tract of young piglets.

    PubMed

    Zhang, Jun; Yin, Yulong; Shu, Xu Gang; Li, Tiejun; Li, Fengna; Tan, Bie; Wu, Zhenlong; Wu, Guoyao

    2013-11-01

    Glutamate receptors and transporters, including T1R1 and T1R3 (taste receptor 1, subtypes 1 and 3), mGluRs (metabotropic glutamate receptors), EAAC-1 (excitatory amino acid carrier-1), GLAST-1 (glutamate-aspartate transporter-1), and GLT-1 (glutamate transporter-1), are expressed in the gastrointestinal tract. This study determined effects of oral administration of monosodium glutamate [MSG; 0, 0.06, 0.5, or 1 g/kg body weight (BW)/day] for 21 days on expression of glutamate receptors and transporters in the stomach and jejunum of sow-reared piglets. Both mRNA and protein levels for gastric T1R1, T1R3, mGluR1, mGluR4, EAAT1, EAAT2, EAAT3, and EAAT4 and mRNA levels for jejunal T1R1, T1R3, EAAT1, EAAT2, EAAT3 and EAAT4 were increased (P < 0.05) by MSG supplementation. Among all groups, mRNA levels for gastric EAAT1, EAAT2, EAAT3, and EAAT4 were highest (P < 0.05) in piglets receiving 1 g MSG/kg BW/day. EAAT1 and EAAT2 mRNA levels in the stomach and jejunum of piglets receiving 0.5 g MSG/kg BW/day, as well as jejunal EAAT3 and EAAT4 mRNA levels in piglets receiving 1 g MSG/kg BW/day, were higher (P < 0.05) than those in the control and in piglets receiving 0.06 g MSG/kg BW/day. Furthermore, protein levels for jejunal T1R1 and EAAT3 were higher (P < 0.05) in piglets receiving 1 g MSG/kg BW/day than those in the control and in piglets receiving 0.06 g MSG/kg BW/day. Collectively, these findings indicate that dietary MSG may beneficially stimulate glutamate signaling and sensing in the stomach and jejunum of young pigs, as well as their gastrointestinal function.

  7. Muscarinic ACh Receptors Contribute to Aversive Olfactory Learning in Drosophila

    PubMed Central

    Silva, Bryon; Molina-Fernández, Claudia; Ugalde, María Beatriz; Tognarelli, Eduardo I.; Angel, Cristian; Campusano, Jorge M.

    2015-01-01

    The most studied form of associative learning in Drosophila consists in pairing an odorant, the conditioned stimulus (CS), with an unconditioned stimulus (US). The timely arrival of the CS and US information to a specific Drosophila brain association region, the mushroom bodies (MB), can induce new olfactory memories. Thus, the MB is considered a coincidence detector. It has been shown that olfactory information is conveyed to the MB through cholinergic inputs that activate acetylcholine (ACh) receptors, while the US is encoded by biogenic amine (BA) systems. In recent years, we have advanced our understanding on the specific neural BA pathways and receptors involved in olfactory learning and memory. However, little information exists on the contribution of cholinergic receptors to this process. Here we evaluate for the first time the proposition that, as in mammals, muscarinic ACh receptors (mAChRs) contribute to memory formation in Drosophila. Our results show that pharmacological and genetic blockade of mAChRs in MB disrupts olfactory aversive memory in larvae. This effect is not explained by an alteration in the ability of animals to respond to odorants or to execute motor programs. These results show that mAChRs in MB contribute to generating olfactory memories in Drosophila. PMID:26380118

  8. Muscarinic receptors in amygdala control trace fear conditioning.

    PubMed

    Baysinger, Amber N; Kent, Brianne A; Brown, Thomas H

    2012-01-01

    Intelligent behavior requires transient memory, which entails the ability to retain information over short time periods. A newly-emerging hypothesis posits that endogenous persistent firing (EPF) is the neurophysiological foundation for aspects or types of transient memory. EPF is enabled by the activation of muscarinic acetylcholine receptors (mAChRs) and is triggered by suprathreshold stimulation. EPF occurs in several brain regions, including the lateral amygdala (LA). The present study examined the role of amygdalar mAChRs in trace fear conditioning, a paradigm that requires transient memory. If mAChR-dependent EPF selectively supports transient memory, then blocking amygdalar mAChRs should impair trace conditioning, while sparing delay and context conditioning, which presumably do not rely upon transient memory. To test the EPF hypothesis, LA was bilaterally infused, prior to trace or delay conditioning, with either a mAChR antagonist (scopolamine) or saline. Computerized video analysis quantified the amount of freezing elicited by the cue and by the training context. Scopolamine infusion profoundly reduced freezing in the trace conditioning group but had no significant effect on delay or context conditioning. This pattern of results was uniquely anticipated by the EPF hypothesis. The present findings are discussed in terms of a systems-level theory of how EPF in LA and several other brain regions might help support trace fear conditioning.

  9. Bovine pancreatic polypeptide as an antagonist of muscarinic cholinergic receptors

    SciTech Connect

    Pan, G.Z.; Lu, L.; Qian, J.; Xue, B.G.

    1987-03-01

    In dispersed acini from rat pancreas, it was found that bovine pancreatic polypeptide (BPP) and its C-fragment hexapeptide amide (PP-6), at concentrations of 0.1 and 30 ..mu..M, respectively, could significantly inhibit amylase secretion stimulated by carbachol, and this inhibition by BPP was dose dependent. /sup 45/Ca outflux induced by carbachol was also inhibited by BPP or PP-6, but they had no effect on cholecystokinin octapeptide- (CCK-8) or A23187-stimulated /sup 45/Ca outflux. BPP was also capable of displacing the specific binding of (/sup 3/H)-quinuclidinyl benzilate to its receptors, and it possessed a higher affinity (K/sub i/35nM) than carbachol (K/sub i/ 1.8 ..mu..M) in binding with M-receptors. It is concluded from this study that BPP acts as an antagonist of muscarinic cholinergic receptors in rat pancreatic acini. In addition, BPP inhibited the potentiation of amylase secretion caused by the combination of carbachol plus secretin or vasoactive intestinal peptide. This may be a possible explanation of the inhibitory effect of BPP on secretin-induced pancreatic enzyme secretion shown in vivo, since pancreatic enzyme secretion stimulated by secretin under experimental conditions may be the result of potentiation of enzyme release produced by the peptide in combination with a cholinergic stimulant.

  10. Muscarinic component of splanchnic-adrenal transmission in the dog

    PubMed Central

    Kayaalp, S. O.; McIsaac, R. J.

    1969-01-01

    1. The effect of atropine on catecholamine release evoked by stimulation of the splanchnic nerve was studied in the adrenal medulla of the dog. The magnitude of the catecholamine release was estimated biologically on the basis of the pressor response occurring in the perfused and acutely sympathectomized hindquarters of the same dog by comparing it with responses elicited by intravenous injection of adrenaline. 2. Atropine reduced the responses to nerve stimulation, and appeared to have a more prominent effect on the responses elicited by stimulation at moderate frequency (10 pulses/sec). 3. Hexamethonium or nicotine caused a more powerful, but not complete, blockade of transmission; the subsequent injection of atropine caused a further inhibition of the residual responses, leading to a complete, or near complete, abolition of the release by nerve stimulation. 4. The data were taken as evidence that transmission of impulses through muscarinic receptors occurs in normal conditions in the adrenal medulla of the dog, though this type of transmission is less prominent than that through nicotinic receptors. PMID:4306886

  11. Autoradiographic analysis of muscarinic receptors in rat nasal glands.

    PubMed

    Van Megen, Y J; Teunissen, M J; Klaassen, A B; Rodrigues de Miranda, J F

    1988-01-01

    An in vitro method was developed for the biochemical and autoradiographic demonstration of low muscarinic receptor densities in peripheral tissue. Histological criteria point clearly to the necessity for fixation to preserve tissue quality. [3H]l-Quinuclidinylbenzilate bound specifically to a homogeneous class of binding sites in 0.5% glutardialdehyde-fixed cryostat sections (10 microns) of rat nasal glands with high affinity (Kd = 0.47 +/- 0.06 nM) and with a receptor density (Bmax) of 41 +/- 1 fmol/mg protein. This binding was linearly dependent on the thickness of the sections. Kinetic experiments resulted in a Kd value of 0.19 nM. Binding was stereoselectively inhibited by benzetimide enantiomers. Autoradiograms, generated after incubation with 0.6 nM [3H]l-quinuclidinylbenzilate and dipping in nuclear K2 emulsion, showed specific labelling of the glandular acini and excretory ducts. These in vitro observations provide conclusive evidence for the presence of acetylcholine receptors in the nasal glands of the rat. PMID:2450760

  12. Regulation of phospholipase D by muscarinic receptors in rat submandibular ductal cells.

    PubMed

    Pochet, Stéphanie; Métioui, Mourad; Grosfils, Katrina; Gómez-Muñoz, Antonio; Marino, Aida; Dehaye, Jean-Paul

    2003-01-01

    The muscarinic agonist carbachol stimulated phospholipase D (PLD) in rat submandibular gland (RSMG) ductal cells in a time and concentration-dependent manner. This effect was inhibited by chelation of extracellular calcium with ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). PLD could also be activated by epinephrine and AlF(4)(-), two polyphosphoinositide-specific phospholipase C (PPI-PLC) activators, and by the phorbol ester o-tetradecanoylphorbol 13-acetate (TPA) which activates protein kinase C (PKC). Ionomycin and thapsigargin only slightly increased PLD activity. Ortho-vanadate, a tyrosine phosphatase inhibitor, also stimulated PLD activity. Both carbachol and o-vanadate increased the formation of inositol phosphates and the tyrosine phosphorylation of at least two proteins (55-60 and 120 kDa). Calphostin C (a PKC inhibitor), U73122 (a PPI-PLC inhibitor) and genistein (a tyrosine kinase inhibitor) blocked the activation of PLD, of PLC and the phosphorylation of tyrosyl residues in response to carbachol and vanadate. Taken together, these results suggest that rat submandibular gland ductal cells express a calcium-dependent PLD activity. This enzyme is regulated by carbachol via a PLC-PKC-tyrosine kinase pathway. PMID:12401525

  13. Na+ activation of the muscarinic K+ channel by a G-protein-independent mechanism

    PubMed Central

    1996-01-01

    Muscarinic potassium channels (KACh) are composed of two subunits, GIRK1 and GIRK4 (or CIR), and are directly gated by G proteins. We have identified a novel gating mechanism of KACh, independent of G-protein activation. This mechanism involved functional modification of KACh which required hydrolysis of physiological levels of intracellular ATP and was manifested by an increase in the channel mean open time. The ATP-modified channels could in turn be gated by intracellular Na+, starting at approximately 3 mM with an EC50 of approximately 40 mM. The Na(+)-gating of KACh was operative both in native atrial cells and in a heterologous system expressing recombinant channel subunits. Block of the Na+/K+ pump (e.g., by cardiac glycosides) caused significant activation of KACh in atrial cells, with a time course similar to that of Na+ accumulation and in a manner indistinguishable from that of Na(+)-mediated activation of the channel, suggesting that cardiac glycosides activated KACh by increasing intracellular Na+ levels. These results demonstrate for the first time a direct effect of cardiac glycosides on atrial myocytes involving ion channels which are critical in the regulation of cardiac rhythm. PMID:8923264

  14. Effects of selected muscarinic cholinergic antagonists on [3H]acetylcholine release from rat hippocampal slices.

    PubMed

    Pohorecki, R; Head, R; Domino, E F

    1988-01-01

    A number of cholinergic muscarinic (M) agonists and antagonists were studied for their ability to enhance tritiated acetylcholine ([3H]ACh) release from electrically field-stimulated rat hippocampal slices. A Ca++-free medium and carbachol, but not nicotine, inhibited [3H]ACh release. Atropine, methylatropine and dexetimide produced concentration-dependent increases in [3H]ACh release to a maximum of about 50% above control. Aprophen and benactyzine produced a maximal response 25 to 35% above control. The selective M1 antagonist pirenzepine had the least effect on [3H]ACh release. Of the nonspecific M1-M2 antagonists studied, benactyzine produced the least amount of [3H]ACh release. The order of potency of the M antagonists in promoting a 15% increase in [3H]ACh release was aprophen greater than benactyzine greater than methylatropine greater than dexetimide greater than pirenzepine greater than atropine. However, the order of promoting maximal release of [3H]ACh was atropine greater than dexetimide greater than methylatropine greater than aprophen greater than benactyzine greater than pirenzepine. PMID:3335998

  15. Spatiotemporal calcium signaling in a Drosophila melanogaster cell line stably expressing a Drosophila muscarinic acetylcholine receptor.

    PubMed

    Cordova, D; Delpech, V Raymond; Sattelle, D B; Rauh, J J

    2003-11-01

    A muscarinic acetylcholine receptor (mAChR), DM1, expressed in the nervous system of Drosophila melanogaster, has been stably expressed in a Drosophila S2 cell line (S2-DM1) and used to investigate spatiotemporal calcium changes following agonist activation. Carbamylcholine (CCh) and oxotremorine are potent agonists, whereas application of the vertebrate M1 mAChR agonist, McN-A-343, results in a weak response. Activation of S2-DM1 receptors using CCh resulted in an increase in intracellular calcium ([Ca(2+)](i)) that was biphasic. Two distinct calcium sources were found to contribute to calcium signaling: (1) internal stores that are sensitive to both thapsigargin and 2-aminoethoxydiphenyl borate and (2) capacitative calcium entry. Spatiotemporal imaging of individual S2-DM1 cells showed that the CCh-induced [Ca(2+)](i) transient resulted from a homogeneous calcium increase throughout the cell, indicative of calcium release from internal stores. In contrast, ionomycin induced the formation of a "calcium ring" at the cell periphery, consistent with external calcium influx. PMID:12827518

  16. Calcium current activated by muscarinic receptors and thapsigargin in neuronal cells

    PubMed Central

    1994-01-01

    The activation of muscarinic receptors in N1E-115 neuroblastoma cells elicits a voltage-independent calcium current. The current turns on slowly, reaches its maximum value approximately 45 s after applying the agonist, is sustained as long as agonist is present, and recovers by one half in approximately 10 s after washing the agonist away. The current density is 0.11 +/- 0.08 pA/pF (mean +/- SD; n = 12). It is absent in zero-Ca++ saline and reduced by Mn++ and Ba++. The I(V) curve characterizing the current has an extrapolated reversal potential > +40 mV. The calcium current is observed in cells heavily loaded with BAPTA indicating that the calcium entry pathway is not directly gated by calcium. In fura-2 experiments, we find that muscarinic activation causes an elevation of intracellular Ca++ that is due to both intracellular calcium release and calcium influx. The component of the signal that requires external Ca++ has the same time course as the receptor operated calcium current. Calcium influx measured in this way elevates (Ca++)i by 89 +/- 41 nM (n = 7). Thapsigargin, an inhibitor of Ca++/ATPase associated with the endoplasmic reticulum (ER), activates a calcium current with similar properties. The current density is 0.22 +/- 0.20 pA/pF (n = 6). Thapsigargin activated current is reduced by Mn++ and Ba++ and increased by elevated external Ca++. Calcium influx activated by thapsigargin elevates (Ca++)i by 82 +/- 35 nM. The Ca++ currents due to agonist and due to thapsigargin do not sum, indicating that these procedures activate the same process. Carbachol and thapsigargin both cause calcium release from internal stores and the calcium current bears strong similarity to calcium-release-activated calcium currents in nonexcitable cells (Hoth, M., and R. Penner. 1993. Journal of Physiology. 465:359-386; Zweifach, A., and R. S. Lewis, 1993. Proceedings of the National Academy of Sciences, USA. 90:6295- 6299). PMID:7964592

  17. The binding of (3H)AF-DX 384 to rat ileal smooth muscle muscarinic receptors

    SciTech Connect

    Entzeroth, M.; Mayer, N. )

    1991-01-01

    The tritiated cardioselective muscarinic antagonist AF-DX 384 (5,11-dihydro-11-(2-(-(8-dipropylamino)methyl)-1-piperidinyl-ethyl-amino-carbonyl)-6H-pyrido (2,3-b) (1,4)benzodiazepin-6-one) was used to label muscarinic receptors in the rat ileum. Saturation binding to membrane suspensions revealed a high affinity binding site with a Kd of 9.2 nM. The maximal number of binding sites labeled in this tissue (Bmax) is 237 fmol/mg protein. The association and dissociation kinetics were well represented by single exponential reactions, and the dissociation constant obtained from the ratio of rate constants was in agreement with that derived from saturation experiments. Specific binding was inhibited by muscarinic antagonists with a rank order of potencies of atropine (pKi: 8.80) greater than 4-DAMP (pKi: 8.23) = AF-DX 384 (pKi: 8.20) greater than AF-DX 116 (pKi: 7.09) = hexahydro-sila-difenidol (pKi: 6.97) greater than pirenzepine (pKi: 6.49) and is consistent with the interaction of (3H)AF-DX 384 with muscarinic receptors of the M2 subtype. It can be concluded that (3H)AF-DX 384 can be used to selectively label M2 muscarinic receptors in heterogeneous receptor populations.

  18. Urtica dioica leaves modulates muscarinic cholinergic system in the hippocampus of streptozotocin-induced diabetic mice.

    PubMed

    Patel, Sita Sharan; Parashar, Arun; Udayabanu, Malairaman

    2015-06-01

    Diabetes mellitus is a chronic metabolic disorder and has been associated with cognitive dysfunction. In our earlier study, chronic Urtica dioica (UD) treatment significantly ameliorated diabetes induced associative and spatial memory deficit in mice. The present study was designed to explore the effect of UD leaves extract on muscarinic cholinergic system, which has long been known to be involved in cognition. Streptozotocin (STZ) (50 mg/kg, i.p., consecutively for 5 days) was used to induce diabetes followed by treatment with UD extract (50 mg/kg, oral) or rosiglitazone (5 mg/kg, oral) for 8 weeks. STZ-induced diabetic mice showed significant reduction in hippocampal muscarinic acetylcholine receptor-1 and choline acetyltransferase expressions. Chronic diabetes significantly up-regulated the protein expression of acetylcholinesterase associated with oxidative stress in hippocampus. Besides, STZ-induced diabetic mice showed hypolocomotion with up-regulation of muscarinic acetylcholine receptor-4 expression in striatum. Chronic UD treatment significantly attenuated the cholinergic dysfunction and oxidative stress in the hippocampus of diabetic mice. UD had no effect on locomotor activity and muscarinic acetylcholine receptor-4 expression in striatum. In conclusion, UD leaves extract has potential to reverse diabetes mediated alteration in muscarinic cholinergic system in hippocampus and thereby improve memory functions. PMID:25514862

  19. Distinct Roles of Bulbar Muscarinic and Nicotinic Receptors in Olfactory Discrimination Learning

    PubMed Central

    Devore, Sasha; de Almeida, Licurgo

    2014-01-01

    The olfactory bulb (OB) and piriform cortex receive dense cholinergic projections from the basal forebrain. Cholinergic modulation within the piriform cortex has long been proposed to serve important functions in olfactory learning and memory. We here investigate how olfactory discrimination learning is regulated by cholinergic modulation of the OB inputs to the piriform cortex. We examined rats' performance on a two-alternative choice odor discrimination task following local, bilateral blockade of cholinergic nicotinic and/or muscarinic receptors in the OB. Results demonstrate that acquisition, but not recall, of novel discrimination problems is impaired following blockade of OB cholinergic receptors, although the relative contribution of muscarinic and nicotinic receptors depends on task difficulty. Blocking muscarinic receptors impairs learning for nearly all odor sets, whereas blocking nicotinic receptors only affects performance for perceptually similar odors. This pattern of behavioral effects is consistent with predictions from a model of cholinergic modulation in the OB and piriform cortex (de Almeida et al., 2013). Model simulations suggest that muscarinic and nicotinic receptors may serve complementary roles in regulating coherence and sparseness of the OB network output, which in turn differentially regulate the strength and overlap in cortical odor representations. Overall, our results suggest that muscarinic receptor blockade results in a bona fide learning impairment that may arise because cortical neurons are activated less often. Behavioral impairment following nicotinic receptor blockade may not be due to the inability of the cortex to learn, but rather arises because the cortex is unable to resolve highly overlapping input patterns. PMID:25143606

  20. Heterogeneous receptor binding of classical quaternary muscarinic antagonists. I. Bovine tissue distribution.

    PubMed

    Roffel, A F; Ensing, K; in 't Hout, W G; de Zeeuw, R A; Zaagsma, J

    1991-01-01

    In competition experiments with the tertiary radioligand [3H]dexetimide, classical quaternary muscarinic antagonists like ipratropium bromide and N-methylscopolamine bromide distinguished two muscarinic binding sites in bovine brain (total brain minus cerebellum) membranes, in contrast to their tertiary analogues, atropine and scopolamine, which recognized only one binding site. This binding behavior was found to be almost identical in bovine striatal membranes, both in terms of binding affinities and proportions of high (Q1) and low (Q2) affinity binding sites. Both in total brain and in striatal membranes, the Q1/Q2 binding heterogeneity was independent of pirenzepine binding heterogeneity (M1/M2). In peripheral tissues, the binding properties of quaternary muscarinic antagonists varied. Whereas tertiary as well as quaternary compounds showed only high affinity binding towards muscarinic receptors in bovine atrial and left ventricular membranes, heterogeneous binding behavior was observed with quaternary but not with tertiary antagonists in bovine tracheal smooth muscle membranes. The tissue distribution found in the present study suggests that bovine tracheal smooth muscle contraction studies might shed light on the functional significance of the anomalous binding behavior of quaternary muscarinic antagonists. PMID:1824191

  1. Effects of extracellular acetylcholine on muscarinic receptor binding assessed by [125I]dexetimide and a simple probe.

    PubMed

    Sánchez-Roa, P M; Wagner, H N; Villemagne, V L; London, E D; Lever, J R

    1998-10-01

    New pharmacologic approaches to enhance brain cholinergic function focus on increasing intrasynaptic acetylcholine. We examined the usefulness of a simple probe and [125I]dexetimide to evaluate in vivo the effects of extracellular acetylcholine on muscarinic receptor binding in the mouse brain. After radiotracer injection continuous time/activity curves were generated over 330 min. [125I]Dexetimide reached a plateau at 90 min post-injection. To increase extracellular acetylcholine, the anticholinesterase physostigmine was administered at 120 min, producing a reversible decrease in [125I]dexetimide specific binding (23%) for 30 min. These findings demonstrate that dynamic changes in extracellular acetylcholine can be evaluated by displacement of [125I]dexetimide binding in vivo using a simple probe system. PMID:9822886

  2. Human electrophysiological correlates of learned irrelevance: effects of the muscarinic M1 antagonist biperiden.

    PubMed

    Klinkenberg, Inge; Blokland, Arjan; Riedel, Wim; Sambeth, Anke

    2012-11-01

    Learned irrelevance (LIrr) refers to a reduction in associative learning after pre-exposure of the conditioned and unconditioned stimulus in a non-contingent fashion. This paradigm might serve as a translational model for (pre)attentive information processing deficits in schizophrenia. This is the first study to investigate the event-related potentials (ERPs) of a within-subject LIrr paradigm in humans. Furthermore, the effects of the muscarinic M1 antagonist biperiden on LIrr were assessed. As expected, LIrr was found to be intact in young healthy volunteers after placebo. Furthermore, in the placebo condition P3b latency was decreased for target stimuli, which were pre-cued. This suggests that the predictability of the occurrence of these stimuli is mainly reflected by this ERP component. Biperiden had no effect on the behavioural LIrr measures, although prolonged reaction times were evident. Biperiden increased the N1 amplitude of the pre-exposed predictor letters, suggesting an effect of this drug on early perceptual processing. In conclusion, the within-subject paradigm used in the current study in combination with electroencephalography can reveal brain mechanisms involved in LIrr. M1 antagonism did not affect LIrr performance but seemed to influence early information processing.

  3. M2 muscarinic receptor activation regulates Schwann cell differentiation and myelin organization.

    PubMed

    Uggenti, Carolina; De Stefano, M Egle; Costantino, Michele; Loreti, Simona; Pisano, Annalinda; Avallone, Bice; Talora, Claudio; Magnaghi, Valerio; Tata, Ada Maria

    2014-07-01

    Glial cells express acetylcholine receptors. In particular, rat Schwann cells express different muscarinic receptor subtypes, the most abundant of which is the M2 subtype. M2 receptor activation causes a reversible arrest of the cell cycle. This negative effect on Schwann cell proliferation suggests that these cells may possibly progress into a differentiating program. In this study we analyzed the in vitro modulation, by the M2 agonist arecaidine, of transcription factors and specific signaling pathways involved in Schwann cell differentiation. The arecaidine-induced M2 receptor activation significantly upregulates transcription factors involved in the promyelinating phase (e.g., Sox10 and Krox20) and downregulates proteins involved in the maintenance of the undifferentiated state (e.g., c-jun, Notch-1, and Jagged-1). Furthermore, arecaidine stimulation significantly increases the expression of myelin proteins, which is accompanied by evident changes in cell morphology, as indicated by electron microscopy analysis, and by substantial cellular re-distribution of actin and cell adhesion molecules. Moreover, ultrastructural and morphometric analyses on sciatic nerves of M2/M4 knockout mice show numerous degenerating axons and clear alterations in myelin organization compared with wild-type mice. Therefore, our data demonstrate that acetylcholine mediates axon-glia cross talk, favoring Schwann cell progression into a differentiated myelinating phenotype and contributing to compact myelin organization.

  4. Subacute Zinc Administration and L-NAME Caused an Increase of NO, Zinc, Lipoperoxidation, and Caspase-3 during a Cerebral Hypoxia-Ischemia Process in the Rat

    PubMed Central

    Blanco-Alvarez, Victor Manuel; Lopez-Moreno, Patricia; Soto-Rodriguez, Guadalupe; Martinez-Fong, Daniel; Rubio, Hector; Gonzalez-Barrios, Juan Antonio; Piña-Leyva, Celia; Torres-Soto, Maricela; Gomez-Villalobos, María de Jesus; Hernandez-Baltazar, Daniel; Eguibar, José Ramon; Ugarte, Araceli; Cebada, Jorge

    2013-01-01

    Zinc or L-NAME administration has been shown to be protector agents, decreasing oxidative stress and cell death. However, the treatment with zinc and L-NAME by intraperitoneal injection has not been studied. The aim of our work was to study the effect of zinc and L-NAME administration on nitrosative stress and cell death. Male Wistar rats were treated with ZnCl2 (2.5 mg/kg each 24 h, for 4 days) and N-ω-nitro-L-arginine-methyl ester (L-NAME, 10 mg/kg) on the day 5 (1 hour before a common carotid-artery occlusion (CCAO)). The temporoparietal cortex and hippocampus were dissected, and zinc, nitrites, and lipoperoxidation were assayed at different times. Cell death was assayed by histopathology using hematoxylin-eosin staining and caspase-3 active by immunostaining. The subacute administration of zinc before CCAO decreases the levels of zinc, nitrites, lipoperoxidation, and cell death in the late phase of the ischemia. L-NAME administration in the rats treated with zinc showed an increase of zinc levels in the early phase and increase of zinc, nitrites, and lipoperoxidation levels, cell death by necrosis, and the apoptosis in the late phase. These results suggest that the use of these two therapeutic strategies increased the injury caused by the CCAO, unlike the alone administration of zinc. PMID:23997853

  5. Axonal transport of muscarinic receptors in vesicles containing noradrenaline and dopamine-beta-hydroxylase.

    PubMed

    Laduron, P M

    1984-01-01

    Presynaptic muscarinic receptors labeled with [3H]dexetimide and noradrenaline in dog splenic nerves accumulated proximally to a ligature at the same rate of axonal transport. After fractionation by differential centrifugation, specific [3H]quinuclidinyl benzilate or [3H]dexetimide binding revealed a distribution profile similar to that of dopamine-beta-hydroxylase and noradrenaline. Subfractionation by density gradient centrifugation showed two peaks of muscarinic receptors; the peak of density 1.17 contained noradrenaline and dopamine-beta-hydroxylase whereas that of density 1.14 was devoid of noradrenaline. Therefore the foregoing experiments provide evidence that presynaptic muscarinic receptors are transported in sympathetic nerves in synaptic vesicles which are similar to those containing noradrenaline and dopamine-beta-hydroxylase. This suggests a possible coexistence of receptor and neurotransmitter in the same vesicle. PMID:6198205

  6. Ovulation requires the activation on proestrus of M₁ muscarinic receptors in the left ovary.

    PubMed

    Cruz, M E; Flores, A; Alvarado, B E; Hernández, C G; Zárate, A; Chavira, R; Cárdenas, M; Arrieta-Cruz, I; Gutiérrez-Juárez, R

    2015-08-01

    We analyzed the effects of chemically blocking type 1 muscarinic receptors (M1R) on either the left or right ovary on ovulation rate, number of ova shed and steroid hormones levels. M1R were unilaterally blocked in ovary with the M1R selective antagonist pirenzepine (PZP). PZP was delivered into the bursa ovarica of the left or right ovary of adult rats at 13:00 h on proestrus day. PZP treatment in the left but not in the right ovary blocked ovulation. PZP did not modify the number of ova shed, nor progesterone or 17β-estradiol serum levels. The surge of luteinizing hormone levels was diminished while that of follicle-stimulating hormone did not change in animals treated with PZP in the left ovary. Interestingly, treatment with either synthetic luteinizing hormone-releasing hormone or human chorionic gonadotropin 1 h after PZP administration in the left ovary restored ovulation in both ovaries. The presence of M1R protein in the theca cells of the ovarian follicles as well as in cells of the corpus luteum was detected on proestrus day. These results suggest that M1R activation in the left ovary is required for pre-ovulatory gonadotropin-releasing hormone (GnRH) secretion and ovulation. Furthermore, these results also suggest that M1R in the left ovary might be regulating ovulation asymmetrically through a stimulatory neural signal relayed to the hypothalamus via the vagus nerve to induce the GnRH secretion which then triggers ovulation.

  7. Structure and dynamics of the M3 muscarinic acetylcholine receptor

    SciTech Connect

    Kruse, Andrew C.; Hu, Jianxin; Pan, Albert C.; Arlow, Daniel H.; Rosenbaum, Daniel M.; Rosemond, Erica; Green, Hillary F.; Liu, Tong; Chae, Pil Seok; Dror, Ron O.; Shaw, David E.; Weis, William I.; Wess, Jürgen; Kobilka, Brian K.

    2012-03-01

    Acetylcholine, the first neurotransmitter to be identified, exerts many of its physiological actions via activation of a family of G-protein-coupled receptors (GPCRs) known as muscarinic acetylcholine receptors (mAChRs). Although the five mAChR subtypes (M1-M5) share a high degree of sequence homology, they show pronounced differences in G-protein coupling preference and the physiological responses they mediate. Unfortunately, despite decades of effort, no therapeutic agents endowed with clear mAChR subtype selectivity have been developed to exploit these differences. We describe here the structure of the G{sub q/11}-coupled M3 mAChR ('M3 receptor', from rat) bound to the bronchodilator drug tiotropium and identify the binding mode for this clinically important drug. This structure, together with that of the G{sub i/o}-coupled M2 receptor, offers possibilities for the design of mAChR subtype-selective ligands. Importantly, the M3 receptor structure allows a structural comparison between two members of a mammalian GPCR subfamily displaying different G-protein coupling selectivities. Furthermore, molecular dynamics simulations suggest that tiotropium binds transiently to an allosteric site en route to the binding pocket of both receptors. These simulations offer a structural view of an allosteric binding mode for an orthosteric GPCR ligand and provide additional opportunities for the design of ligands with different affinities or binding kinetics for different mAChR subtypes. Our findings not only offer insights into the structure and function of one of the most important GPCR families, but may also facilitate the design of improved therapeutics targeting these critical receptors.

  8. Purification of the muscarinic acetylcholine receptor from porcine atria.

    PubMed Central

    Peterson, G L; Herron, G S; Yamaki, M; Fullerton, D S; Schimerlik, M I

    1984-01-01

    The muscarinic acetylcholine receptor from porcine atria has been purified 100,000-fold to homogeneity by solubilization in digitonin/cholate and sequential chromatography on wheat germ agglutinin-agarose, diethylaminoethylagarose, hydroxylapatite, and 3-(2'-aminobenzhydryloxy)tropane-agarose. The yield of purified receptor was 4.3% of that found in the membrane fraction, and the purified receptor bound 11.1-12.8 nmol of L-[3H]quinuclidinyl benzilate per mg of protein, corresponding to a binding component Mr of 78,400-90,000. The purified receptor preparation consisted of two polypeptides in approximately equimolar amounts when examined on silver-stained sodium dodecyl sulfate/polyacrylamide gels. The larger polypeptide (Mr 78,000 on 8% polyacrylamide gels) was specifically alkylated with [3H]propylbenzilylcholine mustard, whereas the smaller polypeptide (Mr 14,800) was not labeled. The possibility that the small polypeptide is a contaminant fortuitously appearing in equimolar amounts with the large polypeptide cannot be ruled out at this time. The purified preparation was highly stable, with no measurable change in the number of ligand binding sites or the gel pattern after 1 month's storage on ice. Scatchard analysis showed a single class of binding sites for the antagonist L-[3H]quinuclidinyl benzilate with a dissociation constant of 61 +/- 4 pM. Equilibrium titration experiments demonstrated that the antagonist L-hyoscyamine displaced L-[3H]quinuclidinyl benzilate from a single class of sites (Kd = 475 +/- 30 pM), whereas the agonist carbamoylcholine interacted at two populations of sites (53% +/- 3% high affinity, Kd = 1.1 +/- 0.3 microM; 47% +/- 3% low affinity, Kd = 67 +/- 14 microM). The ligand binding data were very similar to that for the membrane-bound receptor, suggesting that the receptor has not been altered radically during purification. Images PMID:6589642

  9. Subclassification of muscarinic receptors in the heart, urinary bladder and sympathetic ganglia in the pithed rat. Selectivity of some classical agonists.

    PubMed

    van Charldorp, K J; de Jonge, A; Thoolen, M J; van Zwieten, P A

    1985-12-01

    In pithed normotensive rats muscarinic receptors were characterized in heart, urinary bladder and sympathetic ganglia; the selectivity of some classical muscarinic agents for these subtypes was investigated. The potencies in decreasing heart rate, increasing bladder pressure and increasing diastolic blood pressure were measured for the following, intraarterially administered cholinergic agonists: McN-A-343 ([4-m-chlorophenylcarbamoyloxy]-2-butynyltrimethylammonium), pilocarpine, carbachol, oxotremorine, arecoline, acetyl-beta-methylcholine and acetylcholine. The selective M1-antagonist pirenzepine, the mixed M1/M2-antagonist dexetimide and the cardioselective M2-antagonist gallamine were used as tools for identification of the receptors. All data were obtained after intravenous pretreatment with a high dose of atenolol to eliminate tachycardia induced by stimulating sympathetic ganglionic muscarinic receptors. Dexetimide strongly antagonized the bradycardia as well as the increase in bladder pressure induced by pilocarpine, carbachol, oxotremorine, arecoline, acetyl-beta-methylcholine and acetylcholine, whereas pirenzepine was much less effective. Gallamine antagonized the bradycardia, whereas no influence was found on the bladder contraction. Pilocarpine acted as a partial agonist in reducing heart rate as well as in increasing bladder pressure, whereas McN-A-343 was almost ineffective in doses up to 1 mg/kg. The hypertensive response to pilocarpine and carbachol was less pronounced than that produced by McN-A-343. Pirenzepine and dexetimide significantly antagonized the hypertensive response to McN-A-343 and pilocarpine, whereas gallamine was much less effective. The hypertensive response induced by carbachol was totally blocked by hexamethonium. The other agonists used in this study did not produce a significant increase in diastolic blood pressure in doses that produced a maximal effect on heart rate and urinary bladder pressure.(ABSTRACT TRUNCATED AT 250 WORDS

  10. Number of Academic Freedom Cases Increased by 20 Percent in 1986: Two Institutions Added to AAUP List of Censured Administrations; Three Others Were Removed. AAUP News Release.

    ERIC Educational Resources Information Center

    American Association of Univ. Professors, Washington, DC.

    An update on academic freedom cases and complaints made to the American Association of University Professors (AAUP) is presented. AAUP responded to 1,222 academic freedom cases in 1986, an increase of 20% in the number of cases handled in the 2 preceding years. Two institutions were added to AAUP's list of censured administrations: Clark College…

  11. Inhibition of muscarinic receptor-stimulated phosphoinositide metabolism by cocaine, norcocaine and cocaethylene in rat brain.

    PubMed

    Tan, X X; Costa, L G

    1994-05-13

    The interaction of cocaine, its metabolites norcocaine and benzoylecgonine, and cocaethylene, which is formed following a combined cocaine and ethanol exposure, with muscarinic receptor binding and phosphoinositide metabolism was investigated in brain from immature rats. Cocaine and norcocaine inhibited binding of [3H]telenzepine and carbachol-stimulated phosphoinositide metabolism in cerebral cortex, while benzoylecgonine was devoid of any inhibitory activity. Cocaethylene was the most potent inhibitor of both binding and phosphoinositide metabolism. The effect of cocaine was more pronounced at the muscarinic receptors, but a small inhibition of histamine--and serotonin--stimulated phosphoinositide metabolism was also observed.

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

    SciTech Connect

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

    1985-04-01

    The aim of the present investigation was a comparative studyof muscarinic acetylcholine receptors in human and rat glial cells. (/sup 3/H)Quinuclidinyl-benzylate ((/sup 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 (/sup 3/H)-QB with human and rat glial cell membranes demonstrated the presence of a muscarinic acetylcholine receptor in the glial cells.

  13. Affinity of pyridylalkylamines for nicotinic, muscarinic and histaminic recognition sites in brain tissue preparations.

    PubMed

    Repond, C; Pratt, J A; Stolerman, I P; Mayer, J M; Jenner, P; Marsden, C D; Testa, B

    1986-08-01

    The affinity of 15 regioisomeric and homologous pyridylalkylamines was examined in brain preparations for nicotinic, muscarinic, and H1-histaminic binding sites as labeled by [3H]-nicotine, [3H]-dexetimide and [3H]-mepyramine, respectively. Overall, the compounds show a clear selectivity for the nicotinic versus muscarinic binding sites, and a weak affinity for the H1-histaminic sites. Variations in affinity appear to be partly influenced by steric factors (such as position of attachment, length and rigidity of side-chain) and marginally by lipophilicity. PMID:3778556

  14. Different behavior toward muscarinic receptor binding between quaternary anticholinergics and their tertiary analogues.

    PubMed

    Ensing, K; de Zeeuw, R A

    1986-12-01

    A number of corresponding tertiary and quaternary anticholinergic analogues were examined for their ability to inhibit specific (3)H-dexetimide binding to calf brain muscarinic receptors. In all cases the tertiary antagonists (except pirenzepine) showed steep and monophasic inhibition curves, whereas those of the quaternary derivatives were shallow (thiazinamium, methylbenactyzine) or even biphasic (oxyphenonium, methylatropine, methylscopolamine). These observations show that the addition of a methyl group to the nitrogen atom changes the mode of interaction of the anticholinergics to muscarinic receptor binding sites. Whether there are separate binding sites present or differences in interaction mode for only the quaternary moiety is discussed. PMID:24271831

  15. Muscarinic receptor activation of phosphatidylcholine hydrolysis. Relationship to phosphoinositide hydrolysis and diacylglycerol metabolism

    SciTech Connect

    Martinson, E.A.; Goldstein, D.; Brown, J.H. )

    1989-09-05

    We examined the relationship between phosphatidylcholine (PC) hydrolysis, phosphoinositide hydrolysis, and diacylglycerol (DAG) formation in response to muscarinic acetylcholine receptor (mAChR) stimulation in 1321N1 astrocytoma cells. Carbachol increases the release of (3H)choline and (3H)phosphorylcholine ((3H)Pchol) from cells containing (3H)choline-labeled PC. The production of Pchol is rapid and transient, while choline production continues for at least 30 min. mAChR-stimulated release of Pchol is reduced in cells that have been depleted of intracellular Ca2+ stores by ionomycin pretreatment, whereas choline release is unaffected by this pretreatment. Phorbol 12-myristate 13-acetate (PMA) increases the release of choline, but not Pchol, from 1321N1 cells, and down-regulation of protein kinase C blocks the ability of carbachol to stimulate choline production. Taken together, these results suggest that Ca2+ mobilization is involved in mAChR-mediated hydrolysis of PC by a phospholipase C, whereas protein kinase C activation is required for mAChR-stimulated hydrolysis of PC by a phospholipase D. Both carbachol and PMA rapidly increase the formation of (3H)phosphatidic acid ((3H)PA) in cells containing (3H)myristate-labeled PC. (3H)Diacylglycerol ((3H)DAG) levels increase more slowly, suggesting that the predominant pathway for PC hydrolysis is via phospholipase D. When cells are labeled with (3H)myristate and (14C)arachidonate such that there is a much greater 3H/14C ratio in PC compared with the phosphoinositides, the 3H/14C ratio in DAG and PA increases with PMA treatment but decreases in response to carbachol.

  16. Activation of Muscarinic Acetylcholine Receptor Subtype 4 Is Essential for Cholinergic Stimulation of Gastric Acid Secretion: Relation to D Cell/Somatostatin

    PubMed Central

    Takeuchi, Koji; Endoh, Takuya; Hayashi, Shusaku; Aihara, Takeshi

    2016-01-01

    Background/Aim: Muscarinic acetylcholine receptors exist in five subtypes (M1∼M5), and they are widely expressed in various tissues to mediate diverse autonomic functions, including gastric secretion. In the present study, we demonstrated, using M1∼M5 KO mice, the importance of M4 receptors in carbachol (CCh) stimulation of acid secretion and investigated how the secretion is modulated by the activation of M4 receptors. Methods: C57BL/6J mice of wild-type (WT) and M1–M5 KO were used. Under urethane anesthesia, acid secretion was measured in the stomach equipped with an acute fistula. CCh (30 μg/kg) was given subcutaneously (s.c.) to stimulate acid secretion. Atropine or octreotide (a somatostatin analog) was given s.c. 20 min before the administration of CCh. CYN154806 (a somatostatin SST2 receptor antagonist) was given i.p. 20 min before the administration of octreotide or CCh. Results: CCh caused an increase of acid secretion in WT mice, and the effect was totally inhibited by prior administration of atropine. The effect of CCh was similarly observed in the animals lacking M1, M2 or M5 receptors but significantly decreased in M3 or M4 KO mice. CYN154806, the SST2 receptor antagonist, dose-dependently and significantly reversed the decreased acid response to CCh in M4 but not M3 KO mice. Octreotide, the somatostatin analog, inhibited the secretion of acid under CCh-stimulated conditions in WT mice. The immunohistochemical study showed the localization of M4 receptors on D cells in the stomach. Serum somatostatin levels in M4 KO mice were higher than WT mice under basal conditions, while those in WT mice were significantly decreased in response to CCh. Conclusions: These results suggest that under cholinergic stimulation the acid secretion is directly mediated by M3 receptors and indirectly modified by M4 receptors. It is assumed that the activation of M4 receptors inhibits the release of somatostatin from D cells and minimizes the acid inhibitory effect of

  17. Activation of Muscarinic Acetylcholine Receptor Subtype 4 Is Essential for Cholinergic Stimulation of Gastric Acid Secretion: Relation to D Cell/Somatostatin

    PubMed Central

    Takeuchi, Koji; Endoh, Takuya; Hayashi, Shusaku; Aihara, Takeshi

    2016-01-01

    Background/Aim: Muscarinic acetylcholine receptors exist in five subtypes (M1∼M5), and they are widely expressed in various tissues to mediate diverse autonomic functions, including gastric secretion. In the present study, we demonstrated, using M1∼M5 KO mice, the importance of M4 receptors in carbachol (CCh) stimulation of acid secretion and investigated how the secretion is modulated by the activation of M4 receptors. Methods: C57BL/6J mice of wild-type (WT) and M1–M5 KO were used. Under urethane anesthesia, acid secretion was measured in the stomach equipped with an acute fistula. CCh (30 μg/kg) was given subcutaneously (s.c.) to stimulate acid secretion. Atropine or octreotide (a somatostatin analog) was given s.c. 20 min before the administration of CCh. CYN154806 (a somatostatin SST2 receptor antagonist) was given i.p. 20 min before the administration of octreotide or CCh. Results: CCh caused an increase of acid secretion in WT mice, and the effect was totally inhibited by prior administration of atropine. The effect of CCh was similarly observed in the animals lacking M1, M2 or M5 receptors but significantly decreased in M3 or M4 KO mice. CYN154806, the SST2 receptor antagonist, dose-dependently and significantly reversed the decreased acid response to CCh in M4 but not M3 KO mice. Octreotide, the somatostatin analog, inhibited the secretion of acid under CCh-stimulated conditions in WT mice. The immunohistochemical study showed the localization of M4 receptors on D cells in the stomach. Serum somatostatin levels in M4 KO mice were higher than WT mice under basal conditions, while those in WT mice were significantly decreased in response to CCh. Conclusions: These results suggest that under cholinergic stimulation the acid secretion is directly mediated by M3 receptors and indirectly modified by M4 receptors. It is assumed that the activation of M4 receptors inhibits the release of somatostatin from D cells and minimizes the acid inhibitory effect of

  18. Activation Biosensor for G Protein-Coupled Receptors: A FRET-Based m1 Muscarinic Activation Sensor That Regulates Gq

    PubMed Central

    Chang, Seungwoo; Ross, Elliott M.

    2012-01-01

    We describe the design, construction and validation of a fluorescence sensor to measure activation by agonist of the m1 muscarinic cholinergic receptor, a prototypical class I Gq-coupled receptor. The sensor uses an established general design in which Förster resonance energy transfer (FRET) from a circularly permuted CFP mutant to FlAsH, a selectively reactive fluorescein, is decreased 15–20% upon binding of a full agonist. Notably, the sensor displays essentially wild-type capacity to catalyze activation of Gαq, and the purified and reconstituted sensor displays appropriate regulation of affinity for agonists by Gq. We describe the strategies used to increase the agonist-driven change in FRET while simultaneously maintaining regulatory interactions with Gαq, in the context of the known structures of Class I G protein-coupled receptors. The approach should be generally applicable to other Class I receptors which include numerous important drug targets. PMID:23029161

  19. Effect of paraoxon on muscarinic, dopamine and. gamma. -aminobutyric acid receptors of brain and sensitivity to muscarinic antagonists

    SciTech Connect

    Fernando, J.C.R.; Hoskins, B.; Ho, I.K.

    1986-03-05

    Several acetylcholinesterase (AChE) inhibitors decrease muscarinic cholinergic (mACh) receptors in the brain, alteration of dopamine (DA) and ..gamma..-aminobutyric acid (GABA) receptors after AChE inhibition was also reported. In view of the important interactions among DA, GABA and ACh systems, whether this is a common effect of AChE inhibitors should be established. They report the effect of the AChE inhibitor, paraoxon, on DA, GABA and mACh receptors in the rat. The binding of /sup 3/H-QNB (for mACh), /sup 3/H-spiperone (for DA) and /sup 3/H-muscimol (for GABA) to striatal and hippocampal membranes was analyzed. Also, behavioral sensitivity to atropine was studied. Twenty-four hr after a single dose (0.75 mg/kg, s.c.) of paraoxon, the density of mACh receptors in the striatum was decreased but, at 3 days, no change was seen. In the hippocampus, the mACh receptors were not affected. Repeated treatment with paraoxon (0.3 mg/kg, 48 hourly) for 2 weeks reduced the mACh receptor density in both regions. Neither single nor repeated paraoxon treatment had an effect on DA or GABA receptors. After single or repeated dosing with paraoxon, myoclonus induced by atropine (10 mg/kg, i.p.) was enhanced. The results show rapid downregulation of mACh receptors by paraoxon. DA or GABA, however, appear not to be affected under these treatment regimens.

  20. In vivo biodistribution of two ( sup 18 F)-labelled muscarinic cholinergic receptor ligands: 2-( sup 18 F)- and 4-( sup 18 F)-fluorodexetimide

    SciTech Connect

    Wilson, A.A.; Scheffel, U.A.; Dannals, R.F.; Stathis, M.; Ravert, H.T.; Wagner, H.N. Jr. )

    1991-01-01

    Two ({sup 18}F)-labelled analogues of the potent muscarinic cholinergic receptor (m-AChR) antagonist, dexetimide, were evaluated as potential ligands for imaging m-AChR by positron emission tomography (PET). Intravenous administration of both 2-({sup 18}F)- or 4-({sup 18}F)-fluorodexetimide resulted in high brain uptake of radioactivity in mice. High binding levels were observed in m-AChR rich areas, such as cortex and striatum, with low levels in the receptor-poor cerebellum. Uptake of radioactivity was saturable and could be blocked by pre-administration of dexetimide or atropine. Drugs with different sites of action were ineffective at blocking receptor binding. The results indicate that both radiotracers are promising candidates for use in PET studies.

  1. In vivo biodistribution of two [18F]-labelled muscarinic cholinergic receptor ligands: 2-[18F]- and 4-[18F]-fluorodexetimide.

    PubMed

    Wilson, A A; Scheffel, U A; Dannals, R F; Stathis, M; Ravert, H T; Wagner, H N

    1991-01-01

    Two [18F]-labelled analogues of the potent muscarinic cholinergic receptor (m-AChR) antagonist, dexetimide, were evaluated as potential ligands for imaging m-AChR by positron emission tomography (PET). Intravenous administration of both 2-[18F]- or 4-[18F]-fluorodexetimide resulted in high brain uptake of radioactivity in mice. High binding levels were observed in m-AChR rich areas, such as cortex and striatum, with low levels in the receptor-poor cerebellum. Uptake of radioactivity was saturable and could be blocked by pre-administration of dexetimide or atropine. Drugs with different sites of action were ineffective at blocking receptor binding. The results indicate that both radiotracers are promising candidates for use in PET studies. PMID:2008155

  2. Differential increased survival of staphylococci and limited ultrastructural changes in the core of infected fibrin clots after daptomycin administration.

    PubMed Central

    Michiels, M J; Bergeron, M G

    1996-01-01

    A possible explanation for the difficulties encountered in curing deep fibrin-embedded infections is that antibiotic diffusion inside the infected fibrin matrix is not homogeneous and is insufficient to neutralize the pathogen. To evaluate this conjecture, the differential pharmacodynamics of daptomycin in fibrin clots infected with methicillin-susceptible and -resistant Staphylococcus aureus and Staphylococcus epidermidis was estimated. Daptomycin (20 or 50 mg/kg of body weight) was infused over 30 min. Fibrin clots and blood samples were evaluated from 0.5 to 42 h after the injections. The half-lives of daptomycin in serum and fibrin clot were close to identical after the two doses and averaged 5.4 and 22 h, respectively. The mean areas under the concentration-time curves from 0 to 42 h (AUC0-infinity) for daptomycin concentrations in serum and infected clots were 575 +/- 36.7 and 215 +/- 6.2 micrograms/g/h after administration of 20 mg/kg and 1,089 +/- 39.9 and 326 +/- 16.8 micrograms/g/h after administration of 50 mg/kg. A concentration gradient from the periphery to the core of the clots was observed in many clots up to 18 h after treatment. Mean peak concentrations in the core of the clots reached 60% of the peripheral values (P < 0.05) and were delayed for at least 3 h compared with the peripheral peak concentrations. AUC0-42 h of daptomycin concentration in the periphery and the core of clots were significantly different (P < 0.01). Survival of microorganisms was better in the core than in the periphery, with as much as a 3 log10 CFU/g difference between the center and the surface of the clot. Bacterial examination by transmission electron microscopy also showed noticeable differences in ultrastructural changes between those in the periphery and those in the core of the clots. In conclusion, the pharmacokinetics of daptomycin are significantly different at the periphery and within the core of fibrin clots, which may have led to the higher bacterial survival

  3. Muscarinic acetylcholine receptor subtypes which selectively couple to phospholipase C: Pharmacological and biochemical properties

    SciTech Connect

    Buck, M.A.; Fraser, C.M. )

    1990-12-14

    The pharmacological and biochemical properties of rat m1 and m3 muscarinic acetylcholine receptors (mAChR) stably transfected into Chinese hamster ovary-K1 (CHO) cells were characterized with ligand binding, affinity labeling and biochemical assays. Both mAChR subtypes display saturable, high affinity binding of (3H)-quinuclidinyl benzilate (QNB) and a rank order of antagonist potency of QNB greater than atropine greater than pirenzepine greater than AF-DX 116. Carbachol displacement of (3H)-QNB binding to the m3 mAChR revealed an approximate 17-fold higher affinity than observed with the m1 mAChR. (3H)-propylbenzilylcholine mustard (PrBCM) labeling of mAChR revealed that m1 and m3 mAChR migrated on SDS-polyacrylamide gels with apparent molecular masses of 80,000 and 94,000 daltons, respectively, consistent with the known differences in their molecular sizes. Both m1 and m3 mAChR elicited dose-dependent increases in the hydrolysis of phosphoinositides; however, the maximal increase in total inositol phosphates elicited with the m1 mAChR was approximately 2-fold greater than that observed in cells expressing similar densities of m3 mAChR. Agonist activation of the m1 mAChR also elicited increases in basal and forskolin-stimulated cAMP, whereas the m3 mAChR had no effect on intracellular cAMP levels. These data suggest that although m1 and m3 mAChR display a considerable degree of structural homology, they exhibit distinct pharmacological and biochemical properties.

  4. Administration of orexin receptor 1 antagonist into the rostral ventromedial medulla increased swim stress-induced antinociception in rat

    PubMed Central

    Soliemani, Neda; Moslem, Alireza; Shamsizadeh, Ali; Azhdari-Zarmehri, Hassan

    2016-01-01

    Objective(s): Intracerebroventricular injection of orexin-A (hypocretin-1) antagonist has been shown to inhibit stress-induced analgesia. However the locations of central sites that may mediate these effects have not been totally demonstrated. This study was performed to investigate the role of rostral ventromedial medulla (RVM) orexin receptor 1 in stress-induced analgesia (SIA). Materials and Methods: Forced swim stress in water was employed to adult male rats (200-250 g). Nociceptive responses were measured by formalin test (50 µl injection of formalin 2% subcutaneously into hind paw) and, pain related behaviors were monitored for 90 min following intra-microinjection of SB-334867 (orexin receptor 1 antagonist) into RVM. Results: Exposure to swimming stress test after administration of SB-334867 into RVM significantly reduces the formalin-induced nociceptive behaviors in phase1, interphase, and phase 2 in rats. Conclusion: The result demonstrated the involvement of OXR1 in antinociceptive behaviors induced by swim stress in RVM. PMID:27403261

  5. Radioligand binding to muscarinic receptors of bovine aortic endothelial cells.

    PubMed Central

    Brunner, F.; Kukovetz, W. R.

    1991-01-01

    1. Muscarinic receptors on endothelial cells of bovine thoracic aorta were characterized by binding assays in which (-)-[3H]-N-methyl quinuclidinyl benzilate ([3H]-NMeQNB) was used as radioligand. 2. Binding of [3H]-NMeQNB to crude membranes of freshly isolated endothelial cells was atropine-displaceable and of high affinity (KD = 0.48 nM) to a single class of sites (maximum binding capacity: 14 +/- 3 fmol mg-1 protein). Stereospecificity of the binding sites was demonstrated in experiments in which [3H]-NMeQNB binding was inhibited by dexetimide in the nanomolar range (KI = 0.63 nM) and by levetimide, its stereoisomer in the micromolar range (KI = 3.2 microM) (selectivity factor: approximately 5000). 3. Drug competition curves indicated a single class of binding sites for antagonists and the following apparent affinities (KI, nM): methyl atropine: 1.1: 4-diphenylacetoxy N-methyl piperidine methyl bromide (4-DAMP): 3.4; pirenzepine: 16; 11-[2-diethylamino-methyl)-1-piperidinyl- acetyl]-5,11-dihydro-6H-pyrido(2,3-b)1,4-benzodiazepine-6-one (AF-DX 116); 2.500. Competition of acetylcholine with [3H]-NMeQNB was best described by two affinity sites (or states) (KH = 0.82 microM, KL = 1.6 microM). In the presence of guanylimido diphosphate [Gpp(NH)p] (100 microM), acetylcholine affinity (IC50) was slightly, but significantly reduced (factor approximately 4). 4. Binding of [3H]-NMeQNB to freshly harvested intact cells was also atropine-displaceable, stereospecific (selectivity factor: approximately 3500) and of high affinity (KD = 0.35 nM). The maximum binding capacity (9 +/- 2 fmol mg-1 total cell protein) was comparable to that of membranes and corresponded to approximately 900 binding sites per endothelial cell.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2015420

  6. Long-lasting increase in [³H]CP55,940 binding to CB1 receptors following cocaine self-administration and its withdrawal in rats.

    PubMed

    Adamczyk, Przemysław; Faron-Górecka, Agata; Kuśmider, Maciej; Dziedzicka-Wasylewska, Marta; Papp, Mariusz; Filip, Małgorzata

    2012-04-27

    The present work has aimed on the neuroadaptive changes in CB1 receptor density that are evoked by self-administered cocaine use and subsequent withdrawal in rats. We employed a quantitative autoradiographic analysis using labeled [³H]CP55,940, a CB1 receptor agonist. To distinguish the passive pharmacological effects of cocaine from those related to motivation and the cognitive processes evoked by active cocaine self-administration, the "yoked" procedure was used. Our results demonstrate that repeated cocaine administration over 14 days induced up-regulation of CB1 receptors in the cortical and subcortical brain areas of animals who received cocaine, whether the cocaine was actively self-administered or received passively (the "yoked" control group) and that the neuroadaptation of CB1 receptors persisted after the 10-day extinction phase. On the other hand, we found that only self-administering rats showed CB1 receptor up-regulation in numerous brain areas, which suggests that these structures may be directly linked to CB1 receptor control over motivational and cognitive processes. Moreover, the observed increase in [³H]CP55,940 binding in these brain areas likely indicates long-lasting neurobiological adaptations resulting from chronic cocaine self-administration. In conclusion, we demonstrated that chronic cocaine self-administration leads to increased CB1 receptor levels in numerous brain areas and that this neuroadaptation is maintained over a long-lasting extinction period.

  7. β1-Adrenergic and M2 Muscarinic Autoantibodies and Thyroid Hormone Facilitate Induction of Atrial Fibrillation in Male Rabbits.

    PubMed

    Li, Hongliang; Murphy, Taylor; Zhang, Ling; Huang, Bing; Veitla, Vineet; Scherlag, Benjamin J; Kem, David C; Yu, Xichun

    2016-01-01

    Activating autoantibodies to the β1-adrenergic and M2 muscarinic receptors are present in a very high percentage of patients with Graves' disease and atrial fibrillation (AF). The objective of this study was to develop a reproducible animal model and thereby to examine the impact of these endocrine-like autoantibodies alone and with thyroid hormone on induction of thyroid-associated atrial tachyarrhythmias. Five New Zealand white rabbits were coimmunized with peptides from the second extracellular loops of the β1-adrenergic and M2 muscarinic receptors to produce both sympathomimetic and parasympathomimetic antibodies. A catheter-based electrophysiological study was performed on anesthetized rabbits before and after immunization and subsequent treatment with thyroid hormone. Antibody expression facilitated the induction of sustained sinus, junctional and atrial tachycardias, but not AF. Addition of excessive thyroid hormone resulted in induced sustained AF in all animals. AF induction was blocked acutely by the neutralization of these antibodies with immunogenic peptides despite continued hyperthyroidism. The measured atrial effective refractory period as one parameter of AF propensity shortened significantly after immunization and was acutely reversed by peptide neutralization. No further decrease in the effective refractory period was observed after the addition of thyroid hormone, suggesting other cardiac effects of thyroid hormone may contribute to its role in AF induction. This study demonstrates autonomic autoantibodies and thyroid hormone potentiate the vulnerability of the heart to AF, which can be reversed by decoy peptide therapy. These data help fulfill Witebsky's postulates for an increased autoimmune/endocrine basis for Graves' hyperthyroidism and AF. PMID:26517045

  8. Participation of nitric oxide signaling system in the cardiac muscarinic cholinergic effect of human chagasic IgG.

    PubMed

    Sterin-Borda, L; Leiros, C P; Goin, J C; Cremaschi, G; Genaro, A; Echagüe, A V; Borda, E

    1997-07-01

    The possible role of altered humoral immune response in the pathogenesis of the chronic chagasic cardioneuromyopathy was examined by analyzing the interaction of IgG from T. cruzi infected patients with cardiac muscarinic acetylcholine receptors (mAChR). Human chagasic IgG by activating cardiac M2 mAChR, simulated the agonist actions triggering negative inotropic effect, inositol phosphate accumulation, nitric oxide synthase stimulation and increased production of cyclic GMP. Inhibitors of phospholipase C, protein kinase C, calcium/calmodulin, nitric oxide synthase and guanylate cyclase activities; prevented chagasic IgG effects on signaling pathways involved in M2 mAChR activation. In addition, sodium nitroprusside or 8-bromo cyclic GMP, mimicked the chagasic IgG effect associated with cholinergic-mediated cellular transmembrane signals. Moreover, these chagasic IgG immunoprecipitated the mAChRs solubilized from cardiac membranes. By means of SDS-PAGE and immunoblotting analysis, chagasic sera recognized a band of 70-75 kDa. The major protein recognized by chagasic IgG had an Rf coincident with the peak of [3H] propylbenzilylcholine mustard with an apparent molecular weight similar to that of mAChRs, which disappeared in the presence of atropine. The specificity of this interaction was checked by immunoprecipitation of rat cardiac mAChR and immunoblotting of pure human M2 mAChRs. Chronic interaction of chagasic IgG with myocardial mAChRs, behaving as a muscarinic agonist, might lead to cell dysfunction or tissue damage. Also, these antibodies could produce desensitization, internalization or degradation of mAChRs; explaining the progressive blockade of mAChRs in myocardium with parasympathetic denervation, a phenomenon that has been described in the course of Chagas' cardioneuromyopathy. PMID:9236139

  9. Heterogeneity of binding of muscarinic receptor antagonists in rat brain homogenates

    SciTech Connect

    Lee, J.H.; el-Fakahany, E.E.

    1985-06-01

    The binding properties of (-)-(/sup 3/H)quinuclidinyl benzilate and (/sup 3/H) N-methylscopolamine to muscarinic acetylcholine receptors have been investigated in rat brain homogenates. The binding of both antagonists demonstrated high affinity and saturability. Analysis of the binding data resulted in linear Scatchard plots. However, (-)-(/sup 3/H)quinuclidinyl benzilate showed a significantly higher maximal binding capacity than that of (/sup 3/H)N-methylscopolamine. Displacement of both ligands with several muscarinic receptor antagonists resulted in competition curves in accordance with the law of mass-action for quinuclidinyl benzilate, atropine and scopolamine. A similar profile was found for the quaternary ammonium analogs of atropine and scopolamine when (/sup 3/H)N-methylscopolamine was used to label the receptors. However, when these hydrophilic antagonists were used to displace (-)-(/sup 3/H) quinuclidinyl benzilate binding, they showed interaction with high- and low-affinity binding sites. On the other hand, the nonclassical muscarinic receptor antagonist, pirenzepine, was able to displace both ligands from two binding sites. The present data are discussed in terms of the relationship of this anomalous heterogenity of binding of these hydrophilic muscarinic receptor antagonists and the proposed M1 and M2 receptor subtypes.

  10. Muscarinic receptors are involved in LMM3 tumor cells proliferation and angiogenesis

    SciTech Connect

    Rimmaudo, Laura Elizabeth; Torre, Eulalia de la; Sacerdote de Lustig, Eugenia; Sales, Maria Elena . E-mail: mesales@2vias.com.ar

    2005-09-09

    Angiogenesis is a process of new blood vessel development from pre-existing vasculature and it plays an essential role in tumor growth and metastases. Here, we investigate the expression of muscarinic acetylcholine receptors (mAchR) and their participation in tumor cell proliferation and angiogenesis ability. Saturation binding assays with the tritiated muscarinic antagonist quinuclidinyl benzilate indicate that LMM3 cells derived from a murine mammary adenocarcinoma express a single class of functional mAchR. Competition binding assays with selective muscarinic antagonists indicate a predominance of M{sub 3} receptor subtype. The muscarinic agonist carbachol (CARB) stimulates LMM3 cell proliferation in a concentration dependent manner. The maximal effect induced by 10{sup -9} M CARB was totally blunted by atropine and by the selective M{sub 3} and M{sub 1} antagonists, para-fluoro hexahydro sila-difenidol (pf-HHSiD) and pirenzepine, respectively. In addition, pf-HHSiD completely blocked in vivo CARB-induced neovascular formation and vascular endothelial growth factor-A in LMM3 tumor cells. We can conclude that mAchR expressed in LMM3 mammary tumor cells positively regulate proliferation and angiogenesis required for tumor progression.

  11. Structure of the human M2 muscarinic acetylcholine receptor bound to an antagonist

    SciTech Connect

    Haga, Kazuko; Kruse, Andrew C.; Asada, Hidetsugu; Yurugi-Kobayashi, Takami; Shiroishi, Mitsunori; Zhang, Cheng; Weis, William I.; Okada, Tetsuji; Kobilka, Brian K.; Haga, Tatsuya; Kobayashi, Takuya

    2012-03-15

    The parasympathetic branch of the autonomic nervous system regulates the activity of multiple organ systems. Muscarinic receptors are G-protein-coupled receptors that mediate the response to acetylcholine released from parasympathetic nerves. Their role in the unconscious regulation of organ and central nervous system function makes them potential therapeutic targets for a broad spectrum of diseases. The M2 muscarinic acetylcholine receptor (M2 receptor) is essential for the physiological control of cardiovascular function through activation of G-protein-coupled inwardly rectifying potassium channels, and is of particular interest because of its extensive pharmacological characterization with both orthosteric and allosteric ligands. Here we report the structure of the antagonist-bound human M2 receptor, the first human acetylcholine receptor to be characterized structurally, to our knowledge. The antagonist 3-quinuclidinyl-benzilate binds in the middle of a long aqueous channel extending approximately two-thirds through the membrane. The orthosteric binding pocket is formed by amino acids that are identical in all five muscarinic receptor subtypes, and shares structural homology with other functionally unrelated acetylcholine binding proteins from different species. A layer of tyrosine residues forms an aromatic cap restricting dissociation of the bound ligand. A binding site for allosteric ligands has been mapped to residues at the entrance to the binding pocket near this aromatic cap. The structure of the M2 receptor provides insights into the challenges of developing subtype-selective ligands for muscarinic receptors and their propensity for allosteric regulation.

  12. Muscarinic receptors and amylase secretion of rat pancreatic acini during cerulein-induced acute pancreatitis.

    PubMed

    Morisset, J; Wood, J; Solomon, T E; Larose, L

    1987-08-01

    This study examines the effects of cerulein-induced acute pancreatitis on the secretory response of rat pancreatic acini to carbamylcholine and concentration of acinar muscarinic receptors. Rats were injected subcutaneously every 8 hr with cerulein, 12 micrograms/kg, for two days. They were sacrificed 2 and 4 hr after the first injection, 4 hr after the second and third, and 8 hr after the sixth. By 2 hr after the first injection, carbamylcholine showed decreased potency for stimulating amylase release; decreased potency becomes maximal after the second injection. Four hours after the first injection, carbamylcholine also showed decreased efficacy for causing maximal amylase release. In the course of development of pancreatitis, progressive reductions in muscarinic receptor concentrations were evident from 4 hr after the second injection. Following the complete treatment (8 hr after the sixth injection), no alteration could be observed in the affinity or proportions of each agonist class of muscarinic receptors. These studies indicate that the pancreatic acinar cells still remain functional after acute cerulein-induced pancreatitis, although significant reductions in potency and efficacy of carbamylcholine to cause amylase release and reduced muscarinic receptor concentration occur. PMID:2440647

  13. Quantitative autoradiography of muscarinic and benzodiazepine receptors in the forebrain of the turtle, Pseudemys scripta

    SciTech Connect

    Schlegel, J.R.; Kriegstein, A.R.

    1987-11-22

    The distribution of muscarinic and benzodiazepine receptors was investigated in the turtle forebrain by the technique of in vitro receptor autoradiography. Muscarinic binding sites were labeled with 1 nM /sup 3/H-quinuclidinyl benzilate (/sup 3/H-QNB), and benzodiazepine sites were demonstrated with the aid of 1 nM /sup 3/H-flunitrazepam (/sup 3/H-FLU). Autoradiograms generated on /sup 3/H-Ultrofilm apposed to tissue slices revealed regionally specific distributions of muscarinic and benzodiazepine binding sites that are comparable with those for mammalian brain. Dense benzodiazepine binding was found in the anterior olfactory nucleus, the lateral and dorsal cortices, and the dorsal ventricular ridge (DVR), a structure with no clear mammalian homologue. Muscarinic binding sites were most dense in the striatum, accumbens, DVR, lateral geniculate, and the anterior olfactory nucleus. Cortical binding sites were studied in greater detail by quantitative analysis of autoradiograms generated by using emulsion-coated coverslips. Laminar gradients of binding were observed that were specific for each radioligand; /sup 3/H-QNB sites were most dense in the inner molecular layer in all cortical regions, whereas /sup 3/H-FLU binding was generally most concentrated in the outer molecular layer and was least dense through all layers in the dorsomedial cortex. Because pyramidal cells are arranged in register in turtle cortex, the laminar patterns of receptor binding may reflect different receptor density gradients along pyramidal cell dendrites.

  14. Regional circadian variation of acetylcholine muscarinic receptors in the rat brain

    SciTech Connect

    Por, S.B.; Bondy, S.C.

    1981-01-01

    The level of binding of a labeled acetylcholine muscarinic antagonist (quinuclidinyl benzilate) to different cerebral membranes has been measured. Of the regions examined, circadian rhythmicity of binding could only be detected significantly in the hippocampus and the hypothalamus and not in the cerebral cortex, striatum, or cerebellum.

  15. The effect of oxime reactivators on muscarinic receptors: functional and binding examinations.

    PubMed

    Soukup, O; Kumar, U K; Proska, J; Bratova, L; Adem, A; Jun, D; Fusek, J; Kuca, K; Tobin, G

    2011-05-01

    The antidotal treatment of organophosphorus poisoning is still a problematic issue since no versatile antidote has been developed yet. In our study, we focused on an interesting property, which does not relate to the reactivation of inhibited acetylcholinesterase (AChE) of some oximes, but refers to their anti-muscarinic effects which may contribute considerably to their treatment efficacy. One standard reactivator (HI-6) and two new compounds (K027 and K203) have been investigated for their antimuscarinic properties. Anti-muscarinic effects were studies by means of an in vitro stimulated atrium preparation (functional test), the [(3)H]-QNB binding assay and G-protein coupled receptor assay (GPCR, beta-Arrestin Assay). Based on the functional data HI-6 demonstrates the highest anti-muscarinic effect. However, only when comparing [(3)H]-QNB binding results and GPCR data, K203 shows a very promising compound with regard to anti-muscarinic potency. The therapeutic impact of these findings has been discussed.

  16. Mixed nicotinic and muscarinic features of cholinergic receptor coupled to secretion in bovine chromaffin cells

    SciTech Connect

    Shirvan, M.H.; Pollard, H.B.; Heldman, E. )

    1991-06-01

    Acetylcholine evokes release from cultured bovine chromaffin cells by a mechanism that is believed to be classically nicotinic. However, the authors found that the full muscarinic agonist oxotremorine-M (Oxo-M) induced a robust catecholamine (CA) secretion. By contrast, muscarine, pilocarpine, bethanechol, and McN-A-343 did not elicit any secretory response. Desensitization of the response to nicotine by Oxo-M and desensitization of the response to Oxo-M by nicotine suggest that both nicotine and Oxo-M were acting at the same receptor. Additional experiments supporting this conclusion show that nicotine-induced secretion and Oxo-M-induced secretion were similarly blocked by various muscarinic and nicotinic antagonists. Moreover, secretion induced by nicotine and Oxo-M were Ca{sup 2+} dependent, and both agonists induced {sup 45}Ca{sup 2+} uptake. Equilibrium binding studies showed that ({sup 3}H)Oxo-M bound to chromaffin cell membranes with a K{sub d} value of 3.08 {times} 10{sup {minus}8}M and a Hill coefficient of 1.00, suggesting one binding site for this ligand. Nicotine inhibited Oxo-M binding in a noncompetitive manner, suggesting that both ligands bind at two different sites on the same receptor. They propose that the receptor on bovine chromaffin cells that is coupled to secretion represents an unusual cholinergic receptor that has both nicotinic and muscarinic features.

  17. Mixed nicotinic and muscarinic features of cholinergic receptor coupled to secretion in bovine chromaffin cells.

    PubMed Central

    Shirvan, M H; Pollard, H B; Heldman, E

    1991-01-01

    Acetylcholine evokes release from cultured bovine chromaffin cells by a mechanism that is believed to be classically nicotinic. However, we found that the full muscarinic agonist oxotremorine-M (Oxo-M) induced a robust catecholamine (CA) secretion. By contrast, muscarine, pilocarpine, bethanechol, and McN-A-343 did not elicit any secretory response. Desensitization of the response to nicotine by Oxo-M and desensitization of the response to Oxo-M by nicotine suggest that both nicotine and Oxo-M were acting at the same receptor. Additional experiments supporting this conclusion show that nicotine-induced secretion and Oxo-M-induced secretion were similarly blocked by various muscarinic and nicotinic antagonists. Moreover, secretion induced by nicotine and Oxo-M were Ca2+ dependent, and both agonists induced 45Ca2+ uptake. Equilibrium binding studies showed that [3H]Oxo-M bound to chromaffin cell membranes with a Kd value of 3.08 x 10(-8) M and a Hill coefficient of 1.00, suggesting one binding site for this ligand. Nicotine inhibited Oxo-M binding in a noncompetitive manner, suggesting that both ligands bind at two different sites on the same receptor. We propose that the receptor on bovine chromaffin cells that is coupled to secretion represents an unusual cholinergic receptor that has both nicotinic and muscarinic features. Images PMID:2052567

  18. Muscarinic cholinergic receptors (MR3) in saliva of patients with oral lichen planus.

    PubMed

    Agha-Hosseini, Farzaneh; Mirzaii-Dizgah, Iraj; Mohammadpour, Neda

    2016-09-01

    Oral lichen planus (OLP) is a relatively common, chronic, and inflammatory mucocutaneous disease. Xerostomia is also a common complaint of most OLP patients. Considering the significant role of M3 muscarinic receptors (M3R) in secretion of saliva, this study sought to compare the level of this receptor in saliva between OLP patients and healthy controls. Forty OLP patients and 40 healthy controls filled out two questionnaires regarding xerostomia to assess its degree of severity. Unstimulated and stimulated salivary samples were obtained of both groups and the stimulated and unstimulated salivary flow rates were calculated. Salivary level of M3 muscarinic receptors was measured using the ELISA kit. Data were analyzed and compared using unpaired student's t test. P < 0.05 was considered significant. Stimulated and unstimulated salivary flow rates and M3 muscarinic receptors levels were significantly lower but degree of xerostomia was significantly higher in OLP patients compared to healthy controls. Salivary M3 muscarinic receptor seems to be low in the patients with OLP and these patients suffer from xerostomia and reduced salivary flow rate. PMID:27371099

  19. Monoclonal antibodies to purified muscarinic receptor display agonist-like activity.

    PubMed Central

    Leiber, D; Harbon, S; Guillet, J G; André, C; Strosberg, A D

    1984-01-01

    Monoclonal antibody M-35, which immunoprecipitates native calf brain acetylcholine muscarinic receptor, mimics agonist stimulation of the intact guinea pig myometrium: the antibody, just like carbamoylcholine hydrochloride, causes a rise in intracellular cyclic GMP content, an inhibition of cyclic AMP accumulation due to prostacyclin, and induces uterine contractions. Another antibody, M-23, which reacts with the denatured muscarinic receptor, is devoid of agonist-like activity at the cyclic nucleotide level but is still able to induce contractions of both rat and guinea pig myometrium. The cyclic nucleotide changes caused by both carbamoylcholine and antibody M-35 are inhibited by atropine; this antagonist, which blocks carbamoylcholine-mediated contractions, fails however, to prevent contractions induced by antibodies M-35 and M-23. These results suggest that the information necessary to transmit muscarinic signals is entirely contained in the receptor and that ligands only act to trigger the biological response. The data also imply that the muscarinic receptors of the myometrium are coupled to multiple effector systems. PMID:6087318

  20. Influence of acetylcholine on binding of 4-[125I]iododexetimide to muscarinic brain receptors.

    PubMed

    Weckesser, M; Fixmann, A; Holschbach, M; Müller-Gärtner, H W

    1998-11-01

    The distribution of nicotinic and muscarinic cholinergic receptors in the human brain in vivo has been successfully characterized using radiolabeled tracers and emission tomography. The effect of acetylcholine release into the synaptic cleft on receptor binding of these tracers has not yet been investigated. The present study examined the influence of acetylcholine on binding of 4-[125I]iododexetimide to muscarinic cholinergic receptors of porcine brain synaptosomes in vitro. 4-Iododexetimide is a subtype-unspecific muscarinic receptor antagonist with high affinity. Acetylcholine competed with 4-[125I]iododexetimide in a dose-dependent manner. A concentration of 500 microM acetylcholine inhibited 50% of total specific 4-[125I]iododexetimide binding to synaptosomes when both substances were given simultaneously. An 800 microM acetylcholine solution reduced total specific 4-[125I]iododexetimide binding by about 35%, when acetylcholine was given 60 min after incubation of synaptosomes with 4-[125I]iododexetimide. Variations in the synaptic acetylcholine concentration might influence muscarinic cholinergic receptor imaging in vivo using 4-[123I]iododexetimide. Conversely, 4-[123I]iododexetimide might be an appropriate molecule to investigate alterations of acetylcholine release into the synaptic cleft in vivo using single photon emission computed tomography. PMID:9863566

  1. Does postmenopausal estrogen administration increase the risk of breast cancer? Contributions of animal, biochemical, and clinical investigative studies to a resolution of the controversy.

    PubMed

    Zumoff, B

    1998-01-01

    Despite nearly six decades of epidemiological studies, meta-analyses, and reviews, there is still considerable controversy in the literature about the question, does postmenopausal estrogen administration increase the risk of breast cancer? In an effort to resolve the controversy, a number of animal, biochemical, and clinical investigative studies in this field have been reviewed. The following summary formulation is proposed: 1. Administration of estrogen is inherently capable of promoting the growth of breast cancer, and therefore of increasing the incidence of clinical breast cancer. 2. Human response to estrogen is like that of the low-cancer-incidence strains of mice studied by Lacassagne, in that large doses and prolonged administration are required to induce clinical breast cancer. 3. The blood levels of estradiol produced by the usual doses of postmenopausal estrogen are relatively low, equivalent to those of the follicular phase of the menstrual cycle. These levels may be near the threshold for producing breast-cancer-promoting effects; therefore, the tumor response will vary greatly in different populations, depending on genetic susceptibility factors: a. The prevalence of a family history of premenopausal breast cancer in a first-degree relative. b. The prevalence of abnormal BRCA1, BRCA2, and p53 genes. c. The prevalence of increased 16 alpha-hydroxylation of estradiol. d. The prevalence of smokers who are slow acetylators. 4. Consumption of alcohol (5 grams or more daily) along with the postmenopausal estrogen administration results in elevation of blood estradiol levels to values equivalent to those of the periovulatory peak of the menstrual cycle, which may be well above the threshold for producing breast-cancer-promoting effects in all women. The risk for cancer will therefore be uniformly increased in women who use alcohol and take estrogen. 5. Increased risk of breast cancer from postmenopausal estrogen administration can be eliminated by taking

  2. Increased division of alpha beta TCR+ and gamma delta TCR+ intestinal intraepithelial lymphocytes after oral administration of cholera toxin.

    PubMed Central

    Penney, I; Kilshaw, P J; MacDonald, T T

    1996-01-01

    Cholera toxin (CT) or its subunits were given orally to mice and division of intestinal intraepithelial lymphocytes (IEL) in vivo measured by double immunofluorescence using 5-bromo-2'-deoxyuridine (BRdU) and membrane alpha beta T-cell receptors (TCR) or gamma delta TCR staining in frozen sections. Cholera toxin (10 micrograms) produced a two- to eightfold-increase in the uptake of BRdU in alpha beta TCR+ IEL in the duodenum and a two-to fivefold increase in gamma delta TCR IEL in the ileum. Increased uptake of BRdU was also seen after a dose of 100 micrograms of CT but this dose was also associated with the loss of alpha beta TCR+ IEL and gamma delta TCR+ IEL in the duodenum. CT-A and CT-B subunit produced increased BRdU incorporation by alpha beta TCR in the duodenum and by gamma delta TCR IEL in the ileum. Cholera toxin therefore appears to be mitogenic for IEL probably due to an indirect mechanism. Images Figure 1 PMID:8911140

  3. M1 Muscarinic Receptor Deficiency Attenuates Azoxymethane-Induced Chronic Liver Injury in Mice

    PubMed Central

    Rachakonda, Vikrant; Jadeja, Ravirajsinh N.; Urrunaga, Nathalie H.; Shah, Nirish; Ahmad, Daniel; Cheng, Kunrong; Twaddell, William S.; Raufman, Jean-Pierre; Khurana, Sandeep

    2015-01-01

    Cholinergic nervous system regulates liver injury. However, the role of M1 muscarinic receptors (M1R) in modulating chronic liver injury is uncertain. To address this gap in knowledge we treated M1R-deficient and WT mice with azoxymethane (AOM) for six weeks and assessed liver injury responses 14 weeks after the last dose of AOM. Compared to AOM-treated WT mice, M1R-deficient mice had attenuated liver nodularity, fibrosis and ductular proliferation, α-SMA staining, and expression of α1 collagen, Tgfβ-R, Pdgf-R, Mmp-2, Timp-1 and Timp-2. In hepatocytes, these findings were associated with reductions of cleaved caspase-3 staining and Tnf-α expression. In response to AOM treatment, M1R-deficient mice mounted a vigorous anti-oxidant response by upregulating Gclc and Nqo1 expression, and attenuating peroxynitrite generation. M1R-deficient mouse livers had increased expression of Trail-R2, a promotor of stellate cell apoptosis; dual staining for TUNNEL and α-SMA revealed increased stellate cells apoptosis in livers from M1R-deficient mice compared to those from WT. Finally, pharmacological inhibition of M1R reduced H2O2-induced hepatocyte apoptosis in vitro. These results indicate that following liver injury, anti-oxidant response in M1R-deficient mice attenuates hepatocyte apoptosis and reduces stellate cell activation, thereby diminishing fibrosis. Therefore, targeting M1R expression and activation in chronic liver injury may provide therapeutic benefit. PMID:26374068

  4. Urinary Retention, Incontinence, and Dysregulation of Muscarinic Receptors in Male Mice Lacking Mras

    PubMed Central

    Ehrhardt, Annette; Wang, Bin; Yung, Andrew C.; Wang, Yanni; Kozlowski, Piotr; van Breemen, Cornelis; Schrader, John W.

    2015-01-01

    Here we show that male, but not female mice lacking expression of the GTPase M-Ras developed urinary retention with distention of the bladder that exacerbated with age but occurred in the absence of obvious anatomical outlet obstruction. There were changes in detrusor morphology in Mras-/- males: Smooth muscle tissue, which exhibited a compact organization in WT mice, appeared disorganized and became increasingly ‘layered’ with age in Mras-/- males, but was not fibrotic. Bladder tissue near the apex of bladders of Mras-/- males exhibited hypercontractility in response to the cholinergic agonist carbachol in in vitro, while responses in Mras-/- females were normal. In addition, spontaneous phasic contractions of detrusors from Mras-/- males were increased, and Mras-/- males exhibited urinary incontinence. We found that expression of the muscarinic M2 and M3 receptors that mediate the cholinergic contractile stimuli of the detrusor muscle was dysregulated in both Mras-/- males and females, although only males exhibited a urinary phenotype. Elevated expression of M2R in young males lacking M-Ras and failure to upregulate M3R with age resulted in significantly lower ratios of M3R/M2R expression that correlated with the bladder abnormalities. Our data suggests that M-Ras and M3R are functionally linked and that M-Ras is an important regulator of male bladder control in mice. Our observations also support the notion that bladder control is sexually dimorphic and is regulated through mechanisms that are largely independent of acetylcholine signaling in female mice. PMID:26516777

  5. The restructuring of muscarinic receptor subtype gene transcripts in c-fos knock-out mice.

    PubMed

    Benes, Jan; Mravec, Boris; Kvetnansky, Richard; Myslivecek, Jaromir

    2013-05-01

    Although c-Fos plays a key role in intracellular signalling, the disruption of the c-fos gene has only minor consequences on the central nervous system (CNS) function. As muscarinic receptors (MR) play important roles in many CNS functions (attention, arousal, and cognition), the c-fos knock-out might be compensated through MR changes. The aim of this study was to evaluate changes in the M1-M5 MR mRNA in selected CNS areas: frontal, parietal, temporal and occipital cortex, striatum, hippocampus, hypothalamus and cerebellum (FC, PC, TC, OC, stria, hip, hypo, and crbl, respectively). Knocking out the c-fos gene changed the expression of MR in FC (reduced M1R, M4R and M5R expression), TC (increased M4R expression), OC (decreased M2R and M3R expression) and hippocampus (reduced M3R expression). Moreover, gender differences were observed in WT mice: increased expression of all M1-M5R in the FC in males and M1-M4R in the striatum in females. A detailed analysis of MR transcripts showed pre-existing correlations in the amount of MR-mRNA between specific regions. WT mice showed three major types of cortico-cortical correlations: fronto-occipital, temporo-parietal and parieto-occipital. The cortico-subcortical correlations involved associations between the FC, PC, TC and striatum. In KO mice, a substantial rearrangement of the correlation pattern was observed: only a temporo-parietal correlation and correlations between the FC and striatum remained, and a new correlation between the hypothalamus and cerebellum appeared. Thus, in addition to the previously described dopamine receptor restructuring, the restructuring of MR mRNA correlations reveals an additional mechanism for adaptation to the c-fos gene knockout.

  6. Central amygdala opioid transmission is necessary for increased high-fat intake following 24-h food deprivation, but not following intra-accumbens opioid administration.

    PubMed

    Parker, Kyle E; Johns, Howard W; Floros, Ted G; Will, Matthew J

    2014-03-01

    Previous research has demonstrated a dissociation of certain neural mediators that contribute to the increased consumption of a high-fat diet that follows intra-accumbens (Acb) administration of μ-opioid receptor agonists vs. 24-h food deprivation. These two models, both which induce rapid consumption of the diet, have been shown to involve a distributed corticolimbic circuitry, including the amygdala. Specifically, the central amygdala (CeA) has been shown to be involved in high-fat feeding within both opioid and food-deprivation driven models. The present experiments were conducted to examine the more specific role of CeA opioid transmission in mediating high-fat feeding driven by either intra-Acb administration of the μ-opioid agonist d-Ala2-NMe-Phe4-Glyol5-enkephalin (DAMGO) or 24-h home cage food deprivation. Injection of DAMGO into the Acb (0.25 μg/0.5 μl/side) increased consumption of the high-fat diet, but this feeding was unaffected by administration of opioid antagonist, naltrexone (5 μg/0.25 μl/side) administered into the CeA. In contrast, intra-CeA naltrexone administration attenuated high-fat intake driven by 24-h food deprivation, demonstrating a specific role for CeA opioid transmission in high-fat consumption. Intra-CeA naltrexone administration alone had no effect on baseline feeding levels within either feeding model. These findings suggest that CeA opioid transmission mediates consumption of a palatable high-fat diet driven by short-term negative-energy balance (24-h food deprivation), but not intra-Acb opioid receptor activation.

  7. Repeated methylphenidate administration during lactation reduces maternal behavior, induces maternal tolerance, and increases anxiety-like behavior in pups in adulthood.

    PubMed

    Ponchio, R A; Teodorov, E; Kirsten, T B; Coelho, C P; Oshiro, A; Florio, J C; Bernardi, M M

    2015-01-01

    Methylphenidate (MPD) is a dopamine uptake inhibitor and the most commonly prescribed drug for the treatment of attention-deficit/hyperactivity disorder in children. Several studies have shown that such stimulants as cocaine and amphetamine that are administered during gestation and lactation may disrupt maternal behavior. Also, MPD is used in lactation. Repeated MPD administration can induce either sensitization or tolerance. The aim of the present study was to investigate whether repeated MPD administration alters maternal behavior and promotes tolerance or sensitization in these females. The effects in adult offspring were also examined in models of anxiety. Methylphenidate (5mg/kg) was administered from lactation day 2 to 4, and maternal pup retrieval behavior was assessed. This treatment was continued until lactation day 7. At weaning, the dams received a challenge dose of MPD, and general activity was evaluated in the open field. Striatal monoamine and metabolite levels were also measured to determine whether this treatment promotes behavioral or biochemical plasticity. The long-term behavioral effects of MPD exposure were evaluated in pups in adulthood. The results showed an increase in the latency to retrieve the first, second, and third pups and a decrease in the number of dams that retrieved all pups. After a challenge dose of MPD, the dams exhibited a decrease in locomotion frequency, an increase in immobility duration in the open field, and a decrease in striatal serotonin levels. In pups, anxiety-like behavior increased in the light/dark box test. These results indicate that repeated MPD administration during early lactation impairs maternal behavior, likely by decreasing maternal motivation. Repeated MPD administration induced maternal tolerance at weaning after a challenge dose of MPD, suggesting the development of central nervous system plasticity. In pups, maternal exposure to MPD during early lactation induced long-term effects and increased

  8. Repeated methylphenidate administration during lactation reduces maternal behavior, induces maternal tolerance, and increases anxiety-like behavior in pups in adulthood.

    PubMed

    Ponchio, R A; Teodorov, E; Kirsten, T B; Coelho, C P; Oshiro, A; Florio, J C; Bernardi, M M

    2015-01-01

    Methylphenidate (MPD) is a dopamine uptake inhibitor and the most commonly prescribed drug for the treatment of attention-deficit/hyperactivity disorder in children. Several studies have shown that such stimulants as cocaine and amphetamine that are administered during gestation and lactation may disrupt maternal behavior. Also, MPD is used in lactation. Repeated MPD administration can induce either sensitization or tolerance. The aim of the present study was to investigate whether repeated MPD administration alters maternal behavior and promotes tolerance or sensitization in these females. The effects in adult offspring were also examined in models of anxiety. Methylphenidate (5mg/kg) was administered from lactation day 2 to 4, and maternal pup retrieval behavior was assessed. This treatment was continued until lactation day 7. At weaning, the dams received a challenge dose of MPD, and general activity was evaluated in the open field. Striatal monoamine and metabolite levels were also measured to determine whether this treatment promotes behavioral or biochemical plasticity. The long-term behavioral effects of MPD exposure were evaluated in pups in adulthood. The results showed an increase in the latency to retrieve the first, second, and third pups and a decrease in the number of dams that retrieved all pups. After a challenge dose of MPD, the dams exhibited a decrease in locomotion frequency, an increase in immobility duration in the open field, and a decrease in striatal serotonin levels. In pups, anxiety-like behavior increased in the light/dark box test. These results indicate that repeated MPD administration during early lactation impairs maternal behavior, likely by decreasing maternal motivation. Repeated MPD administration induced maternal tolerance at weaning after a challenge dose of MPD, suggesting the development of central nervous system plasticity. In pups, maternal exposure to MPD during early lactation induced long-term effects and increased

  9. Increased susceptibility to hyperoxic lung injury and alveolar simplification in newborn rats by prenatal administration of benzo[a]pyrene

    PubMed Central

    Thakur, Vijay S.; Liang, Yanhong W.; Lingappan, Krithika; Jiang, Weiwu; Wang, Lihua; Barrios, Roberto; Zhou, Guodong; Guntupalli, Bharath; Shivanna, Binoy; Maturu, Paramahamsa; Welty, Stephen E.; Moorthy, Bhagavatula; Couroucli, Xanthi I.

    2014-01-01

    Maternal smoking is one of the risk factors for preterm birth and for the development of bronchopulmonary dysplasia (BPD). In this study, we tested the hypothesis that prenatal exposure of rats to benzo[a]pyrene (BP), a component of cigarette smoke, will result in increased susceptibility of newborns to oxygen-mediated lung injury and alveolar simplification, and that cytochrome P450 (CYP)1A and 1B1 enzymes and oxidative stress mechanistically contribute to this phenomenon. Timed pregnant Fisher 344 rats were administered BP (25 mg/Kg) or the vehicle corn oil (CO) on gestational days 18, 19 and 20, and newborn were either maintained in room air or exposed to hyperoxia (85% O2) for 7 or 14 days. Hyperoxic newborn rats prenatally exposed to the vehicle CO showed lung injury and alveolar simplification, and inflammation, and these effects were potentiated in rats that were prenatally exposed to BP. Prenatal exposure to BP, followed by hyperoxia, also resulted in significant modulation of hepatic and pulmonary cytochrome P450 (CYP)1A and 1B1 enzymes at PND 7-14. These rats displayed significant oxidative stress in lungs at postnatal day (PND) 14, as evidenced by increased levels of the F2-isoprostane 8-iso-PGF2α. Furthermore, these animals showed BP-derived DNA adducts and oxidative DNA adducts in the lung. In conclusion, our results show increased susceptibility of newborns to oxygen-mediated lung injury and alveolar simplification following maternal exposure to BP, and our results suggest that modulation of CYP1A/1B1 enzymes, increases in oxidative stress, and BP-DNA adducts contributed to this phenomenon. PMID:24657529

  10. Shift Work in Rats Results in Increased Inflammatory Response after Lipopolysaccharide Administration: A Role for Food Consumption.

    PubMed

    Guerrero-Vargas, Natalí N; Guzmán-Ruiz, Mara; Fuentes, Rebeca; García, Joselyn; Salgado-Delgado, Roberto; Basualdo, María del Carmen; Escobar, Carolina; Markus, Regina P; Buijs, Ruud M

    2015-08-01

    The suprachiasmatic nucleus (SCN) drives circadian rhythms in behavioral and physiological variables, including the inflammatory response. Shift work is known to disturb circadian rhythms and is associated with increased susceptibility to develop disease. In rodents, circadian disruption due to shifted light schedules (jet lag) induced increased innate immune responses. To gain more insight into the influence of circadian disruption on the immune response, we characterized the inflammatory response in a model of rodent shift work and demonstrated that circadian disruption affected the inflammatory response to lipopolysaccharide (LPS) both in vivo and in vitro. Since food consumption is a main disturbing element in the shift work schedule, we also evaluated the inflammatory response to LPS in a group of rats that had no access to food during their working hours. Our results demonstrated that the shift work schedule decreased basal TNF-α levels in the liver but not in the circulation. Despite this, we observed that shift work induced increased cytokine response after LPS stimulation in comparison to control rats. Also, Kupffer cells (liver macrophages) isolated from shift work rats produced more TNF-α in response to in vitro LPS stimulation, suggesting important effects of circadian desynchronization on the functionality of this cell type. Importantly, the effects of shift work on the inflammatory response to LPS were prevented when food was not available during the working schedule. Together, these results show that dissociating behavior and food intake from the synchronizing drive of the SCN severely disturbs the immune response. PMID:26017928

  11. Shift Work in Rats Results in Increased Inflammatory Response after Lipopolysaccharide Administration: A Role for Food Consumption.

    PubMed

    Guerrero-Vargas, Natalí N; Guzmán-Ruiz, Mara; Fuentes, Rebeca; García, Joselyn; Salgado-Delgado, Roberto; Basualdo, María del Carmen; Escobar, Carolina; Markus, Regina P; Buijs, Ruud M

    2015-08-01

    The suprachiasmatic nucleus (SCN) drives circadian rhythms in behavioral and physiological variables, including the inflammatory response. Shift work is known to disturb circadian rhythms and is associated with increased susceptibility to develop disease. In rodents, circadian disruption due to shifted light schedules (jet lag) induced increased innate immune responses. To gain more insight into the influence of circadian disruption on the immune response, we characterized the inflammatory response in a model of rodent shift work and demonstrated that circadian disruption affected the inflammatory response to lipopolysaccharide (LPS) both in vivo and in vitro. Since food consumption is a main disturbing element in the shift work schedule, we also evaluated the inflammatory response to LPS in a group of rats that had no access to food during their working hours. Our results demonstrated that the shift work schedule decreased basal TNF-α levels in the liver but not in the circulation. Despite this, we observed that shift work induced increased cytokine response after LPS stimulation in comparison to control rats. Also, Kupffer cells (liver macrophages) isolated from shift work rats produced more TNF-α in response to in vitro LPS stimulation, suggesting important effects of circadian desynchronization on the functionality of this cell type. Importantly, the effects of shift work on the inflammatory response to LPS were prevented when food was not available during the working schedule. Together, these results show that dissociating behavior and food intake from the synchronizing drive of the SCN severely disturbs the immune response.

  12. Direct binding and functional studies on muscarinic cholinoceptors in porcine coronary artery.

    PubMed

    Yamada, S; Yamazawa, T; Nakayama, K

    1990-02-01

    The muscarinic cholinoceptors in porcine coronary artery were identified and characterized by a binding assay using (-)-[3H]quinuclidinyl benzilate (QNB) and also by pharmacological method. Specific (-)-[3H]QNB binding in the coronary artery was saturable and of high affinity (Kd = 0.08 nM), and it showed a pharmacological specificity as well as stereoselectivity which characterized muscarinic receptors. Muscarinic antagonists competed with the (-)-[3H]QNB binding in order: nonlabeled QNB greater than dexetimide greater than atropine greater than pirenzepine greater than AF-DX 116 greater than levetimide greater than gallamine. Dexetimide was approximately 2000 times as potent as levetimide. The potencies (pKi) of these muscarinic antagonists in competing for (-)-[3H]QNB binding sites in porcine coronary artery correlated well with their pharmacological potencies (pA2 for antagonistic effect of acetylcholine-induced contraction of coronary artery). The decrease in the (-)-[3H]QNB binding by atropine and pirenzepine was due to a reduction in the apparent affinity with little change in the number of maximal binding sites, suggesting a competitive antagonism. Specific (-)-[3H]QNB binding (Kd and maximal number of binding sites) in porcine coronary artery was not changed by the removal of endothelium. We conclude: 1) (-)-[3H]QNB selectively labels the physiologically relevant muscarinic receptors in porcine coronary artery and 2) the majority of these receptors is localized on vascular smooth muscles and the receptors mediate the acetylcholine-induced contractile response of coronary artery. PMID:2313599

  13. Muscarinic receptors of the vascular bed: radioligand binding studies on bovine splenic veins.

    PubMed

    Brunner, F; Kukovetz, W R

    1986-01-01

    Despite an obvious lack of parasympathetic innervation to the spleen, pharmacological evidence suggests the presence of cholinergic receptors in isolated bovine splenic veins. We therefore studied muscarinic cholinergic binding sites in a bovine splenic vein preparation by direct radioligand binding techniques using [3H]quinuclidinyl benzilate ([3H]QNB) as radioactive probe. Saturation experiments indicated one homogeneous class of high-affinity binding sites, with a KD of 0.11 nM and a binding site density Bmax of 55 fmol/mg protein. The rate constants at 37 degrees C for formation and dissociation of the [3H]QNB receptor complex were 2.7 X 10(9) M-1 h-1 and 0.38 h-1, respectively, yielding a KD of 0.14 nM. The binding sites showed a high stereospecificity, which was evident from competition experiments with dexetimide (KI = 1.3 nM) and levetimide (KI = 4.6 microM). In competition experiments with muscarinic and nicotinic antagonists and some antidepressants, only one binding site was found, whereas with muscarinic agonists, two binding sites were detected. In the presence of 0.1 mM guanyl-imido-diphosphate, only one binding site could be identified with the muscarinic agonist carbamylcholine. The affinity of [3H]QNB, on the other hand, was slightly decreased, and Bmax values were unchanged. It is concluded that specific, saturable, high-affinity muscarinic binding sites in the bovine splenic vein have been identified and characterized that exhibit properties similar to cholinergic receptors of brain and peripheral tissues and probably mediate acetylcholine-induced relaxation of splenic veins. PMID:2427809

  14. Administration of the non-steroidal anti-inflammatory drug ibuprofen increases macrophage concentrations but reduces necrosis during modified muscle use

    NASA Technical Reports Server (NTRS)

    Cheung, E. V.; Tidball, J. G.

    2003-01-01

    OBJECTIVE: To test the hypothesis that ibuprofen administration during modified muscle use reduces muscle necrosis and invasion by select myeloid cell populations. METHODS: Rats were subjected to hindlimb unloading for 10 days, after which they experienced muscle reloading by normal weight-bearing to induce muscle inflammation and necrosis. Some animals received ibuprofen by intraperitoneal injection 8 h prior to the onset of muscle reloading, and then again at 8 and 16 h following the onset of reloading. Other animals received buffer injection at 8 h prior to reloading and then ibuprofen at 8 and 16 h following the onset of reloading. Control animals received buffer only at each time point. Quantitative immunohistochemical analysis was used to assess the presence of necrotic muscle fibers, total inflammatory infiltrate, neutrophils, ED1+ macrophages and ED2+ macrophages at 24 h following the onset of reloading. RESULT: Administration of ibuprofen beginning 8 h prior to reloading caused significant reduction in the concentration of necrotic fibers, but increased the concentration of inflammatory cells in muscle. The increase in inflammatory cells was attributable to a 2.6-fold increase in the concentration of ED2+ macrophages. Animals treated with ibuprofen 8 h following the onset of reloading showed no decrease in muscle necrosis or increase in ED2+ macrophage concentrations. CONCLUSION: Administration of ibuprofen prior to increased muscle loading reduces muscle damage, but increases the concentration of macrophages that express the ED2 antigen. The increase in ED2+ macrophage concentration and decrease in necrosis may be mechanistically related because ED2+ macrophages have been associated with muscle regeneration and repair.

  15. Amphiphysin I but not dynamin I nor synaptojanin mRNA expression increased after repeated methamphetamine administration in the rat cerebrum and cerebellum.

    PubMed

    Hamamura, Mitsuko; Okouchi, Jiro; Ozawa, Hidetoshi; Kimuro, Yoshihiko; Iwaki, Akiko; Fukumaki, Yasuyuki

    2013-07-01

    Dopamine increases/decreases synaptic vesicle recycling and in schizophrenia the proteins/mRNA is decreased. We isolated cDNA clone, similar to amphiphysin 1 (vesicle protein) mRNA from the neocortex of rats injected repeatedly with methamphetamine using polymerase chain reaction (PCR) differential display. This clone is highly homologous to the 3' region of the human amphiphysin gene. PCR extension study using a primer specific for the rat amphiphysin 1 gene and a primer located within the clone revealed that it is the 3' UTR region of the rat amphiphysin 1 gene. Furthermore, in situ hybridization revealed that amphiphysin 1 mRNA is expressed in the cerebrum, medial thalamus, hippocampus and cerebellum. In the cerebellum, amphiphysin mRNA expression was confined to upper granule cell layer. Repeated methamphetamine administration increased amphiphysin I mRNA expression in both anterior part of the cerebrum, and the cerebellum. However, the repeated administration did not alter mRNA expression of the other vesicle proteins, synaptotagmin I, synapsin I, synaptojanin and dynamin I, we conclude that the repeated administration selectively increased amphiphysin 1 mRNA expression. Thus, amphiphysin 1 does not work as synaptic recycling, but it is suggested, as a part of pathogenesis of brain tissue injury (under Ca²⁺ and Mg²⁺ devoid environment) in repeated methamphetamine-injected states, the gene regulate actin-asssembly, learning, cell stress signaling and cell polarity.

  16. Suspension-Expansion of Bone Marrow Results in Small Mesenchymal Stem Cells Exhibiting Increased Transpulmonary Passage Following Intravenous Administration.

    PubMed

    Zanetti, Andrea; Grata, Michelle; Etling, Emily B; Panday, Regeant; Villanueva, Flordeliza S; Toma, Catalin

    2015-07-01

    Mesenchymal stem cells (MSCs) have been extensively explored in a variety of regenerative medicine applications. The relatively large size of MSCs expanded in tissue culture flasks leads to retention in the microcirculation of the lungs following intravenous delivery, reducing their capacity to reach target sites. We explored whether the expansion of whole marrow in suspension cultures would yield smaller MSCs with increased capacity to traverse the pulmonary microcirculation compared with traditional monolayer cultures. We tested this hypothesis using rat marrow in a suspension bioreactor culture with fibronectin-coated microcarriers, leading to sustained expansion of both the microbead-adherent cells, as well as of a nonadherent cell fraction. Magnetic depletion of CD45(+) cells from the bioreactor cultures after 5 weeks led to a highly enriched CD73(+)/CD90(+)/CD105(+) MSC population. The bioreactor-grown MSCs were significantly smaller than parallel monolayer MSCs (15.1 ± 0.9 μm vs. 18.5 ± 2.3 μm diameter, p<0.05). When fluorescently labeled bioreactor-grown MSCs were intravenously injected into rats, the peak cell concentration in the arterial circulation was an order of magnitude higher than similarly delivered monolayer-grown MSCs (94.8 ± 29.6 vs. 8.2 ± 5.6/10(6) nucleated blood cells, respectively, p<0.05). At 24 h after intravenous injection of the LacZ-labeled bioreactor-grown MSCs, there was a significant threefold decrease in the LacZ-labeled MSCs trapped in the lungs, with a significant increase in the cells reaching the spleen and liver in comparison to their monolayer MSC counterparts. Bioreactor-grown whole marrow cell cultures yielded smaller MSCs with increased capacity to traverse the pulmonary microcirculation compared with traditionally expanded monolayer MSCs. This may significantly improve the capacity and efficiency of these cells to home to injury sites downstream of the lungs. PMID:25567723

  17. Activation of the Endogenous Renin-Angiotensin-Aldosterone System or Aldosterone Administration Increases Urinary Exosomal Sodium Channel Excretion.

    PubMed

    Qi, Ying; Wang, Xiaojing; Rose, Kristie L; MacDonald, W Hayes; Zhang, Bing; Schey, Kevin L; Luther, James M

    2016-02-01

    Urinary exosomes secreted by multiple cell types in the kidney may participate in intercellular signaling and provide an enriched source of kidney-specific proteins for biomarker discovery. Factors that alter the exosomal protein content remain unknown. To determine whether endogenous and exogenous hormones modify urinary exosomal protein content, we analyzed samples from 14 mildly hypertensive patients in a crossover study during a high-sodium (HS, 160 mmol/d) diet and low-sodium (LS, 20 mmol/d) diet to activate the endogenous renin-angiotensin-aldosterone system. We further analyzed selected exosomal protein content in a separate cohort of healthy persons receiving intravenous aldosterone (0.7 μg/kg per hour for 10 hours) versus vehicle infusion. The LS diet increased plasma renin activity and aldosterone concentration, whereas aldosterone infusion increased only aldosterone concentration. Protein analysis of paired urine exosome samples by liquid chromatography-tandem mass spectrometry-based multidimensional protein identification technology detected 2775 unique proteins, of which 316 exhibited significantly altered abundance during LS diet. Sodium chloride cotransporter (NCC) and α- and γ-epithelial sodium channel (ENaC) subunits from the discovery set were verified using targeted multiple reaction monitoring mass spectrometry quantified with isotope-labeled peptide standards. Dietary sodium restriction or acute aldosterone infusion similarly increased urine exosomal γENaC[112-122] peptide concentrations nearly 20-fold, which correlated with plasma aldosterone concentration and urinary Na/K ratio. Urine exosomal NCC and αENaC concentrations were relatively unchanged during these interventions. We conclude that urinary exosome content is altered by renin-angiotensin-aldosterone system activation. Urinary measurement of exosomal γENaC[112-122] concentration may provide a useful biomarker of ENaC activation in future clinical studies.

  18. Association of Opioids and Sedatives with Increased Risk of In-Hospital Cardiopulmonary Arrest from an Administrative Database

    PubMed Central

    Overdyk, Frank J.; Dowling, Oonagh; Marino, Joseph; Qiu, Jiejing; Chien, Hung-Lun; Erslon, Mary; Morrison, Neil; Harrison, Brooke; Dahan, Albert; Gan, Tong J.

    2016-01-01

    Background While opioid use confers a known risk for respiratory depression, the incremental risk of in-hospital cardiopulmonary arrest, respiratory arrest, or cardiopulmonary resuscitation (CPRA) has not been studied. Our aim was to investigate the prevalence, outcomes, and risk profile of in-hospital CPRA for patients receiving opioids and medications with central nervous system sedating side effects (sedatives). Methods A retrospective analysis of adult inpatient discharges from 2008–2012 reported in the Premier Database. Patients were grouped into four mutually exclusive categories: (1) opioids and sedatives, (2) opioids only, (3) sedatives only, and (4) neither opioids nor sedatives. Results Among 21,276,691 inpatient discharges, 53% received opioids with or without sedatives. A total of 96,554 patients suffered CPRA (0.92 per 1000 hospital bed-days). Patients who received opioids and sedatives had an adjusted odds ratio for CPRA of 3.47 (95% CI: 3.40–3.54; p<0.0001) compared with patients not receiving opioids or sedatives. Opioids alone and sedatives alone were associated with a 1.81-fold and a 1.82-fold (p<0.0001 for both) increase in the odds of CPRA, respectively. In opioid patients, locations of CPRA were intensive care (54%), general care floor (25%), and stepdown units (15%). Only 42% of patients survived CPRA and only 22% were discharged home. Opioid patients with CPRA had mean increased hospital lengths of stay of 7.57 days and mean increased total hospital costs of $27,569. Conclusions Opioids and sedatives are independent and additive risk factors for in-hospital CPRA. The impact of opioid sparing analgesia, reduced sedative use, and better monitoring on CPRA incidence deserves further study. PMID:26913753

  19. Role of M2 Muscarinic Receptor in the Airway Response to Methacholine of Mice Selected for Minimal or Maximal Acute Inflammatory Response

    PubMed Central

    Castro, Juciane Maria de Andrade; Resende, Rodrigo R.; Florsheim, Esther; Albuquerque, Layra Lucy; Lino-dos-Santos-Franco, Adriana; Gomes, Eliane; Tavares de Lima, Wothan; de Franco, Marcelo; Ribeiro, Orlando Garcia

    2013-01-01

    Airway smooth muscle constriction induced by cholinergic agonists such as methacholine (MCh), which is typically increased in asthmatic patients, is regulated mainly by muscle muscarinic M3 receptors and negatively by vagal muscarinic M2 receptors. Here we evaluated basal (intrinsic) and allergen-induced (extrinsic) airway responses to MCh. We used two mouse lines selected to respond maximally (AIRmax) or minimally (AIRmin) to innate inflammatory stimuli. We found that in basal condition AIRmin mice responded more vigorously to MCh than AIRmax. Treatment with a specific M2 antagonist increased airway response of AIRmax but not of AIRmin mice. The expression of M2 receptors in the lung was significantly lower in AIRmin compared to AIRmax animals. AIRmax mice developed a more intense allergic inflammation than AIRmin, and both allergic mouse lines increased airway responses to MCh. However, gallamine treatment of allergic groups did not affect the responses to MCh. Our results confirm that low or dysfunctional M2 receptor activity is associated with increased airway responsiveness to MCh and that this trait was inherited during the selective breeding of AIRmin mice and was acquired by AIRmax mice during allergic lung inflammation. PMID:23691511

  20. Continuous administration of heparin in patients with deep vein thrombosis can increase plasma levels of diamine oxidase.

    PubMed

    Klocker, Josef; Perkmann, Reinhold; Klein-Weigel, Peter; Mörsdorf, Gabriele; Drasche, Astrid; Klingler, Anton; Fraedrich, Gustav; Schwelberger, Hubert G

    2004-01-01

    Besides its anticoagulant activity, the sulfated polysaccharide heparin has numerous other biological effects. Especially the antiinflammatory and immunoregulatory properties of heparin may be associated with its ability to release the histamine-degrading enzyme diamine oxidase (DAO) from tissue-bound sites into the circulation. Whereas DAO activity is at the limits of detection in normal human plasma, the application of heparin leads to a significant increase of plasma DAO activity. However, previously, only the effect of bolus injection of unfractionated heparin (UFH) had been studied. To investigate DAO release during continuous heparin infusion, 28 patients with deep vein thrombosis (DVT) undergoing heparin therapy were analyzed. Whereas continuous heparin infusion did not lead to any increase of plasma DAO activity in 12 patients (43%), 6 patients (21%) showed a single elevated and 10 patients (36%) permanently elevated plasma DAO activity. The groups of patients exhibiting different DAO release responses did not differ in age, sex, body weight, concomitant diseases, heparin infusion rates, coagulation indices, location and extension of thrombosis, or clinical outcome. However, the rate of idiopathic DVT was significantly higher in the group of patients releasing DAO. This study shows, for the first time, that continuous heparin infusion can lead to DAO release and that individuals exhibit considerable differences in their release response. Although the significance of heparin-induced DAO release needs further clarification, our results indicate that postheparin plasma DAO activity could be an interesting parameter correlated with idiopathic DVT.

  1. Muscarinic and opioid receptor modulation of release of (Met/sup 5/-enkephalin immunoreactive material and catecholamines from the bovine adrenal gland

    SciTech Connect

    Barron, B.A.

    1985-01-01

    Retrogradely perfused bovine adrenal glands were stimulated by acetylcholine (ACh) and 1,1-dimethyl-4-phenyl-piperazinium (DMPP), with or without: hexamethonium (C-6), atropine, imipramine, methacholine, pilocarpine, etorphine, or diprenorphine. Stimulation by either ACh DMPP resulted in an increased release of both (Met/sup 5/)-enkephalin immunoreactive material (ME-IRM) and catecholamines as measured by radioimmunoassay and high performance liquid chromatography with electrochemical detection, respectively. ACh (5 x 10/sup -5/ M) and DMPP (5 x 10/sup -5/ M) stimulated the release of norepinephrine greater than the release of epinephrine. The action of these agents was antagonized by C-6(5 x 10/sup -4/ M). Atropine (5 x 10/sup -7/ M) antagonized the action of ACh to stimulate norepinephrine and MI-IRM release while having no effect on DMPP-stimulated release. Imipramine (5 x 10/sup -6/ M) had no effect on either ACh or DMPP-stimulated release. Methacholine (4 x 10/sup -5/ M) potentiated the DMPP (1 x 10/sup -5/ M) stimulation of ME-IRM and catecholamine release; pilocarpine (4 x 10/sup -5/ M) significantly potentiated only the DMPP-stimulated release of norepinephrine. Pilocarpine (5 x 10/sup -5/ M) and muscarine (5 x 10/sup -5/ M) had no effect on the secretion of MI-IRM and catecholamines from the bovine adrenal gland. Etorphine (5 x 10/sup -7/ M) significantly decreased the ACh and DMPP stimulation ME-IRM and catecholamine release. The activity of a muscarinic cholinergic receptor in the bovine adrenal medulla in stimulus-secretion coupling has been controversial. The binding of /sup 3/H-quinuclidinyl benzilate to chromaffin granule membranes was investigated to further characterize muscarinic receptors in the bovine adrenal gland.

  2. Increased CD4+ T cell levels during IL-7 administration of antiretroviral therapy-treated simian immunodeficiency virus-positive macaques are not dependent on strong proliferative responses.

    PubMed

    Leone, Amanda; Rohankhedkar, Mukta; Okoye, Afam; Legasse, Alfred; Axthelm, Michael K; Villinger, Francois; Piatak, Michael; Lifson, Jeffrey D; Assouline, Brigitte; Morre, Michel; Picker, Louis J; Sodora, Donald L

    2010-08-01

    CD4(+) T cell depletion is a fundamental component of HIV infection and AIDS pathogenesis and is not always reversed following antiretroviral therapy (ART). In this study, the SIV-infected rhesus macaque model was used to assess recombinant simian IL-7 in its glycosylated form (rsIL-7gly) to enhance regeneration of CD4(+) T cells, particularly the crucial central memory compartment, after ART. We assessed the impact of rsIL-7gly administration as single injections and as a cluster of three doses. Irrespective of the dosing strategy used, the rsIL-7gly administration transiently increased proliferation of both central memory and naive cells, in both CD4(+) and CD8(+) subsets, without increasing SIV levels in the blood. Administration of rsIL-7gly at intervals of 4-6 wk maximized the proliferative response to therapy but resulted in only transient increases in peripheral blood T cell counts. Although more frequent rsIL-7gly "clustered" dosing (three times weekly with 2 wk of rest and then repeat) induced only an initial proliferative burst by CD4(+) T cells, this dosing strategy resulted in sustained increases in peripheral blood CD4(+) T cell counts. The clustered rsIL-7gly treatment regimen was shown to increase the half-life of a BrdU label among memory T cells in the blood when compared with that of macaques treated with ART alone, which is consistent with enhanced cell survival. These results indicate that dosing intervals have a major impact on the response to rsIL-7gly in SIV-positive ART-treated rhesus macaques and that optimum dosing strategies may be ones that induce CD4(+) T cell proliferation initially and provide increased CD4(+) T cell survival.

  3. Potentiation of Muscarinic and α -adrenergic Responses by an Analogue of Guanosine 5'-triphosphate

    NASA Astrophysics Data System (ADS)

    Evans, M. G.; Marty, A.

    1986-06-01

    Ca2+-dependent K+ and Cl- currents were recorded in isolated and dialyzed rat lacrimal gland cells by use of the tight-seal whole-cell recording technique. Under control conditions, application of acetylcholine (0.5-1.0 μ M) resulted in the full activation of both types of current. When 50-200 μ M guanosine 5'-[γ -thio]triphosphate (GTP[S], a nonhydrolyzable GTP analogue) was added to the intracellular solution, activation of both currents was seen with 1 nM acetylcholine, a dose 1/100th that needed under control conditions. Dialysis with solutions containing 200 μ M GTP or cAMP had no, or only slight, potentiation effects. The effects of GTP[S] were obtained only when ATP was included in the intracellular solution. The potentiated responses to acetylcholine were blocked by increasing 10-fold the intracellular Ca2+-buffering capacity and were not dependent on external Ca2+. Thus, the potentiated responses appeared to result from a release of Ca2+ from internal stores. GTP[S] also greatly potentiated the Ca2+-dependent adrenergic (norepinephrine) response of this preparation. In addition, GTP[S] elicited in some cells transient responses without application of acetylcholine or norepinephrine. Finally, rapid and sustained responses were seen as soon as the cells were dialyzed with inositol trisphosphate (20 μ M). These findings are discussed in terms of a possible role of a GTP-binding protein as a link between activation of muscarinic or adrenergic receptors and initiation of Ca2+ release by inositol trisphosphate.

  4. The Role of Hippocampal NMDA Receptors in Long-Term Emotional Responses following Muscarinic Receptor Activation.

    PubMed

    Hoeller, Alexandre A; Costa, Ana Paula R; Bicca, Maíra A; Matheus, Filipe C; Lach, Gilliard; Spiga, Francesca; Lightman, Stafford L; Walz, Roger; Collingridge, Graham L; Bortolotto, Zuner A; de Lima, Thereza C M

    2016-01-01

    Extensive evidence indicates the influence of the cholinergic system on emotional processing. Previous findings provided new insights into the underlying mechanisms of long-term anxiety, showing that rats injected with a single systemic dose of pilocarpine--a muscarinic receptor (mAChR) agonist--displayed persistent anxiogenic-like responses when evaluated in different behavioral tests and time-points (24 h up to 3 months later). Herein, we investigated whether the pilocarpine-induced long-term anxiogenesis modulates the HPA axis function and the putative involvement of NMDA receptors (NMDARs) following mAChRs activation. Accordingly, adult male Wistar rats presented anxiogenic-like behavior in the elevated plus-maze (EPM) after 24 h or 1 month of pilocarpine injection (150 mg/kg, i.p.). In these animals, mAChR activation disrupted HPA axis function inducing a long-term increase of corticosterone release associated with a reduced expression of hippocampal GRs, as well as consistently decreased NMDAR subunits expression. Furthermore, in another group of rats injected with memantine--an NMDARs antagonist (4 mg/kg, i.p.)--prior to pilocarpine, we found inhibition of anxiogenic-like behaviors in the EPM but no further alterations in the pilocarpine-induced NMDARs downregulation. Our data provide evidence that behavioral anxiogenesis induced by mAChR activation effectively yields short- and long-term alterations in hippocampal NMDARs expression associated with impairment of hippocampal inhibitory regulation of HPA axis activity. This is a novel mechanism associated with anxiety-like responses in rats, which comprise a putative target to future translational studies. PMID:26795565

  5. Mechanistic insights into allosteric structure-function relationships at the M1 muscarinic acetylcholine receptor.

    PubMed

    Abdul-Ridha, Alaa; Lane, J Robert; Mistry, Shailesh N; López, Laura; Sexton, Patrick M; Scammells, Peter J; Christopoulos, Arthur; Canals, Meritxell

    2014-11-28

    Benzylquinolone carboxylic acid (BQCA) is the first highly selective positive allosteric modulator (PAM) for the M1 muscarinic acetylcholine receptor (mAChR), but it possesses low affinity for the allosteric site on the receptor. More recent drug discovery efforts identified 3-((1S,2S)-2-hydroxycyclohexyl)-6-((6-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)methyl)benzo[h]quinazolin-4(3H)-one (referred to herein as benzoquinazolinone 12) as a more potent M1 mAChR PAM with a structural ancestry originating from BQCA and related compounds. In the current study, we optimized the synthesis of and fully characterized the pharmacology of benzoquinazolinone 12, finding that its improved potency derived from a 50-fold increase in allosteric site affinity as compared with BQCA, while retaining a similar level of positive cooperativity with acetylcholine. We then utilized site-directed mutagenesis and molecular modeling to validate the allosteric binding pocket we previously described for BQCA as a shared site for benzoquinazolinone 12 and provide a molecular basis for its improved activity at the M1 mAChR. This includes a key role for hydrophobic and polar interactions with residues Tyr-179, in the second extracellular loop (ECL2) and Trp-400(7.35) in transmembrane domain (TM) 7. Collectively, this study highlights how the properties of affinity and cooperativity can be differentially modified on a common structural scaffold and identifies molecular features that can be exploited to tailor the development of M1 mAChR-targeting PAMs. PMID:25326383

  6. The Role of Hippocampal NMDA Receptors in Long-Term Emotional Responses following Muscarinic Receptor Activation.

    PubMed

    Hoeller, Alexandre A; Costa, Ana Paula R; Bicca, Maíra A; Matheus, Filipe C; Lach, Gilliard; Spiga, Francesca; Lightman, Stafford L; Walz, Roger; Collingridge, Graham L; Bortolotto, Zuner A; de Lima, Thereza C M

    2016-01-01

    Extensive evidence indicates the influence of the cholinergic system on emotional processing. Previous findings provided new insights into the underlying mechanisms of long-term anxiety, showing that rats injected with a single systemic dose of pilocarpine--a muscarinic receptor (mAChR) agonist--displayed persistent anxiogenic-like responses when evaluated in different behavioral tests and time-points (24 h up to 3 months later). Herein, we investigated whether the pilocarpine-induced long-term anxiogenesis modulates the HPA axis function and the putative involvement of NMDA receptors (NMDARs) following mAChRs activation. Accordingly, adult male Wistar rats presented anxiogenic-like behavior in the elevated plus-maze (EPM) after 24 h or 1 month of pilocarpine injection (150 mg/kg, i.p.). In these animals, mAChR activation disrupted HPA axis function inducing a long-term increase of corticosterone release associated with a reduced expression of hippocampal GRs, as well as consistently decreased NMDAR subunits expression. Furthermore, in another group of rats injected with memantine--an NMDARs antagonist (4 mg/kg, i.p.)--prior to pilocarpine, we found inhibition of anxiogenic-like behaviors in the EPM but no further alterations in the pilocarpine-induced NMDARs downregulation. Our data provide evidence that behavioral anxiogenesis induced by mAChR activation effectively yields short- and long-term alterations in hippocampal NMDARs expression associated with impairment of hippocampal inhibitory regulation of HPA axis activity. This is a novel mechanism associated with anxiety-like responses in rats, which comprise a putative target to future translational studies.

  7. Mechanistic Insights into Allosteric Structure-Function Relationships at the M1 Muscarinic Acetylcholine Receptor*

    PubMed Central

    Abdul-Ridha, Alaa; Lane, J. Robert; Mistry, Shailesh N.; López, Laura; Sexton, Patrick M.; Scammells, Peter J.; Christopoulos, Arthur; Canals, Meritxell

    2014-01-01

    Benzylquinolone carboxylic acid (BQCA) is the first highly selective positive allosteric modulator (PAM) for the M1 muscarinic acetylcholine receptor (mAChR), but it possesses low affinity for the allosteric site on the receptor. More recent drug discovery efforts identified 3-((1S,2S)-2-hydroxycyclohexyl)-6-((6-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)methyl)benzo[h]quinazolin-4(3H)-one (referred to herein as benzoquinazolinone 12) as a more potent M1 mAChR PAM with a structural ancestry originating from BQCA and related compounds. In the current study, we optimized the synthesis of and fully characterized the pharmacology of benzoquinazolinone 12, finding that its improved potency derived from a 50-fold increase in allosteric site affinity as compared with BQCA, while retaining a similar level of positive cooperativity with acetylcholine. We then utilized site-directed mutagenesis and molecular modeling to validate the allosteric binding pocket we previously described for BQCA as a shared site for benzoquinazolinone 12 and provide a molecular basis for its improved activity at the M1 mAChR. This includes a key role for hydrophobic and polar interactions with residues Tyr-179, in the second extracellular loop (ECL2) and Trp-4007.35 in transmembrane domain (TM) 7. Collectively, this study highlights how the properties of affinity and cooperativity can be differentially modified on a common structural scaffold and identifies molecular features that can be exploited to tailor the development of M1 mAChR-targeting PAMs. PMID:25326383

  8. Upregulation of M3 muscarinic receptor inhibits cardiac hypertrophy induced by angiotensin II

    PubMed Central

    2013-01-01

    Background M3 muscarinic acetylcholine receptor (M3-mAChR) is stably expressed in the myocardium, but its pathophysiological role remains largely undefined. This study aimed to investigate the role of M3-mAChR in cardiac hypertrophy induced by angiotensin II (Ang II) and elucidate the underlying mechanisms. Methods Cardiac-specific M3-mAChR overexpression transgenic (TG) mice and rat H9c2 cardiomyoblasts with ectopic expression of M3-mAChR were established. Models of cardiac hypertrophy were induced by transverse aortic constriction (TAC) or Ang II infusion in the mice in vivo, and by isoproterenol (ISO) or Ang II treatment of H9c2 cells in vitro. Cardiac hypertrophy was evaluated by electrocardiography (ECG) measurement, hemodynamic measurement and histological analysis. mRNA and protein expression were detected by real-time RT-PCR and Western blot analysis. Results M3-mAChR was upregulated in hypertrophic heart, while M2-mAChR expression did not change significantly. M3-mAChR overexpression significantly attenuated the increased expression of atrial natriuretic peptide and β-myosin heavy chain induced by Ang II both in vivo and in vitro. In addition, M3-mAChR overexpression downregulated AT1 receptor expression and inhibited the activation of MAPK signaling in the heart. Conclusion The upregulation of M3-mAChR during myocardial hypertrophy could relieve the hypertrophic response provoked by Ang II, and the mechanism may involve the inhibition of MAPK signaling through the downregulation of AT1 receptor. PMID:24028210

  9. Acute Ethanol Administration Rapidly Increases Phosphorylation of Conventional Protein Kinase C in Specific Mammalian Brain Regions in Vivo

    PubMed Central

    Wilkie, Mary Beth; Besheer, Joyce; Kelley, Stephen P.; Kumar, Sandeep; O’Buckley, Todd K.; Morrow, A. Leslie; Hodge, Clyde W.

    2010-01-01

    Background Protein kinase C (PKC) is a family of isoenzymes that regulate a variety of functions in the central nervous system including neurotransmitter release, ion channel activity, and cell differentiation. Growing evidence suggests that specific isoforms of PKC influence a variety of behavioral, biochemical, and physiological effects of ethanol in mammals. The purpose of this study was to determine whether acute ethanol exposure alters phosphorylation of conventional PKC isoforms at a threonine 674 (p-cPKC) site in the hydrophobic domain of the kinase, which is required for its catalytic activity. Methods Male rats were administered a dose range of ethanol (0, 0.5, 1, or 2 g/kg, intragastric) and brain tissue was removed 10 minutes later for evaluation of changes in p-cPKC expression using immunohistochemistry and Western blot methods. Results Immunohistochemical data show that the highest dose of ethanol (2 g/kg) rapidly increases p-cPKC immunoreactivity specifically in the nucleus accumbens (core and shell), lateral septum, and hippocampus (CA3 and dentate gyrus). Western blot analysis further showed that ethanol (2 g/kg) increased p-cPKC expression in the P2 membrane fraction of tissue from the nucleus accumbens and hippocampus. Although p-cPKC was expressed in numerous other brain regions, including the caudate nucleus, amygdala, and cortex, no changes were observed in response to acute ethanol. Total PKCγ immunoreactivity was surveyed throughout the brain and showed no change following acute ethanol injection. Conclusions These results suggest that ethanol rapidly promotes phosphorylation of cPKC in limbic brain regions, which may underlie effects of acute ethanol on the nervous system and behavior. PMID:17511744

  10. Significant increase of salivary testosterone levels after single therapeutic transdermal administration of testosterone: suitability as a potential screening parameter in doping control.

    PubMed

    Thieme, Detlef; Rautenberg, Claudia; Grosse, Joachim; Schoenfelder, Martin

    2013-01-01

    The legally defensible proof of the abuse of endogenous steroids in sports is currently based on carbon isotope ratio mass spectrometry (IRMS), i.e. a comparison between (13)C/(12)C ratios of diagnostic precursors and metabolites of testosterone. The application of this technique requires a chromatographic baseline separation of respective steroids prior to IRMS detection and hence laborious sample pre-processing of the urinary steroid extracts including clean up by solid-phase extraction and/or liquid chromatography. Consequently, an efficient pre-selection of suspicious control urine samples is essential for appropriate follow up confirmation by IRMS and effective doping control. Two single transdermal administration studies of testosterone (50 mg Testogel® and Testopatch® at 3.8 mg in 16 h, respectively) were conducted and resulting profiles of salivary testosterone and urinary steroid profiles and corresponding carbon isotope ratios were determined. Conventional doping control markers (testosterone/epitestosterone ratio, threshold concentrations of androsterone, etiocholanolone, or androstanediols) did not approach or exceed critical thresholds. In contrast to these moderate variations, the testosterone concentration in oral fluid increased from basal values (30-142 pg/mg) to peak concentrations above 1000 pg/mg. It is likely that this significant increase in oral fluid is due to a pulsatile elevation of free (protein unbound) circulating testosterone after transdermal administration and may be assumed to represent a more diagnostic marker for transdermal testosterone administration.

  11. Effects of temperature and ethanol on agonist and antagonist binding to rat heart muscarinic receptors in the absence and presence of GTP.

    PubMed Central

    Waelbroeck, M; Robberecht, P; Chatelain, P; De Neef, P; Christophe, J

    1985-01-01

    The effect of temperature on the binding of four agonists and three antagonists to rat heart muscarinic receptors was studied in the absence and presence of GTP. The binding of agonists to two states (or classes) of receptors, in the absence of GTP, led to enthalpy and entropy changes that decreased sharply above 25 degrees C, suggesting that agonists induced 'isomerization' reactions (large conformational changes and/or receptor-effector association). Both temperature increase and ethanol decreased hydrophobic interactions, thereby hindering binding and/or agonist-induced 'isomerization' reactions. Addition of GTP to the incubation medium also appeared to reverse (or prevent) 'isomerization' reactions. For agonist binding to the low-affinity state, in the presence of GTP, and for antagonist binding, the thermodynamic parameters observed could be readily explained by simple receptor-ligand associations; large entropy increases and small enthalpy increases, provoked by hydrophobic and ionic interactions, were partly neutralized by entropy and enthalpy decreases, due to hydrogen bonds and van der Waals interactions. The muscarinic antagonists used (atropine, n-methylscopolamine and dexetimide), being more hydrophobic molecules than the agonists tested (carbamylcholine, oxotremorine and pilocarpine), induced larger entropy changes or more negative enthalpy changes. PMID:4062907

  12. Effects of temperature and ethanol on agonist and antagonist binding to rat heart muscarinic receptors in the absence and presence of GTP.

    PubMed

    Waelbroeck, M; Robberecht, P; Chatelain, P; De Neef, P; Christophe, J

    1985-10-15

    The effect of temperature on the binding of four agonists and three antagonists to rat heart muscarinic receptors was studied in the absence and presence of GTP. The binding of agonists to two states (or classes) of receptors, in the absence of GTP, led to enthalpy and entropy changes that decreased sharply above 25 degrees C, suggesting that agonists induced 'isomerization' reactions (large conformational changes and/or receptor-effector association). Both temperature increase and ethanol decreased hydrophobic interactions, thereby hindering binding and/or agonist-induced 'isomerization' reactions. Addition of GTP to the incubation medium also appeared to reverse (or prevent) 'isomerization' reactions. For agonist binding to the low-affinity state, in the presence of GTP, and for antagonist binding, the thermodynamic parameters observed could be readily explained by simple receptor-ligand associations; large entropy increases and small enthalpy increases, provoked by hydrophobic and ionic interactions, were partly neutralized by entropy and enthalpy decreases, due to hydrogen bonds and van der Waals interactions. The muscarinic antagonists used (atropine, n-methylscopolamine and dexetimide), being more hydrophobic molecules than the agonists tested (carbamylcholine, oxotremorine and pilocarpine), induced larger entropy changes or more negative enthalpy changes. PMID:4062907

  13. Acute Central Neuropeptide Y Administration Increases Food Intake but Does Not Affect Hepatic Very Low-Density Lipoprotein (Vldl) Production in Mice

    PubMed Central

    Havekes, Louis M.; Romijn, Johannes A.; Rensen, Patrick C. N.

    2013-01-01

    Objective Central neuropeptide Y (NPY) administration stimulates food intake in rodents. In addition, acute modulation of central NPY signaling increases hepatic production of very low-density lipoprotein (VLDL)-triglyceride (TG) in rats. As hypertriglyceridemia is an important risk factor for atherosclerosis, for which well-established mouse models are available, we set out to validate the effect of NPY on hepatic VLDL-TG production in mice, to ultimately investigate whether NPY, by increasing VLDL production, contributes to the development of atherosclerosis. Research Design and Methods Male C57Bl/6J mice received an intracerebroventricular (i.c.v.) cannula into the lateral (LV) or third (3V) ventricle of the brain. One week later, after a 4 h fast, the animals received an intravenous (i.v.) injection of Tran35S (100 µCi) followed by tyloxapol (500 mg/kg body weight; BW), enabling the study of hepatic VLDL-apoB and VLDL-TG production, respectively. Immediately after the i.v. injection of tyloxapol, the animals received either an i.c.v. injection of NPY (0.2 mg/kg BW in artificial cerebrospinal fluid; aCSF), synthetic Y1 receptor antagonist GR231118 (0.5 mg/kg BW in aCSF) or vehicle (aCSF), or an i.v. injection of PYY3–36 (0.5 mg/kg BW in PBS) or vehicle (PBS). Results Administration of NPY into both the LV and 3V increased food intake within one hour after injection (+164%, p<0.001 and +367%, p<0.001, respectively). NPY administration neither in the LV nor in the 3V affected hepatic VLDL-TG or VLDL-apoB production. Likewise, antagonizing central NPY signaling by either PYY3–36 or GR231118 administration did not affect hepatic VLDL production. Conclusion In mice, as opposed to rats, acute central administration of NPY increases food intake without affecting hepatic VLDL production. These results are of great significance when extrapolating findings on the central regulation of hepatic VLDL production between species. PMID:23460782

  14. Increased platelet aggregation and in vivo platelet activation after granulocyte colony-stimulating factor administration. A randomised controlled trial.

    PubMed

    Spiel, Alexander O; Bartko, Johann; Schwameis, Michael; Firbas, Christa; Siller-Matula, Jolanta; Schuetz, Matthias; Weigl, Manuela; Jilma, Bernd

    2011-04-01

    Granulocyte colony-stimulating factor (G-CSF) stimulates the bone marrow to produce granulocytes and stem cells and is widely used to accelerate neutrophil recovery after chemotherapy. Interestingly, specific G-CSF receptors have been demonstrated not only on myeloid cells, but also on platelets. Data on the effects of G-CSF on platelet function are limited and partly conflicting. The objective of this study was to determine the effect of G-CSF on platelet aggregation and in vivo platelet activation. Seventy-eight, healthy volunteers were enrolled into this randomised, placebo-controlled trial. Subjects received 5 μg/kg methionyl human granulocyte colony-stimulating factor (r-metHuG-CSF, filgrastim) or placebo subcutaneously for four days. We determined platelet aggregation with a whole blood impedance aggregometer with various, clinically relevant platelet agonists (adenosine diphosphate [ADP], collagen, arachidonic acid [AA], ristocetin and thrombin receptor activating peptide 6 [TRAP]). Filgrastim injection significantly enhanced ADP (+40%), collagen (+60%) and AA (+75%)-induced platelet aggregation (all p<0.01 as compared to placebo and p<0.001 as compared to baseline). In addition, G-CSF enhanced ristocetin-induced platelet aggregation (+18%) whereas TRAP-induced platelet aggregation decreased slightly (-14%) in response to filgrastim. While baseline aggregation with all agonists was only slightly but insignificantly higher in women than in men, this sex difference was enhanced by G-CSF treatment, and became most pronounced for ADP after five days (p<0.001). Enhanced platelet aggregation translated into a 75% increase in platelet activation as measured by circulating soluble P-selectin. G-CSF enhances platelet aggregation and activation in humans. This may put patients suffering from cardiovascular disease and cancer at risk for thrombotic events. PMID:21301783

  15. Increased docosahexaenoic acid levels in human newborn infants by administration of sardines and fish oil during pregnancy.

    PubMed

    Connor, W E; Lowensohn, R; Hatcher, L

    1996-03-01

    In rhesus monkeys, maternal n-3 fatty acid deficiency during pregnancy produces infant monkeys deficient in n-3 fatty acids at birth. These results stimulated current experiments to find out if n-3 fatty acids from fish in the diets of pregnant women would influence the concentration of docosahexaenoic acid (DHA, 22:6 n-3) in the newborn human infant. Fifteen healthy pregnant women were enrolled to receive a 9-wk dietary supplementation of n-3 fatty acids from the 26th to the 35th wk of pregnancy. Sixteen pregnant women were not supplemented and served as controls. n-3 Fatty acid supplementation consisted of sardines and additional fish oil, which provided a total of 2.6 g of n-3 fatty acids per day (d) for the 9-wk period of supplementation. This included 1.01 g DHA. The end point of this study was the blood concentrations of DHA in the newborn infant. DHA in maternal red blood cells increased from 4.6% of total fatty acids to 7.15% at the end of the supplement period and at the time of delivery decreased (as expected) to 5.97% of total fatty acids. Maternal plasma showed a similar change from 2.12 to 3.51% of total fatty acids and then decreased to 2.35%. Levels of DHA in plasma and red blood cells of unsupplemented mothers did not change during the same time period. Levels of DHA in blood of newborn infants differed greatly in infants born from n-3-supplemented mothers compared with control infants. In red blood cells, DHA was 7.92% of total fatty acids compared with 5.86% (control infants). Plasma values showed a similar difference: 5.05% vs. 3.47% (controls). In n-3-supplemented infants, DHA concentrations were 35.2% higher than in control infants in red blood cells and 45.5% higher in plasma. These data indicate the importance of maternal dietary n-3 fatty acids and, in particular, maternal dietary DHA in promoting higher concentrations of DHA in the blood of the newborn infant.

  16. Adolescent alcohol exposure reduces behavioral flexibility, promotes disinhibition, and increases resistance to extinction of ethanol self-administration in adulthood.

    PubMed

    Gass, Justin T; Glen, William Bailey; McGonigal, Justin T; Trantham-Davidson, Heather; Lopez, Marcelo F; Randall, Patrick K; Yaxley, Richard; Floresco, Stan B; Chandler, L Judson

    2014-10-01

    The prefrontal cortex (PFC) is a brain region that is critically involved in cognitive function and inhibitory control of behavior, and adolescence represents an important period of continued PFC development that parallels the maturation of these functions. Evidence suggests that this period of continued development of the PFC may render it especially vulnerable to environmental insults that impact PFC function in adulthood. Experimentation with alcohol typically begins during adolescence when binge-like consumption of large quantities is common. In the present study, we investigated the effects of repeated cycles of adolescent intermittent ethanol (AIE) exposure (postnatal days 28-42) by vapor inhalation on different aspects of executive functioning in the adult rat. In an operant set-shifting task, AIE-exposed rats exhibited deficits in their ability to shift their response strategy when the rules of the task changed, indicating reduced behavioral flexibility. There were no differences in progressive ratio response for the reinforcer suggesting that AIE did not alter reinforcer motivation. Examination of performance on the elevated plus maze under conditions designed to minimize stress revealed that AIE exposure enhanced the number of entries into the open arms, which may reflect either reduced anxiety and/or disinhibition of exploratory-like behavior. In rats that trained to self-administer ethanol in an operant paradigm, AIE increased resistance to extinction of ethanol-seeking behavior. This resistance to extinction was reversed by positive allosteric modulation of mGluR5 during extinction training, an effect that is thought to reflect promotion of extinction learning mechanisms within the medial PFC. Consistent with this, CDPPB was also observed to reverse the deficits in behavioral flexibility. Finally, diffusion tensor imaging with multivariate analysis of 32 brain areas revealed that while there were no differences in the total brain volume, the volume of

  17. Activation of muscarinic cholinoceptor ameliorates tumor necrosis factor-α-induced barrier dysfunction in intestinal epithelial cells.

    PubMed

    Khan, Md Rafiqul Islam; Uwada, Junsuke; Yazawa, Takashi; Islam, Md Tariqul; Krug, Susanne M; Fromm, Michael; Karaki, Shin-ichiro; Suzuki, Yuichi; Kuwahara, Atsukazu; Yoshiki, Hatsumi; Sada, Kiyonao; Muramatsu, Ikunobu; Anisuzzaman, Abu Syed Md; Taniguchi, Takanobu

    2015-11-30

    Impaired intestinal barrier function is one of the critical issues in inflammatory bowel diseases. The aim of this study is to investigate muscarinic cholinoceptor (mAChR)-mediated signaling for the amelioration of cytokine-induced barrier dysfunction in intestinal epithelium. Rat colon challenged with TNF-α and interferon γ reduced transepithelial electrical resistance (TER). This barrier injury was attenuated by muscarinic stimulation. In HT-29/B6 intestinal epithelial cells, muscarinic stimulation suppressed TNF-α-induced activation of NF-κB signaling and barrier disruption. Finally, muscarinic stimulation promoted the shedding of TNFR1, which would be a mechanism for the attenuation of TNF-α/NF-κB signaling and barrier disruption via mAChR.

  18. Differentiation of muscarinic cholinergic receptor subtypes in human cortex and pons - Implications for anti-motion sickness therapy

    NASA Technical Reports Server (NTRS)

    Mccarthy, Bruce G.; Peroutka, Stephen J.

    1988-01-01

    Radioligand binding studies were used to analyze muscarinic cholinergic receptor subtypes in human cortex and pons. Muscarinic cholinergic receptors were labeled by H-3-quinuclidinyl benzilate (H-3-QNB). Scopolamine was equipotent in both brain regions and did not discriminate subtypes of H-3-QNB binding. By contrast, the M1 selective antagonist pirenzepine was approximately 33-fold more potent in human cortex than pons. Carbachol, a putative M2 selective agonist, was more than 100-fold more potent in human pons than cortex. These results demonstrate that the human pons contains a relatively large proportion of carbachol-sensitive muscarinic cholinergic receptors. Drugs targeted to this subpopulation of muscarinic cholinergic receptors may prove to be effective anti-motion sickness agents with less side effects than scopolamine.

  19. Superovulation Using the Combined Administration of Inhibin Antiserum and Equine Chorionic Gonadotropin Increases the Number of Ovulated Oocytes in C57BL/6 Female Mice

    PubMed Central

    Takeo, Toru; Nakagata, Naomi

    2015-01-01

    Superovulation is a reproductive technique generally used to produce genetically engineered mice. Superovulation in mice involves the administration of equine chorionic gonadotropin (eCG) to promote follicle growth and then that of human chorionic gonadotropin (hCG) to induce ovulation. Previously, some published studies reported that inhibin antiserum (IAS) increased the number of ovulated oocytes in ddY and wild-derived strains of mice. However, the effect of IAS on the C57BL/6 strain, which is the most widely used inbred strain for the production of genetically engineered mice, has not been investigated. In addition, the combined effect of IAS and eCG (IASe) on the number of ovulated oocytes in superovulation treatment has not been examined. In this study, we examined the effect of IAS and eCG on the number of ovulated oocytes in immature female mice of the C57BL/6 strain in superovulation treatment. Furthermore, we evaluated the quality of obtained oocytes produced by superovulation using IASe by in vitro fertilization (IVF) with sperm from C57BL/6 or genetically engineered mice. The developmental ability of fresh or cryopreserved embryos was examined by embryo transfer. The administration of IAS or eCG had a similar effect on the number of ovulated oocytes in C57BL/6 female mice. The number of ovulated oocytes increased to about 3-fold by the administration of IASe than by the administration of IAS or eCG alone. Oocytes derived from superovulation using IASe normally developed into 2-cell embryos by IVF using sperm from C57BL/6 mice. Fresh or cryopreserved 2-cell embryos produced by IVF between oocytes of C57BL/6 mice and sperm from genetically engineered mice normally developed into live pups following embryo transfer. In summary, a novel technique of superovulation using IASe is extremely useful for producing a great number of oocytes and offspring from genetically engineered mice. PMID:26024317

  20. Synthesis and biological evaluation of [125I]- and [123I]-4-iododexetimide, a potent muscarinic cholinergic receptor antagonist.

    PubMed

    Wilson, A A; Dannals, R F; Ravert, H T; Frost, J J; Wagner, H N

    1989-05-01

    A series of halogenated racemic analogues of dexetimide (1) was synthesized and their affinity for the muscarinic cholinergic receptor measured. One analogue, 4-iododexetimide (21), was efficiently labeled with 125I and 123I at high specific activity. In vitro binding studies and in vivo biodistribution studies suggest that 123I-labeled 21 may be useful for imaging muscarinic cholinergic receptors in the living human brain with single photon emission computed tomography. PMID:2785211

  1. The 5XFAD Mouse Model of Alzheimer's Disease Exhibits an Age-Dependent Increase in Anti-Ceramide IgG and Exogenous Administration of Ceramide Further Increases Anti-Ceramide Titers and Amyloid Plaque Burden.

    PubMed

    Dinkins, Michael B; Dasgupta, Somsankar; Wang, Guanghu; Zhu, Gu; He, Qian; Kong, Ji Na; Bieberich, Erhard

    2015-01-01

    We present evidence that 5XFAD Alzheimer's disease model mice develop an age-dependent increase in antibodies against ceramide, suggesting involvement of autoimmunity against ceramide in Alzheimer's disease pathology. To test this, we increased serum anti-ceramide IgG (2-fold) by ceramide administration and analyzed amyloid plaque formation in 5XFAD mice. There were no differences in soluble or total amyloid-β levels. However, females receiving ceramide had increased plaque burden (number, area, and size) compared to controls. Ceramide-treated mice showed an increase of serum exosomes (up to 3-fold using Alix as marker), suggesting that systemic anti-ceramide IgG and exosome levels are correlated with enhanced plaque formation. PMID:25720409

  2. Potentiation of M1 Muscarinic Receptor Reverses Plasticity Deficits and Negative and Cognitive Symptoms in a Schizophrenia Mouse Model.

    PubMed

    Ghoshal, A; Rook, J M; Dickerson, J W; Roop, G N; Morrison, R D; Jalan-Sakrikar, N; Lamsal, A; Noetzel, M J; Poslusney, M S; Wood, M R; Melancon, B J; Stauffer, S R; Xiang, Z; Daniels, J S; Niswender, C M; Jones, C K; Lindsley, C W; Conn, P J

    2016-01-01

    Schizophrenia patients exhibit deficits in signaling of the M1 subtype of muscarinic acetylcholine receptor (mAChR) in the prefrontal cortex (PFC) and also display impaired cortical long-term depression (LTD). We report that selective activation of the M1 mAChR subtype induces LTD in PFC and that this response is completely lost after repeated administration of phencyclidine (PCP), a mouse model of schizophrenia. Furthermore, discovery of a novel, systemically active M1 positive allosteric modulator (PAM), VU0453595, allowed us to evaluate the impact of selective potentiation of M1 on induction of LTD and behavioral deficits in PCP-treated mice. Interestingly, VU0453595 fully restored impaired LTD as well as deficits in cognitive function and social interaction in these mice. These results provide critical new insights into synaptic changes that may contribute to behavioral deficits in this mouse model and support a role for selective M1 PAMs as a novel approach for the treatment of schizophrenia.

  3. Antipsychotic-like effect of the muscarinic acetylcholine receptor agonist BuTAC in non-human primates.

    PubMed

    Andersen, Maibritt B; Croy, Carrie Hughes; Dencker, Ditte; Werge, Thomas; Bymaster, Frank P; Felder, Christian C; Fink-Jensen, Anders

    2015-01-01

    Cholinergic, muscarinic receptor agonists exhibit functional dopamine antagonism and muscarinic receptors have been suggested as possible future targets for the treatment of schizophrenia and drug abuse. The muscarinic ligand (5R,6R)-6-(3-butylthio-1,2,5-thiadiazol-4-yl)-1-azabicyclo[3.2.1]octane (BuTAC) exhibits high affinity for muscarinic receptors with no or substantially less affinity for a large number of other receptors and binding sites, including the dopamine receptors and the dopamine transporter. In the present study, we wanted to examine the possible antipsychotic-like effects of BuTAC in primates. To this end, we investigated the effects of BuTAC on d-amphetamine-induced behaviour in antipsychotic-naive Cebus paella monkeys. Possible adverse events of BuTAC, were evaluated in the same monkeys as well as in monkeys sensitized to antipsychotic-induced extrapyramidal side effects. The present data suggests that, the muscarinic receptor ligand BuTAC exhibits antipsychotic-like behaviour in primates. The behavioural data of BuTAC as well as the new biochemical data further substantiate the rationale for the use of muscarinic M1/M2/M4-preferring receptor agonists as novel pharmacological tools in the treatment of schizophrenia. PMID:25880220

  4. Antipsychotic-like effect of the muscarinic acetylcholine receptor agonist BuTAC in non-human primates.

    PubMed

    Andersen, Maibritt B; Croy, Carrie Hughes; Dencker, Ditte; Werge, Thomas; Bymaster, Frank P; Felder, Christian C; Fink-Jensen, Anders

    2015-01-01

    Cholinergic, muscarinic receptor agonists exhibit functional dopamine antagonism and muscarinic receptors have been suggested as possible future targets for the treatment of schizophrenia and drug abuse. The muscarinic ligand (5R,6R)-6-(3-butylthio-1,2,5-thiadiazol-4-yl)-1-azabicyclo[3.2.1]octane (BuTAC) exhibits high affinity for muscarinic receptors with no or substantially less affinity for a large number of other receptors and binding sites, including the dopamine receptors and the dopamine transporter. In the present study, we wanted to examine the possible antipsychotic-like effects of BuTAC in primates. To this end, we investigated the effects of BuTAC on d-amphetamine-induced behaviour in antipsychotic-naive Cebus paella monkeys. Possible adverse events of BuTAC, were evaluated in the same monkeys as well as in monkeys sensitized to antipsychotic-induced extrapyramidal side effects. The present data suggests that, the muscarinic receptor ligand BuTAC exhibits antipsychotic-like behaviour in primates. The behavioural data of BuTAC as well as the new biochemical data further substantiate the rationale for the use of muscarinic M1/M2/M4-preferring receptor agonists as novel pharmacological tools in the treatment of schizophrenia.

  5. Antipsychotic-Like Effect of the Muscarinic Acetylcholine Receptor Agonist BuTAC in Non-Human Primates

    PubMed Central

    Dencker, Ditte; Werge, Thomas; Bymaster, Frank P.; Felder, Christian C.; Fink-Jensen, Anders

    2015-01-01

    Cholinergic, muscarinic receptor agonists exhibit functional dopamine antagonism and muscarinic receptors have been suggested as possible future targets for the treatment of schizophrenia and drug abuse. The muscarinic ligand (5R,6R)-6-(3-butylthio-1,2,5-thiadiazol-4-yl)-1-azabicyclo[3.2.1]octane (BuTAC) exhibits high affinity for muscarinic receptors with no or substantially less affinity for a large number of other receptors and binding sites, including the dopamine receptors and the dopamine transporter. In the present study, we wanted to examine the possible antipsychotic-like effects of BuTAC in primates. To this end, we investigated the effects of BuTAC on d-amphetamine-induced behaviour in antipsychotic-naive Cebus paella monkeys. Possible adverse events of BuTAC, were evaluated in the same monkeys as well as in monkeys sensitized to antipsychotic-induced extrapyramidal side effects. The present data suggests that, the muscarinic receptor ligand BuTAC exhibits antipsychotic-like behaviour in primates. The behavioural data of BuTAC as well as the new biochemical data further substantiate the rationale for the use of muscarinic M1/M2/M4-preferring receptor agonists as novel pharmacological tools in the treatment of schizophrenia. PMID:25880220

  6. Mitochondrial Ultrastructural Alterations and Declined M2 Receptor Density Were Involved in Cardiac Dysfunction in Rats after Long Term Treatment with Autoantibodies against M2 Muscarinic Receptor

    PubMed Central

    Wang, Jin; Wang, Li; Wu, Ye; Wang, Jie; Lv, Tingting; Liu, Huirong

    2015-01-01

    Background Previous studies showed that autoantibodies (M2-AA) against the second extracellular loop of M2 muscarinic receptor (M2AChR-el2) from dilated cardiomyopathy (DCM) serum could induce DCM-like morphological changes in mice hearts. However, the effects of M2-AA on the cardiac function during the process of DCM and the potential mechanisms are not fully known. The present study was designed to dynamically observe the cardiac function, mitochondrial changes, and M2 receptor binding characteristics in rats long-term stimulated with M2-AA in vivo. Methods M2-AA-positive model was established by actively immunizing healthy male Wistar rats with synthetic M2AChR-el2 peptide for 18 months. Meanwhile, vehicle group rats were administrated with physiological saline. The change of mitochondrial membrane potential (ΔΨm) was detected by radionuclide imaging. The ultrastructure of mitochondria was observed under electron microscopy. The M2 receptor binding characteristics were determined by radioactive ligand binding assay. Results After immunization for 12 months, compared with vehicle group, M2AChR-el2-immunized rats showed decreased myocardial contractility and cardiac diastolic function evidenced by declined maximal rate of rise of ventricular pressure and increased left ventricular end-diastolic pressure, respectively. Additionally, mitochondrial swelling and vacuolation were observed. At 18 months, M2AChR-el2-immunized rats manifested significant decreased cardiac systolic and diastolic function and pathological changes such as enlargement of right ventricular cavity and wall thinning; and the mitochondrial damage was aggravated. Furthermore, the M2 receptor maximum binding capacity (Bmax) of the M2AChR-el2-immunized rats significantly decreased, while the M2 receptor dissociation constant (Kd) was increased. Conclusions Our study suggested that long-term stimulation with M2-AA leaded to the ventricular dilatation and gradual deterioration of cardiac dysfunction

  7. Repeated administration of aripiprazole produces a sensitization effect in the suppression of avoidance responding and phencyclidine-induced hyperlocomotion and increases D2 receptor-mediated behavioral function.

    PubMed

    Gao, Jun; Qin, Rongyin; Li, Ming

    2015-04-01

    The present study investigated how repeated administration of aripiprazole (a novel antipsychotic drug) alters its behavioral effects in two behavioral tests of antipsychotic activity and whether this alteration is correlated with an increase in dopamine D2 receptor function. Male adult Sprague-Dawley rats were first repeatedly tested with aripiprazole (3, 10 and 30 mg/kg, subcutaneously (sc)) or vehicle in a conditioned avoidance response (CAR) test or a phencyclidine (PCP) (3.20 mg/kg, sc)-induced hyperlocomotion test daily for five consecutive days. After 2-3 days of drug-free retraining or resting, all rats were then challenged with aripiprazole (1.5 or 3.0 mg/kg, sc). Repeated administration of aripiprazole progressively increased its inhibition of avoidance responding and PCP-induced hyperlocomotion. More importantly, rats previously treated with aripiprazole showed significantly lower avoidance response and lower PCP-induced hyperlocomotion than those previously treated with vehicle in the challenge tests. An increased sensitivity to quinpirole (a selective D2/3 agonist) in prior aripiprazole-treated rats was also found in the quinpirole-induced hyperlocomotion test, suggesting an enhanced D2/3-mediated function. These findings suggest that aripiprazole, despite its distinct receptor mechanisms of action, induces a sensitization effect similar to those induced by other antipsychotic drugs and this effect may be partially mediated by brain plasticity involving D2/3 receptor systems.

  8. Diet, physical exercise and Orlistat administration increase serum anti-Müllerian hormone (AMH) levels in women with polycystic ovary syndrome (PCOS).

    PubMed

    Vosnakis, Christos; Georgopoulos, Neoklis A; Rousso, David; Mavromatidis, Georgios; Katsikis, Ilias; Roupas, Nikolaos D; Mamali, Irene; Panidis, Dimitrios

    2013-03-01

    The present study investigates the combined effect of diet, physical exercise and Orlistat for 24 weeks, on serum anti-Müllerian hormone (AMH) levels in overweight and obese women with polycystic ovary syndrome (PCOS) and in overweight and obese controls. Sixty-one (61) selected women with PCOS and 20 overweight and obese controls followed an energy-restricted diet, physical exercise plus Orlistat administration (120 mg, 3 times per day) for 24 weeks. At baseline, week 12 and week 24, serum levels of AMH, FSH, LH, PRL, androgens, sex hormone-binding globulin (SHBG), glucose, and insulin were measured and Free Androgen Index (FAI) and Insulin Resistance (IR) indices were calculated. In PCOS women, serum AMH levels increased after 12 and 24 weeks of treatment. After 12 weeks LH and SHBG were increased, while Testosterone decreased. After 12 and 24 weeks, FAI was decreased and all indices of IR were significantly improved. We concluded that in overweight and obese women with PCOS Orlistat administration, combined with diet and physical exercise, for 24 weeks, resulted in significant weight loss, improvement of hyperandrogenism and insulin sensitivity, and increased serum AMH levels.

  9. Pharmacokinetics of repeated sodium salicylate administration to laying hens: evidence for time dependent increase in drug elimination from plasma and eggs.

    PubMed

    Poźniak, Błażej; Grabowski, Tomasz; Motykiewicz-Pers, Karolina; Bobrek, Kamila; Rak, Lech; Bobusia, Katarzyna; Gaweł, Andrzej; Świtała, Marcin

    2015-01-01

    Salicylates were the first non-steroid anti-inflammatory drugs (NSAIDs) to be used in any species and are still widely used in humans and livestock. However, the data on their pharmacokinetics in animals is limited, especially after repeated administration. Evidence exist that in chickens (Gallus gallus) salicylate (SA) may induce its own elimination. The aim of this study was to investigate salicylate pharmacokinetics and egg residues during repeated administration of sodium salicylate (SS) to laying hens. Pharmacokinetics of SA was assessed during 14 d oral administration of SS at daily doses of 50 mg/kg and 200 mg/kg body weight to laying hens. On the 1st, 7th and 14th d a 24 h-long pharmacokinetic study was carried out, whereas eggs were collected daily. Salicylate concentrations in plasma and eggs were determined using high-performance liquid chromatography with ultraviolet detection and pharmacokinetic variables were calculated using a non-compartmental model. Mean residence time (MRT), minimal plasma concentration (Cmin, C16h) and elimination half-life (T1/2el) of SA showed gradual decrease in layers administered with a lower dose. Total body clearance (ClB) increased. Layers administered with the higher dose showed a decrease only in the T1/2el. In the low dose group, SA was found only in the egg white and was low throughout the experiment. Egg whites from the higher dose group showed initially high SA levels which significantly decreased during the experiment. Yolk SA levels were lower and showed longer periods of accumulation and elimination. Repeated administration of SS induces SA elimination, although this effect may differ depending on the dose and production type of a chicken. Decreased plasma drug concentration may have clinical implications during prolonged SS treatment. PMID:25893240

  10. Increased sensitivity to cocaine self-administration in HIV-1 transgenic rats is associated with changes in striatal dopamine transporter binding

    PubMed Central

    McIntosh, Scot; Sexton, Tammy; Pattison, Lindsey P.; Childers, Steven R.; Hemby, Scott E.

    2015-01-01

    Cocaine abuse in HIV patients accelerates the progression and severity of neuropathology, motor impairment and cognitive dysfunction compared to non-drug using HIV patients. Cocaine and HIV interact with the dopamine transporter (DAT); however, the effect of their interaction on DAT binding remains understudied. The present study compared the dose-response functions for intravenous self-administration of cocaine and heroin between male HIV-1 transgenic (HIV-1 Tg) and Fischer 344 rats. The cocaine and heroin dose-response functions exhibit an inverted U-shape for both HIV-1 Tg and F344 rats. For cocaine, the number of infusions for each dose on the ascending limb was greater for HIV-1 Tg versus F344 rats. No significant changes in the heroin dose-response function were observed in HIV-1 Tg animals. Following the conclusion of self-administration experiments, DAT binding was assessed in striatal membranes. Saturation binding of the cocaine analog [125I] 3β-(4-iodophenyl)tropan-2β-carboxylic acid methyl ester ([125I]RTI-55) in rat striatal membranes resulted in binding curves that were best fit to a two-site binding model, allowing for calculation of dissociation constant (Kd) and binding density (Bmax) values that correspond to high- and low-affinity DAT binding sites. Control HIV-1 Tg rats exhibited a significantly greater affinity (i.e., decrease in Kd value) in the low-affinity DAT binding site compared to control F344 rats. Furthermore, cocaine self-administration in HIV-1 Tg rats increased low-affinity Kd (i.e., decreased affinity) compared to levels observed in control F344 rats. Cocaine also increased low-affinity Bmax in HIV-1 Tg rats as compared to controls, indicating an increase in the number of low-affinity DAT binding sites. F344 rats did not exhibit any change in high- or low-affinity Kd or Bmax values following cocaine or heroin self-administration. The increase in DAT affinity in cocaine HIV-1 Tg rats is consistent with the leftward shift of the

  11. Increased Sensitivity to Cocaine Self-Administration in HIV-1 Transgenic Rats is Associated with Changes in Striatal Dopamine Transporter Binding.

    PubMed

    McIntosh, Scot; Sexton, Tammy; Pattison, Lindsey P; Childers, Steven R; Hemby, Scott E

    2015-09-01

    Cocaine abuse in HIV patients accelerates the progression and severity of neuropathology, motor impairment and cognitive dysfunction compared to non-drug using HIV patients. Cocaine and HIV interact with the dopamine transporter (DAT); however, the effect of their interaction on DAT binding remains understudied. The present study compared the dose-response functions for intravenous self-administration of cocaine and heroin between male HIV-1 transgenic (HIV-1 Tg) and Fischer 344 rats. The cocaine and heroin dose-response functions exhibit an inverted U-shape for both HIV-1 Tg and F344 rats. For cocaine, the number of infusions for each dose on the ascending limb was greater for HIV-1 Tg versus F344 rats. No significant changes in the heroin dose-response function were observed in HIV-1 Tg animals. Following the conclusion of self-administration experiments, DAT binding was assessed in striatal membranes. Saturation binding of the cocaine analog [(125)I] 3β-(4-iodophenyl)tropan-2β-carboxylic acid methyl ester ([(125)I]RTI-55) in rat striatal membranes resulted in binding curves that were best fit to a two-site binding model, allowing for calculation of dissociation constant (Kd) and binding density (Bmax) values that correspond to high- and low-affinity DAT binding sites. Control HIV-1 Tg rats exhibited a significantly greater affinity (i.e., decrease in Kd value) in the low-affinity DAT binding site compared to control F344 rats. Furthermore, cocaine self-administration in HIV-1 Tg rats increased low-affinity Kd (i.e., decreased affinity) compared to levels observed in control F344 rats. Cocaine also increased low-affinity Bmax in HIV-1 Tg rats as compared to controls, indicating an increase in the number of low-affinity DAT binding sites. F344 rats did not exhibit any change in high- or low-affinity Kd or Bmax values following cocaine or heroin self-administration. The increase in DAT affinity in cocaine HIV-1 Tg rats is consistent with the leftward shift of the

  12. The relation between dopamine oxidation currents in the nucleus accumbens and conditioned increases in motor activity in rats following repeated administration of d-amphetamine or cocaine.

    PubMed

    Di Ciano, P; Blaha, C D; Phillips, A G

    1998-03-01

    Chronoamperometric recording techniques were used to monitor extracellular dopamine efflux in the nucleus accumbens associated with unconditioned and conditioned increases in motor activity in rats, following the intravenous administration of either d-amphetamine (0.63 mg/kg) or cocaine (3 mg/kg), or the presentation of a conditioned stimulus paired repeatedly with one of these psychostimulants. Each drug was administered daily for 7 days, either in the home cage or an environment in which a compound stimulus (light offset, odour) was presented. Rats in control groups received saline instead of drug in the distinctive test environment. On day 7 of training, significant increases in unconditioned motor activity were observed in the 30 min session following infusions of either d-amphetamine or cocaine. Associated dopamine oxidation currents in the nucleus accumbens increased immediately following administration of either drug and remained significantly elevated above baseline during the entire 30 min recording period. On the test day, presentation of the conditioned stimulus with vehicle infusions, in the distinct environment, was accompanied by an increase in dopamine oxidation currents and a conditioned increase in motor activity, only in the groups in which these stimuli had been paired with d-amphetamine or cocaine. Neither the magnitude or duration of the conditioned motor activity matched the corresponding change in extracellular dopamine efflux in the nucleus accumbens. Accordingly, it is argued that the increase in dopamine concentration serves as a neurochemical correlate of the unconditioned and conditioned stimuli. The change in motor activity constitutes the unconditioned and conditioned responses that are subserved by the neural systems activated by the initial rise in extracellular dopamine. PMID:9753179

  13. Affinities of brompheniramine, chlorpheniramine, and terfenadine at the five human muscarinic cholinergic receptor subtypes.

    PubMed

    Yasuda, S U; Yasuda, R P

    1999-04-01

    Anticholinergic effects are presumed to be the mechanism for the efficacy of chlorpheniramine in symptomatic relief of the common cold. Terfenadine, a second-generation antihistamine, reportedly lacks anticholinergic side effects. We evaluated affinities of two commonly used over-the-counter antihistamines, brompheniramine and chlorpheniramine, as well as terfenadine in comparison with atropine at the five human muscarinic cholinergic receptor subtypes using CHO cells stably transfected with the individual subtypes. Atropine was more potent than all three drugs at m1-m5 (p<0.01). No significant difference was observed between chlorpheniramine and brompheniramine. Atropine, brompheniramine, and chlorpheniramine could not discriminate between m1-m5. Terfenadine demonstrated subtype selectivity at m3. In vitro comparisons in human muscarinic receptor subtypes could potentially be used to predict clinical anticholinergic effects of antihistamines and to target receptor-specific effects of such agents.

  14. Conformational and stereoeletronic investigations of muscarinic agonists of acetylcholine by NMR and theoretical calculations

    NASA Astrophysics Data System (ADS)

    da Silva, Julio Cesar A.; Ducati, Lucas C.; Rittner, Roberto

    2012-05-01

    NMR solvent effects and theoretical calculations showed muscarinic agonists present a large stability for their near synclinal conformations, indicating the presence of significant stabilization factors. Analysis of the results clearly indicated that this stability is not determined by the dihedral around the substituted C-C ethane bond, as stated by some authors, but a consequence of the geometry adopted in order to maximize N+/O interactions in this type of molecules. It can be assumed that acetylcholine and its muscarinic agonists exhibit their biologic activity when the positively charged nitrogen and the oxygen atoms are in the same side of the molecule within an interatomic distance ranging from 3.0 to 6.0 Å.

  15. Parallel maturation of the pancreatic secretory response to cholinergic stimulation and the muscarinic receptor population.

    PubMed Central

    Dumont, Y.; Larose, L.; Morisset, J.; Poirier, G. G.

    1981-01-01

    1 The appearance of pancreatic muscarinic receptors during development has been measured by use of the specific ligand [3H]-quinuclidinyl benzilate ([3H]-QNB). 2 QNB binding sites are present in foetal pancreas; their maximal concentration is attained at the age of 30 days and a significant decrease is observed in one year old animals. 3 Affinity of [3H]-QNB for the muscarinic receptor does not change with age. 4 An evaluation of the pancreatic secretory response to a cholinoceptor agonist as a function of age indicates that the development of this response parallels that of the receptor population. 5 It is suggested that, at all ages from 3 days after birth onwards, the maximal secretory response of the exocrine pancreas to a cholinoceptor agonist mobilizes the same proportion of the total population of QNB binding sites. PMID:6165420

  16. Solubilization and characterization of guanine nucleotide-sensitive muscarinic agonist binding sites from rat myocardium.

    PubMed Central

    Berrie, C. P.; Birdsall, N. J.; Hulme, E. C.; Keen, M.; Stockton, J. M.

    1984-01-01

    Muscarinic receptors from rat myocardial membranes may be solubilized by digitonin in good yield at low temperatures in the presence of Mg2+. Under these conditions, up to 60% of the soluble receptors show high affinity binding for the potent agonist [3H]-oxotremorine-M (KA = 10(9)M-1), which is inhibited by 5'-guanylylimidodiphosphate. The muscarinic binding site labelled with [3H]-oxotremorine-M has a higher sedimentation coefficient (13.4 s) than sites labelled with a 3H antagonist in the presence of guanylylimidodiphosphate (11.6 s) and probably represents a complex between the ligand binding subunit of the receptor and a guanine nucleotide binding protein. PMID:6478115

  17. Monoclonal antibodies against the native or denatured forms of muscarinic acetylcholine receptors.

    PubMed Central

    André, C; Guillet, J G; De Backer, J P; Vanderheyden, P; Hoebeke, J; Strosberg, A D

    1984-01-01

    BALB/c mice were immunized with affinity-purified muscarinic acetylcholine receptors from calf brain and their splenocytes fused with NS1 myeloma cells. Hybrid cultures were grown and selected for production of antibodies on the basis of enzyme immunoassays on calf and rat forebrain membrane preparations. Thirty-four clones were retained and six of them further subcloned. Two of these subclones produced antibodies that selectively recognized muscarinic acetylcholine receptor-bearing membranes. The M-35b antibodies interacted only with native digitonin-solubilized receptors, and not with denatured receptors. The M-23c antibodies did not react with active digitonin-solubilized receptors but recognized the denatured form. The M-23c antibodies should thus be useful in the purification of the receptor and its precursor translation products, while the M-35b antibodies could be used for the immunocytochemical localization of the receptor in cells and tissues of different species. Images Fig. 2. Fig. 3. PMID:6200320

  18. Muscarinic M(3) facilitation of acetylcholine release from rat myenteric neurons depends on adenosine outflow leading to activation of excitatory A(2A) receptors.

    PubMed

    Vieira, C; Duarte-Araújo, M; Adães, S; Magalhães-Cardoso, T; Correia-de-Sá, P

    2009-10-01

    Acetylcholine (ACh) is a major excitatory neurotransmitter in the myenteric plexus, and it regulates its own release acting via muscarinic autoreceptors. Adenosine released from stimulated myenteric neurons modulates ACh release preferentially via facilitatory A(2A) receptors. In this study, we investigated how muscarinic and adenosine receptors interplay to regulate ACh from the longitudinal muscle-myenteric plexus of the rat ileum. Blockade of the muscarinic M(2) receptor with 11-[[2-1[(diethylamino) methyl-1-piperidinyl]- acetyl

  19. Effects of atropine treatment on in vitro and in vivo binding of 4-[125I]-dexetimide to central and myocardial muscarinic receptors.

    PubMed

    Uno, Y; Matsumura, K; Scheffel, U; Wilson, A A; Dannals, R F; Wagner, H N

    1991-01-01

    Upregulation of muscarinic cholinergic receptors (mAChR) after chronic atropine treatment has been described previously. The present study was designed to evaluate 4-iodine-125 dexetimide as an agent to determine changes in the number of mAChR. Rats were injected subcutaneously with atropine (500 mg/kg) either once or chronically, once daily for 10 days, and sacrificed 24 h later. In vitro binding assays with 4-[125I]-dexetimide showed significant increases in the number of mAChR in cerebra (21%) and ventricles (45%) after chronic atropine treatment but not after acute treatment. The affinity of binding to cerebral and ventricular mAChR declined after acute and chronic atropine treatment. In vivo studies were carried out involving intravenous injection of 4-[125I]-dexetimide 24 h after atropine treatment. Binding was markedly reduced in the brain and heart. Upregulation of mAChR, as seen in in vitro studies, could not be observed because of the remaining atropine. Occupancy of mAChR by atropine persisted as long as 7 days after one dose. The results of these studies indicate that 4-[125I]-dexetimide binding reflects the effects of atropine on central and peripheral muscarinic cholinergic receptors in vitro and in vivo. PMID:1915471

  20. Chronic administration of nicotine-free cigarette smoke extract impaired endothelium-dependent vascular relaxation in rats via increased vascular oxidative stress.

    PubMed

    Shimosato, Takashi; Geddawy, Ayman; Tawa, Masashi; Imamura, Takeshi; Okamura, Tomio

    2012-01-01

    Cigarette smoking has been implicated in the initiation and progression of cardiovascular disorders and atherosclerosis. Here, we examined the effects of nicotine-free cigarette smoke extract (CSE) on the regulation of cardiovascular function. Rats were subcutaneously administered PBS or nicotine-free CSE at 0.05 to 1.5 mL/day per rat for 4 weeks. Blood pressure, cardiac function, and vascular responsiveness were measured at 4 weeks after administration. Furthermore, acute effects of nicotine-free CSE were also studied in the aorta isolated from normal rats. Blood pressure and left ventricular systolic pressure (LVSP) were significantly increased in the nicotine-free CSE-administered rats, but heart rate, dP/dt(max), and dP/dt(min) were not affected. Endothelium-dependent relaxation by acetylcholine (ACh) in the nicotine-free CSE-treated rats was significantly attenuated compared to PBS-treated rats, but endothelium-independent relaxation by sodium nitroprusside (SNP) did not differ. Pretreatment with superoxide dismutase restored the attenuated ACh-induced relaxation. Contractions by phenylephrine, angiotensin II, and KCl did not differ between two groups. In vitro acute nicotine-free CSE treatment did not alter the response to ACh or SNP. These results suggest that chronic nicotine-free CSE administration impairs endothelial function by increased production of superoxide derived from the vascular wall components other than smooth muscles and induces slight hypertension accompanied with LVSP elevation.

  1. Memory improving actions of gabapentin in mice: possible involvement of central muscarinic cholinergic mechanism.

    PubMed

    Boccia, M M; Acosta, G B; Baratti, C M

    2001-10-01

    Male CF-1 mice were tested 48 h after training on a one trial step-through inhibitory avoidance task. Immediately post-training, intraperitoneal (i.p.) injections of the antiepileptic gabapentin (1-(aminomethyl) cyclohexaneacetic acid) (GBP, 10 mg/kg) enhanced retention performance. The effect was prevented by atropine, a central muscarinic cholinergic receptor antagonist (0.5 mg/kg, i.p.) administered after training but 10 min prior to GBP treatment. In contrast, neither methylatropine (0.5 mg/kg, i.p.), a peripherally acting muscarinic receptor blocker, nor mecamylamine (5 mg/kg, i.p.) or hexamethonium (5 mg/kg, i.p.), two cholinergic nicotinic receptor antagonists, prevented the effects of post-training GBP on retention performance. Low subeffective doses of the central acting anticholinesterase physostigmine (35 mg/kg, i.p.) administered immediately after training, and GBP (5 mg/kg, i.p.), given 10 min after training, significantly enhanced retention performance. The effects of GBP (5 mg/kg, i.p.) were not influenced by the peripherally acting anticholinesterase neostigmine (150 mg/kg, i.p.). Considered together, these findings suggest a disinhibitory action of GBP on the activity of central muscarinic cholinergic mechanisms that are involved in memory consolidation. PMID:11578817

  2. Quantitative ARG microimaging studies of two muscarinic antagonist isomers: Blocking and the effects of cocaine

    SciTech Connect

    Som, P.; Wang, G.J.; Oster, Z.H.

    1994-05-01

    The distribution of the racemic mixture of IQNP(1-Azabicyclo [2-2-2] oct-3-yl alpha-hydroxy-alpha-(1-iodo-propen-3-yl)-alpha-phenylacetate), a muscarinic antagonist was described earlier. Recently, the radioiodinated Z and E-(R,R) IQNP isomers have been prepared. Quantitative ARG studies using the Z and E isomers were performed in control rats and after pretreatment with ({plus_minus}) QNB or cocaine. High uptake of (Z)-IQNP was seen in the heart and brain with GI and urinary excretion. Lung uptake was lower than with the racemic IQNP. (Z)-IQNP uptake was maximal at 15 min p.i. with homogeneous distribution in the heart. In the brain, highest uptake was in the caudate, cortex, hippocampus, pons and thalamus. (Z)-IQNP showed higher cerebellar uptake and lower cortical uptake compared to (E)-IQNP. Clearance from brain was slower than bean. Heart and brain uptake of (E)-IQNP were markedly lower than the Z isomer. After QNB pretreatment, almost complete blocking of (Z)-IQNP uptake in heart and brain occurred. Cocaine did not significantly affect the distribution of IQNP. These data indicate that (Z)-IQNP has high affinity for the M2 muscarinic receptor with potential for brain and heart imaging. Cocaine appears to have little effect on the muscarinic-cholinergic receptors in the brain and heart.

  3. Carbamylcholine and phorbol esters desensitize muscarinic receptors by different mechanisms in rat pancreatic acini.

    PubMed

    Blanchard, L M; Paquette, B; Larose, L; Morisset, J

    1990-01-01

    Pretreatment of rat pancreatic acini with phorbol 12-myristate, 13-acetate (PMA), a protein kinase C (PK-C) activator, caused the desensitization of carbamylcholine (CBC)-induced amylase release in a concentration- and time-dependent fashion. The less potent phorbol-12, 13-dibutyrate (PDBu) also provoked a desensitization, but the inactive 4-alpha-phorbol-12,13-didecanoate had no effect. PMA or PDBu also significantly reduced subsequent amylase release induced by caerulein or secretin in contrast to CBC, which only reduced amylase release induced by CBC or secretin. Preincubation of acini with PMA did not lead to a decrease in PMA or A23187-stimulated amylase release. A 3 h resting period did not restore the desensitization induced by PMA or PDBu. Pretreatment with PMA did not cause changes in muscarinic receptor high- and low-affinity populations as observed with CBC pretreatment. The PK-C inhibitor H-7 completely prevented the desensitization induced by PDBu but not that induced by CBC. TMB-8, another PK-C inhibitor, also completely prevented the desensitization induced by PDBu but only partially that induced by CBC. These results suggest that phorbol esters can induce desensitization of muscarinic receptor-stimulated amylase release by a different mechanism than that involved in muscarinic agonist-induced desensitization.

  4. Motor activity-induced dopamine release in the substantia nigra is regulated by muscarinic receptors.

    PubMed

    Andersson, Daniel R; Björnsson, Evelina; Bergquist, Filip; Nissbrandt, Hans

    2010-01-01

    Nigro-striatal neurons release dopamine not only from their axon terminals in the striatum, but also from somata and dendrites in the substantia nigra. Somatodendritic dopamine release in the substantia nigra can facilitate motor function by mechanisms that may act independently of axon terminal dopamine release in the striatum. The dopamine neurons in the substantia nigra receive a cholinergic input from the pedunculopontine nucleus. Despite recent efforts to introduce this nucleus as a potential target for deep brain stimulation to treat motor symptoms in Parkinson's disease; and the well-known antiparkinsonian effects of anticholinergic drugs; the cholinergic influence on somatodendritic dopamine release is not well understood. The aim of this study was to investigate the possible regulation of locomotor-induced dopamine release in the substantia nigra by endogenous acetylcholine release. In intact and 6-OHDA hemi-lesioned animals alike, the muscarinic antagonist scopolamine, when perfused in the substantia nigra, amplified the locomotor-induced somatodendritic dopamine release to approximately 200% of baseline, compared to 120-130% of baseline in vehicle-treated animals. A functional importance of nigral muscarinic receptor activation was demonstrated in hemi-lesioned animals, where motor performance was significantly improved by scopolamine to 82% of pre-lesion performance, as compared to 56% in vehicle-treated controls. The results indicate that muscarinic activity in the substantia nigra is of functional importance in an animal Parkinson's disease model, and strengthen the notion that nigral dopaminergic regulation of motor activity/performance is independent of striatal dopamine release.

  5. Verrucotoxin inhibits KATP channels in cardiac myocytes through a muscarinic M3 receptor-PKC pathway.

    PubMed

    Wang, Jian-Wu; Yazawa, Kazuto; Hao, Li-Ying; Onoue, Yoshio; Kameyama, Masaki

    2007-06-01

    Verrucotoxin is the major component of venom from the stonefish (Synanceia verrucosa). Stings from the dorsal spines of the stonefish produce intensive pain, convulsions, hypotension, paralysis, respiratory weakness and collapse of the cardiovascular system, occasionally leading to death. It has been reported that verrucotoxin might modulate ATP-sensitive K+ (KATP) current in frog atrial fibers. However, the mechanism by which verrucotoxin acts on KATP current remains unclear. In this study, we examined whether verrucotoxin inhibited KATP current in guinea pig ventricular myocytes, using the patch clamp method. Verrucotoxin suppressed KATP current induced by pinacidil (KATP channel opener) in a concentration-dependent manner, with a half maximum concentration of 16.3 microg/ml. The effect of verrucotoxin on KATP current was suppressed by atropine (1 microM), a muscarinic receptor antagonist, or by 4-diphenylacetoxy-N-methylpiperidine (100 nM), a muscarinic M3 receptor antagonist. Furthermore, the effect of verrucotoxin on KATP current was attenuated by the protein kinase C (PKC) inhibitor chelerythrine (10 microM) and calphostin C (10 microM), yet not by the cAMP-dependent protein kinase (PKA) inhibitor H-89 (0.5 microM). These results suggest that verrucotoxin inhibits KATP current through the muscarinic M3 receptor-PKC pathway. These findings enhance our understanding of the toxic effects of verrucotoxin from the stonefish. PMID:17362922

  6. Expression of muscarinic acetylcholine and dopamine receptor mRNAs in rat basal ganglia

    SciTech Connect

    Weiner, D.M. Howard Hughes Medical Inst., Bethesda, MD ); Levey, A.I. Johns Hopkins Univ., Baltimore, MD ); Brann, M.R. )

    1990-09-01

    Within the basal ganglia, acetylcholine and dopamine play a central role in the extrapyramidal control of motor function. The physiologic effects of these neurotransmitters are mediated by a diversity of receptor subtypes, several of which have now been cloned. Muscarinic acetylcholine receptors are encoded by five genes (m1-m5), and of the two known dopamine receptor subtypes (D1 and D2) the D2 receptor gene has been characterized. To gain insight into the physiological roles of each of these receptor subtypes, the authors prepared oligodeoxynucleotide probes to localize receptor subtype mRNAs within the rat striatum and substantia nigra by in situ hybridization histochemistry. Within the striatum, three muscarinic (m1, m2, m4) receptor mRNAs and the D2 receptor mRNA were detected. The m1 mRNA was expressed in most neurons; the m2 mRNA, in neurons which were both very large and rare; and the m4 and D2 mRNAs, in 40-50% of the neurons, one-third of which express both mRNAs. Within the substantia nigra, pars compacta, only the m5 and D2 mRNAs were detected, and most neurons expressed both mRNAs. These data provide anatomical evidence for the identity of the receptor subtypes which mediate the diverse effects of muscarinic and dopaminergic drugs on basal ganglia function.

  7. Activation of muscarinic receptors inhibits glutamate-induced GSK-3β overactivation in PC12 cells

    PubMed Central

    Ma, Ke; Yang, Li-min; Chen, Hong-zhuan; Lu, Yang

    2013-01-01

    Aim: To investigate the actions of the muscarinic agonist carbachol on glutamate-induced neurotoxicity in PC12 cells, and the underlying mechanisms. Methods: PC12 cells were treated with different concentrations of glutamate for 24 or 48 h. The cell viability was measured using MTT assay, and the expression and activation of GSK-3β were detected with Western blot. β-Catenin translocation was detected using immunofluorescence. Luciferase reporter assay and real-time PCR were used to analyze the transcriptional activity of β-catenin. Results: Glutamate (1, 3, and 10 mmol/L) induced PC12 cell death in a dose-dependent manner. Moreover, treatment of the cells with glutamate (1 mmol/L) caused significant overactivation of GSK-3β and prevented β-catenin translocation to the nucleus. Pretreatment with carbachol (0.01 μmol/L) blocked glutamate-induced cell death and GSK-3β overactivation, and markedly enhanced β-catenin transcriptional activity. Conclusion: Activation of muscarinic receptors exerts neuroprotection in PC12 cells by attenuating glutamate-induced GSK-3β overactivation, suggesting potential benefits of muscarinic agonists for Alzheimer's disease. PMID:23685950

  8. ATP-dependent desensitization of the muscarinic K+ channel in rat atrial cells.

    PubMed Central

    Shui, Z; Boyett, M R; Zang, W J

    1997-01-01

    1. Fast desensitization of the muscarinic K+ channel has been studied in excised patches from rat atrial cells. 2. In inside-out patches, ACh was present in the pipette and GTP was applied via the bath to activate the channel. In outside-out patches, GTP was present in the pipette and ACh was applied via the bath to activate the channel. In both cases, during a 30 s exposure to GTP or ACh there was a decline in channel activity as a result of fast desensitization if ATP was present. 3. In inside-out patches, fast desensitization was still observed if the muscarinic ACh receptor was bypassed and the channel was activated by GTP gamma S. This suggests that fast desensitization is a result of a modification of the channel (or the connecting G protein) and not the receptor. 4. In both inside-out and outside-out patches, channel activity was depressed and fast desensitization was reduced or absent, if ATP was not present. 5. The non-hydrolysable analogue of ATP, AMP-PNP, did not substitute for ATP in its effects on the channel. 6. The results are consistent with the hypothesis that fast desensitization of the muscarinic K+ channel is the result of a dephosphorylation of the channel. Images Figure 12 PMID:9409473

  9. Long-acting muscarinic receptor antagonists for the treatment of chronic airway diseases

    PubMed Central

    Palot, Alain; Sofalvi, Tunde; Pahus, Laurie; Gouitaa, Marion; Tummino, Celine; Martinez, Stephanie; Charpin, Denis; Bourdin, Arnaud; Chanez, Pascal

    2014-01-01

    Acetylcholine (neuronal and non-neuronal origin) regulates bronchoconstriction, and mucus secretion. It has an inflammatory effect by inducing attraction, survival and cytokine release from inflammatory cells. Muscarinic receptors throughout the bronchial tree are mainly restricted to muscarinic M1, M2 and M3 receptors. Three long-acting muscarinic receptor antagonists (LAMAs) were approved for the treatment of chronic obstructive pulmonary disease (COPD) in Europe: once-daily tiotropium bromide; once-daily glycopyrronium bromide; and twice-daily aclidinium bromide. All have higher selectivity for M3 receptors than for M2 receptors, and dissociate more slowly from the M3 receptors than they do from the M2 receptors. Some LAMAs showed anti-inflammatory effects [inhibition of neutrophil chemotactic activity and migration of alveolar neutrophils, decrease of several cytokines in the bronchoalveolar lavage (BAL) including interleukin (IL)-6, tumor necrosis factor (TNF)-α and leukotriene (LT)B4] and antiremodeling effects (inhibition of mucus gland hypertrophy and decrease in MUC5AC-positive goblet cell number, decrease in MUC5AC overexpression). In the clinic, LAMAs showed a significant improvement of forced expiratory volume in 1 second (FEV1), quality of life, dyspnea and reduced the number of exacerbations in COPD and more recently in asthma. This review will focus on the three LAMAs approved in Europe in the treatment of chronic airway diseases. PMID:24587893

  10. Memory improving actions of gabapentin in mice: possible involvement of central muscarinic cholinergic mechanism.

    PubMed

    Boccia, M M; Acosta, G B; Baratti, C M

    2001-10-01

    Male CF-1 mice were tested 48 h after training on a one trial step-through inhibitory avoidance task. Immediately post-training, intraperitoneal (i.p.) injections of the antiepileptic gabapentin (1-(aminomethyl) cyclohexaneacetic acid) (GBP, 10 mg/kg) enhanced retention performance. The effect was prevented by atropine, a central muscarinic cholinergic receptor antagonist (0.5 mg/kg, i.p.) administered after training but 10 min prior to GBP treatment. In contrast, neither methylatropine (0.5 mg/kg, i.p.), a peripherally acting muscarinic receptor blocker, nor mecamylamine (5 mg/kg, i.p.) or hexamethonium (5 mg/kg, i.p.), two cholinergic nicotinic receptor antagonists, prevented the effects of post-training GBP on retention performance. Low subeffective doses of the central acting anticholinesterase physostigmine (35 mg/kg, i.p.) administered immediately after training, and GBP (5 mg/kg, i.p.), given 10 min after training, significantly enhanced retention performance. The effects of GBP (5 mg/kg, i.p.) were not influenced by the peripherally acting anticholinesterase neostigmine (150 mg/kg, i.p.). Considered together, these findings suggest a disinhibitory action of GBP on the activity of central muscarinic cholinergic mechanisms that are involved in memory consolidation.

  11. Acute Nicotine Administration Increases BOLD fMRI Signal in Brain Regions Involved in Reward Signaling and Compulsive Drug Intake in Rats

    PubMed Central

    Alexander, Jon C.; Perez, Pablo D.; Bauzo-Rodriguez, Rayna; Hall, Gabrielle; Klausner, Rachel; Guerra, Valerie; Zeng, Huadong; Igari, Moe; Febo, Marcelo

    2015-01-01

    Background: Acute nicotine administration potentiates brain reward function and enhances motor and cognitive function. These studies investigated which brain areas are being activated by a wide range of doses of nicotine, and if this is diminished by pretreatment with the nonselective nicotinic receptor antagonist mecamylamine. Methods: Drug-induced changes in brain activity were assessed by measuring changes in the blood oxygen level dependent (BOLD) signal using an 11.1-Tesla magnetic resonance scanner. In the first experiment, nicotine naïve rats were mildly anesthetized and the effect of nicotine (0.03–0.6mg/kg) on the BOLD signal was investigated for 10min. In the second experiment, the effect of mecamylamine on nicotine-induced brain activity was investigated. Results: A high dose of nicotine increased the BOLD signal in brain areas implicated in reward signaling, such as the nucleus accumbens shell and the prelimbic area. Nicotine also induced a dose-dependent increase in the BOLD signal in the striato-thalamo-orbitofrontal circuit, which plays a role in compulsive drug intake, and in the insular cortex, which contributes to nicotine craving and relapse. In addition, nicotine induced a large increase in the BOLD signal in motor and somatosensory cortices. Mecamylamine alone did not affect the BOLD signal in most brain areas, but induced a negative BOLD response in cortical areas, including insular, motor, and somatosensory cortices. Pretreatment with mecamylamine completely blocked the nicotine-induced increase in the BOLD signal. Conclusions: These studies demonstrate that acute nicotine administration activates brain areas that play a role in reward signaling, compulsive behavior, and motor and cognitive function. PMID:25552431

  12. Rhythmic patterns evoked in locust leg motor neurons by the muscarinic agonist pilocarpine.

    PubMed

    Ryckebusch, S; Laurent, G

    1993-05-01

    1. When an isolated metathoracic ganglion of the locust was superfused with the muscarinic cholinergic agonist pilocarpine, rhythmic activity was induced in leg motor neurons. The frequency of this induced rhythm increased approximately linearly from 0 to 0.2 Hz with concentrations of pilocarpine from 10(-5) to 10(-4) M. Rhythmic activity evoked by pilocarpine could be completely and reversibly blocked by 3 x 10(-5) M atropine, but was unaffected by 10(-4) M d-tubocurarine. 2. For each hemiganglion, the observed rhythm was characterized by two main phases: a levator phase, during which the anterior coxal rotator, levators of the trochanter, flexors of the tibia, and common inhibitory motor neurons were active; and a depressor phase, during which depressors of the trochanter, extensors of the tibia, and depressors of the tarsus were active. Activity in depressors of the trochanter followed the activity of the levators of the trochanter with a short, constant interburst latency. Activity in the levator of the tarsus spanned both phases. 3. The levator phase was short compared with the period (0.5-2 s, or 10-20% of the period) and did not depend on the period. The interval between the end of a levator burst and the beginning of the following one thus increased with cycle period. The depressor phase was more variable, and was usually shorter than the interval between successive levator bursts. 4. Motor neurons in a same pool often received common discrete synaptic potentials (e.g., levators of trochanter or extensors of tibia), suggesting common drive during the rhythm. Coactive motor neurons on opposite sides (such as left trochanteral depressors and right trochanteral levators), however, did not share obvious common postsynaptic potentials. Depolarization of a pool of motor neurons during its phase of activity was generally accompanied by hyperpolarization of its antagonist(s) on the same side. 5. Rhythmic activity was generally evoked in both hemiganglia of the

  13. Muscarinic excitation of parvalbumin-positive interneurons contributes to the severity of pilocarpine-induced seizures

    PubMed Central

    Yi, Feng; DeCan, Evan; Stoll, Kurt; Marceau, Eric; Deisseroth, Karl; Lawrence, J. Josh

    2014-01-01

    SUMMARY Objective A common rodent model in epilepsy research employs the muscarinic acetylcholine receptor (mAChR) agonist pilocarpine, yet the mechanisms underlying the induction of pilocarpine-induced seizures (PISs) remain unclear. Global M1 mAChR (M1R) knockout mice are resistant to PISs, implying that M1R activation disrupts excitation/inhibition balance. Parvalbumin-positive (PV) inhibitory neurons express M1 mAChRs, participate in cholinergically-induced oscillations, and can enter a state of depolarization block (DB) during epileptiform activity. Here, we test the hypothesis that pilocarpine activation of M1Rs expressed on PV cells contributes to PISs. Methods CA1 PV cells in PV-CRE mice were visualized with a floxed YFP or hM3Dq-mCherry adeno-associated virus, or by crossing PV-CRE mice with the RosaYFP reporter line. To eliminate M1Rs from PV cells, we generated PV-M1KO mice by crossing PV-CRE and floxed M1 mice. Action potential (AP) frequency was monitored during application of pilocarpine (200 µM). In behavioral experiments, locomotion and seizure symptoms were recorded in WT or PV-M1KO mice during PISs. Results Pilocarpine significantly increased AP frequency in CA1 PV cells into the gamma range. In the continued presence of pilocarpine, a subset (5/7) of PV cells progressed to DB, which was mimicked by hM3Dq activation of Gq-receptor signaling. Pilocarpine-induced depolarization, AP firing at gamma frequency, and progression to DB were prevented in CA1 PV cells of PV-M1KO mice. Finally, compared to WT mice, PV-M1KO mice were associated with reduced severity of PISs. Significance Pilocarpine can directly depolarize PV+ cells via M1R activation but a subset of these cells progress to DB. Our electrophysiological and behavioral results suggest that this mechanism is active during PISs, contributing to a collapse of PV-mediated GABAergic inhibition, dysregulation of excitation/inhibition balance, and increased susceptibility to PISs. PMID:25495999

  14. Autoantibodies enhance agonist action and binding to cardiac muscarinic receptors in chronic Chagas' disease.

    PubMed

    Hernandez, Ciria C; Nascimento, Jose H; Chaves, Elen A; Costa, Patricia C; Masuda, Masako O; Kurtenbach, Eleonora; Campos DE Carvalho, Antonio C; Gimenez, Luis E

    2008-01-01

    Chronic Chagasic patient immunoglobulins (CChP-IgGs) recognize an acidic amino acid cluster at the second extracellular loop (el2) of cardiac M(2)-muscarinic acetylcholine receptors (M(2)AChRs). These residues correspond to a common binding site for various allosteric agents. We characterized the nature of the M(2)AChR/CChP-IgG interaction in functional and radioligand binding experiments applying the same mainstream strategies previously used for the characterization of other allosteric agents. Dose-response curves of acetylcholine effect on heart rate were constructed with data from isolated heart experiments in the presence of CChP or normal blood donor (NBD) sera. In these experiments, CChP sera but not NBD sera increased the efficacy of agonist action by augmenting the onset of bradyarrhythmias and inducing a Hill slope of 2.5. This effect was blocked by gallamine, an M(2)AChR allosteric antagonist. Correspondingly, CChP-IgGs increased acetylcholine affinity twofold and showed negative cooperativity for [(3)H]-N-methyl scopolamine ([(3)H]-NMS) in allosterism binding assays. A peptide corresponding to the M(2)AChR-el2 blocked this effect. Furthermore, dissociation assays showed that the effect of gallamine on the [(3)H]-NMS off-rate was reverted by CChP-IgGs. Finally, concentration-effect curves for the allosteric delay of W84 on [(3)H]-NMS dissociation right shifted from an IC(50) of 33 nmol/L to 78 nmol/L, 992 nmol/L, and 1670 nmol/L in the presence of 6.7 x 10(- 8), 1.33 x 10(- 7), and 2.0 x 10(- 7) mol/L of anti-el2 affinity-purified CChP-IgGs. Taken together, these findings confirmed a competitive interplay of these ligands at the common allosteric site and revealed the novel allosteric nature of the interaction of CChP-IgGs at the M(2)AChRs as a positive cooperativity effect on acetylcholine action. PMID:18702010

  15. Autoantibodies Enhance Agonist Action and Binding to Cardiac Muscarinic Receptors in Chronic Chagas’ Disease

    PubMed Central

    Hernández, Ciria C.; Nascimento, José H.; Chaves, Elen A.; Costa, Patrícia C.; Masuda, Masako O.; Kurtenbach, Eleonora; Campos de Carvalho, Antônio C.; Giménez, Luis E.

    2009-01-01

    Chronic Chagasic patient immunoglobulins (CChP-IgGs) recognize an acidic amino acid cluster at the second extracellular loop (el2) of cardiac M2-muscarinic acetylcholine receptors (M2AChRs). These residues correspond to a common binding site for various allosteric agents. We characterized the nature of the M2AChR/CChP-IgG interaction in functional and radioligand binding experiments applying the same mainstream strategies previously used for the characterization of other allosteric agents. Dose-response curves of acetylcholine effect on heart rate were constructed with data from isolated heart experiments in the presence of CChP or normal blood donor (NBD) sera. In these experiments, CChP sera but not NBD sera increased the efficacy of agonist action by augmenting the onset of bradyarrhythmias and inducing a Hill slope of 2.5. This effect was blocked by gallamine, an M2AChR allosteric antagonist. Correspondingly, CChP-IgGs increased acetylcholine affinity twofold and showed negative cooperativity for [3H]-N-methyl scopolamine ([3H]-NMS) in allosterism binding assays. A peptide corresponding to the M2AChR-el2 blocked this effect. Furthermore, dissociation assays showed that the effect of gallamine on the [3H]-NMS off-rate was reverted by CChP-IgGs. Finally, concentration-effect curves for the allosteric delay of W84 on [3H]-NMS dissociation right shifted from an IC50 of 33 nmol/L to 78 nmol/L, 992 nmol/L, and 1670 nmol/L in the presence of 6.7 × 10−8, 1.33 × 10−7, and 2.0 × 10−7 mol/L of anti-el2 affinity-purified CChP-IgGs. Taken together, these findings confirmed a competitive interplay of these ligands at the common allosteric site and revealed the novel allosteric nature of the interaction of CChP-IgGs at the M2AChRs as a positive cooperativity effect on acetylcholine action. PMID:18702010

  16. Oral administration of D-alanine in monkeys robustly increases plasma and cerebrospinal fluid levels but experimental D-amino acid oxidase inhibitors had minimal effect.

    PubMed

    Rojas, Camilo; Alt, Jesse; Ator, Nancy A; Wilmoth, Heather; Rais, Rana; Hin, Niyada; DeVivo, Michael; Popiolek, Michael; Tsukamoto, Takashi; Slusher, Barbara S

    2016-09-01

    Hypofunction of the N-methyl-d-aspartate (NMDA) receptor is thought to exacerbate psychosis in patients diagnosed with schizophrenia. Consistent with this hypothesis, D-alanine, a co-agonist at the glycine site of the NMDA receptor, was shown to improve positive and cognitive symptoms when used as add-on therapy for schizophrenia treatment. However, D-alanine had to be administered at high doses (~7 g) to observe clinical effects. One possible reason for the high dose is that D-alanine could be undergoing oxidation by D-amino acid oxidase (DAAO) before it reaches the brain. If this is the case, the dose could be reduced by co-administration of D-alanine with a DAAO inhibitor (DAAOi). Early studies with rodents showed that co-administration of D-alanine with 5-chloro-benzo[d]isoxazol-3-ol (CBIO), a prototype DAAOi, significantly enhanced the levels of extracellular D-alanine in the frontal cortex compared with D-alanine alone. Further, the use of CBIO reduced the dose of D-alanine needed to attenuate prepulse inhibition deficits induced by dizocilpine. The objective of the work reported herein was to confirm the hypothesis that DAAO inhibition can enhance D-alanine exposure in a species closer to humans: non-human primates. We report that while oral D-alanine administration to baboons (10 mg/kg) enhanced D-alanine plasma and CSF levels over 20-fold versus endogenous levels, addition of experimental DAAOi to the regimen exhibited a 2.2-fold enhancement in plasma and no measurable effect on CSF levels. The results provide caution regarding the utility of DAAO inhibition to increase D-amino acid levels as treatment for patients with schizophrenia. PMID:27287825

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

    PubMed

    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.

  18. Interleukin-1 administration before lethal irradiation and allogeneic bone marrow transplantation: early transient increase of peripheral granulocytes and successful engraftment with accelerated leukocyte, erythrocyte, and platelet recovery.

    PubMed

    Tiberghien, P; Laithier, V; Mabed, M; Racadot, E; Reynolds, C W; Angonin, R; Loumi, R; Pavy, J J; Cahn, J Y; Noir, A

    1993-04-01

    Administration of interleukin-1 beta (IL-1 beta) before a lethal irradiation with or without allogeneic bone marrow transplantation (BMT) protects greater than 90% of the irradiated mice. To approach the mechanisms responsible for the radioprotective effect of IL-1, we examined the effects of IL-1 pretreatment on engraftment and kinetics of peripheral blood, spleen, and marrow cell reconstitution after irradiation and BMT. Although the BMT was not necessary for the survival of the IL-1-pretreated lethally irradiated mice, allogeneic marrow did engraft in these mice as evaluated in the spleen and marrow 2 months after BMT. IL-1 pretreatment significantly accelerated hematopoietic recovery versus transplanted saline-treated controls with a pronounced enhancement of peripheral leukocyte, platelet, and erythrocyte recovery. Leukocyte recovery in IL-1-pretreated mice was unique in that IL-1 first induced an early transient (maximum at day 7) increase of peripheral granulocytes before accelerating leukocyte recovery after day 11. IL-1 pretreatment also significantly enhanced marrow cell recovery after allogeneic BMT with an eightfold increase in marrow cellularity from day 4 to 11 versus control transplanted mice. When lethal irradiation was not followed by allogeneic BMT. IL-1 pretreatment also affected the peripheral reconstitution of leukocytes, platelets, and erythrocytes. Interestingly, in the absence of BMT, IL-1 also induced an early circulation of peripheral granulocytes. Overall, our data demonstrate that a single administration of IL-1 before lethal irradiation and allogeneic BMT can induce an early transient increase of circulating granulocytes, followed by an accelerated multilineage recovery and long-term allogeneic engraftment. PMID:8461477

  19. Tolerance to Ethanol or Nicotine Results in Increased Ethanol Self-Administration and Long-Term Depression in the Dorsolateral Striatum.

    PubMed

    Abburi, Chandrika; Wolfman, Shannon L; Metz, Ryan A E; Kamber, Rinya; McGehee, Daniel S; McDaid, John

    2016-01-01

    Ethanol (EtOH) and nicotine are the most widely coabused drugs. Tolerance to EtOH intoxication, including motor impairment, results in greater EtOH consumption and may result in a greater likelihood of addiction. Previous studies suggest that cross-tolerance between EtOH and nicotine may contribute to the abuse potential of these drugs. Here we demonstrate that repeated intermittent administration of either EtOH or nicotine in adult male Sprague Dawley rats results in tolerance to EtOH-induced motor impairment and increased EtOH self-administration. These findings suggest that nicotine and EtOH cross-tolerance results in decreased aversive and enhanced rewarding effects of EtOH. Endocannabinoid signaling in the dorsolateral striatum (DLS) has been implicated in both EtOH tolerance and reward, so we investigated whether nicotine or EtOH pretreatment might modulate endocannabinoid signaling in this region. Using similar EtOH and nicotine pretreatment methods resulted in increased paired-pulse ratios of evoked EPSCs in enkephalin-positive medium spiny neurons in DLS slices. Thus, EtOH and nicotine pretreatment may modulate glutamatergic synapses in the DLS presynaptically. Bath application of the CB1 receptor agonist Win 55,2-212 increased the paired-pulse ratio of evoked EPSCs in control slices, while Win 55,2-212 had no effect on paired-pulse ratio in slices from either EtOH- or nicotine-pretreated rats. Consistent with these effects, nicotine pretreatment occluded LTD induction by high-frequency stimulation of the corticostriatal inputs to the dorsolateral striatum. These results suggest that nicotine and EtOH pretreatment modulates striatal synapses to induce tolerance to the motor-impairing effects of EtOH, which may contribute to nicotine and EtOH coabuse. PMID:27517088

  20. Tolerance to Ethanol or Nicotine Results in Increased Ethanol Self-Administration and Long-Term Depression in the Dorsolateral Striatum

    PubMed Central

    Abburi, Chandrika; Wolfman, Shannon L.; Metz, Ryan A. E.; Kamber, Rinya

    2016-01-01

    Abstract Ethanol (EtOH) and nicotine are the most widely coabused drugs. Tolerance to EtOH intoxication, including motor impairment, results in greater EtOH consumption and may result in a greater likelihood of addiction. Previous studies suggest that cross-tolerance between EtOH and nicotine may contribute to the abuse potential of these drugs. Here we demonstrate that repeated intermittent administration of either EtOH or nicotine in adult male Sprague Dawley rats results in tolerance to EtOH-induced motor impairment and increased EtOH self-administration. These findings suggest that nicotine and EtOH cross-tolerance results in decreased aversive and enhanced rewarding effects of EtOH. Endocannabinoid signaling in the dorsolateral striatum (DLS) has been implicated in both EtOH tolerance and reward, so we investigated whether nicotine or EtOH pretreatment might modulate endocannabinoid signaling in this region. Using similar EtOH and nicotine pretreatment methods resulted in increased paired-pulse ratios of evoked EPSCs in enkephalin-positive medium spiny neurons in DLS slices. Thus, EtOH and nicotine pretreatment may modulate glutamatergic synapses in the DLS presynaptically. Bath application of the CB1 receptor agonist Win 55,2-212 increased the paired-pulse ratio of evoked EPSCs in control slices, while Win 55,2-212 had no effect on paired-pulse ratio in slices from either EtOH- or nicotine-pretreated rats. Consistent with these effects, nicotine pretreatment occluded LTD induction by high-frequency stimulation of the corticostriatal inputs to the dorsolateral striatum. These results suggest that nicotine and EtOH pretreatment modulates striatal synapses to induce tolerance to the motor-impairing effects of EtOH, which may contribute to nicotine and EtOH coabuse. PMID:27517088

  1. Methylphenidate treatment beyond adolescence maintains increased cocaine self-administration in the spontaneously hypertensive rat model of attention deficit/hyperactivity disorder.

    PubMed

    Baskin, Britahny M; Dwoskin, Linda P; Kantak, Kathleen M

    2015-04-01

    extending methylphenidate treatment beyond adolescence does not ameliorate explicitly the long-term consequences of adolescent methylphenidate treatment. Pre-session methylphenidate may mask temporarily the detection of an increase in cocaine self-administration following chronic methylphenidate treatment.

  2. Stereoselective recognition of the enantiomers of phenglutarimide and of six related compounds by four muscarinic receptor subtypes.

    PubMed Central

    Waelbroeck, M.; Lazareno, S.; Pfaff, O.; Friebe, T.; Tastenoy, M.; Mutschler, E.; Lambrecht, G.

    1996-01-01

    0.02% of the eutomer. 6. In contrast with the eutomer binding site, replacement of the phenyl ring by a thienyl or cyclohexyl ring did not affect binding of the low affinity enantiomers to the muscarinic receptor or the [3H]-NMS-receptor complex. The replacement of the diethylamino group by a piperidine ring, and N-methylation of the piperidine-2,6 dione moiety increased slightly these enantiomers' potencies. 7. The muscarinic receptors were extremely stereoselective, and had up to 20000 fold lower affinity for the less active enantiomers. However, the stereochemical requirements of the muscarinic receptor subtypes were different for the enantiomers of compounds 1-7, being most stringent at M1 receptors. 8. The weaker enantiomers behaved as competitive antagonists in pharmacological studies, at least in the concentration-range investigated. PMID:8968538

  3. Thermodynamics of antagonist binding to rat muscarinic M2 receptors: antimuscarinics of the pridinol, sila-pridinol, diphenidol and sila-diphenidol type.

    PubMed Central

    Waelbroeck, M.; Camus, J.; Tastenoy, M.; Lambrecht, G.; Mutschler, E.; Kropfgans, M.; Sperlich, J.; Wiesenberger, F.; Tacke, R.; Christophe, J.

    1993-01-01

    1. We studied the effect of temperature on the binding to rat heart M2 muscarinic receptors of antagonists related to the carbon/silicon pairs pridinol/sila-pridinol and diphenidol/sila-diphenidol (including three germanium compounds) and six structurally related pairs of enantiomers [(R)- and (S)-procyclidine, (R)- and (S)-trihexyphenidyl, (R)- and (S)-tricyclamol, (R)- and (S)-trihexyphenidyl methiodide, (R)- and (S)-hexahydro-diphenidol and (R)- and (S)-hexbutinol]. Binding affinities were determined in competition experiments using [3H]-N-methyl-scopolamine chloride as radioligand. The reference drugs were scopolamine and N-methyl-scopolamine bromide. 2. The affinity of the antagonists either increased or decreased with temperature. van't Hoff plots were linear in the 278-310 degrees K temperature range. Binding of all antagonists was entropy driven. Enthalpy changes varied from large negative values (down to -29 kJ mol-1) to large positive values (up to +30 kJ mol-1). 3. (R)-configurated drugs had a 10 to 100 fold greater affinity for M2 receptors than the corresponding (S)-enantiomers. Enthalpy and entropy changes of the respective enantiomers were different but no consistent pattern was observed. 4. When silanols (R3SiOH) were compared to carbinols (R3COH), the affinity increase caused by C/Si exchange varied between 3 and 10 fold for achiral drugs but was negligible in the case of chiral drugs. Silanols induced more favourable enthalpy and less favourable entropy changes than the corresponding carbinols when binding. Organogermanium compounds (R4Ge) when compared to their silicon counterparts (R4Si) showed no significant difference in affinity as well as in enthalpy and entropy changes. 5. Exchange of a cyclohexyl by a phenyl moiety was associated with an increase or a decrease in drug affinity (depending on the absolute configuration in the case of chiral drugs) and generally also with a more favourable enthalpy change and a less favourable entropy change

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

    PubMed Central

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

    2014-01-01

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

  5. Intrathecal Administration of Tempol Reduces Chronic Constriction Injury-Induced Neuropathic Pain in Rats by Increasing SOD Activity and Inhibiting NGF Expression.

    PubMed

    Zhao, Baisong; Pan, Yongying; Wang, Zixin; Tan, Yonghong; Song, Xingrong

    2016-08-01

    We investigate the antinociceptive effect of intrathecal and intraperitoneal tempol administration in a rat model of chronic constriction injury (CCI)-induced neuropathic pain and explore the underlying antinociceptive mechanisms of tempol. Rats were randomly assigned to four groups (n = 8 per group): sham group, CCI group, Tem1 group (intrathecal injection of tempol), and Tem2 group (intraperitoneal injection of tempol). Neuropathic pain was induced by CCI of the sciatic nerve. Tempol was intrathecally or intraperitoneally administered daily for 7 days beginning on postoperative day one. The mechanical withdrawal threshold and thermal withdrawal latency were tested on preoperative day 3 and postoperative days 1, 3, 5, 7, 10, 14, and 21. Structural changes were examined by hematoxylin and eosin staining, toluidine blue staining, and electron microscopy. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were determined using the thiobarbituric acid and nitroblue tetrazolium methods, respectively. Nerve growth factor (NGF) expression levels were determined by immunohistochemistry and Western blot. Intrathecal, but not intraperitoneal, injection of tempol produced a persistent antinociceptive effect. Intraperitoneal injection of tempol did not result in high enough concentration of tempol in the cerebrospinal fluid. Intrathecal, but not intraperitoneal, injection of tempol inhibited CCI-induced structural damage in the spinal cord reduced MDA levels, and increased SOD activities in the spinal cord. Furthermore, intrathecal, but not intraperitoneal, injection of tempol further downregulated the expression of NGF in the spinal cord following CCI, and this effect was blocked by p38MAPK inhibitor. Intrathecal injection of tempol produces antinociceptive effects and reduces CCI-induced structural damage in the spinal cord by increasing SOD activities and downregulating the expression of NGF via the p38MAPK pathway. Intraperitoneal administration of tempol does

  6. Administration of kisspeptin-54 into discrete regions of the hypothalamus potently increases plasma luteinising hormone and testosterone in male adult rats.

    PubMed

    Patterson, M; Murphy, K G; Thompson, E L; Patel, S; Ghatei, M A; Bloom, S R

    2006-05-01

    Kisspeptin-54 is the peptide product of the KiSS-1 gene and an endogenous agonist of the GPR54 receptor. KiSS-1 was initially discovered as a metastasis suppressor gene, but recent studies demonstrate that the kisspeptin/GPR54 system is a key regulator of the reproductive system. Disrupted GPR54 signalling causes hypogonadotrophic hypogonadism in rodents and man. Intracerebroventricular or peripheral administration of kisspeptin potently stimulates the hypothalamic-pituitary-gonadal (HPG) axis via the hypothalamic gonadotrophin-releasing hormone system. We have investigated the effect of injection of kisspeptin-54 into discrete hypothalamic regions on the HPG axis. To construct a dose-response curve for the effects of intrahypothalamic kisspeptin administration, adult male Wistar rats were cannulated into the medial preoptic area (MPOA) at the level of the organum vasculosum laminae terminalis (OVLT). Kisspeptin-54 was injected into the MPOA at doses of 0.01, 0.1, 1, 10 and 100 pmol. At 60 min following injection of 1, 10 or 100 pmol kisspeptin-54, plasma luteinising hormone (LH) and total testosterone levels were significantly increased. Adult male Wistar rats were then cannulated into the rostral preoptic area at the level of the OVLT (RPOA), the MPOA, the paraventricular (PVN), dorsomedial (DMN) and arcuate hypothalamic nuclei, and the lateral hypothalamic area. A dose of 1 pmol kisspeptin-54 was administered into all areas. The circulating levels of LH and total testosterone were significantly increased 60 min postinjection of kisspeptin-54 into the RPOA, MPOA, PVN and arcuate nucleus. Our results suggest that kisspeptin may mediate its effects on the HPG axis via these regions of the hypothalamus.

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

    PubMed

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

    2015-02-01

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

  8. Administration of thimerosal to infant rats increases overflow of glutamate and aspartate in the prefrontal cortex: protective role of dehydroepiandrosterone sulfate.

    PubMed

    Duszczyk-Budhathoki, Michalina; Olczak, Mieszko; Lehner, Malgorzata; Majewska, Maria Dorota

    2012-02-01

    Thimerosal, a mercury-containing vaccine preservative, is a suspected factor in the etiology of neurodevelopmental disorders. We previously showed that its administration to infant rats causes behavioral, neurochemical and neuropathological abnormalities similar to those present in autism. Here we examined, using microdialysis, the effect of thimerosal on extracellular levels of neuroactive amino acids in the rat prefrontal cortex (PFC). Thimerosal administration (4 injections, i.m., 240 μg Hg/kg on postnatal days 7, 9, 11, 15) induced lasting changes in amino acid overflow: an increase of glutamate and aspartate accompanied by a decrease of glycine and alanine; measured 10-14 weeks after the injections. Four injections of thimerosal at a dose of 12.5 μg Hg/kg did not alter glutamate and aspartate concentrations at microdialysis time (but based on thimerosal pharmacokinetics, could have been effective soon after its injection). Application of thimerosal to the PFC in perfusion fluid evoked a rapid increase of glutamate overflow. Coadministration of the neurosteroid, dehydroepiandrosterone sulfate (DHEAS; 80 mg/kg; i.p.) prevented the thimerosal effect on glutamate and aspartate; the steroid alone had no influence on these amino acids. Coapplication of DHEAS with thimerosal in perfusion fluid also blocked the acute action of thimerosal on glutamate. In contrast, DHEAS alone reduced overflow of glycine and alanine, somewhat potentiating the thimerosal effect on these amino acids. Since excessive accumulation of extracellular glutamate is linked with excitotoxicity, our data imply that neonatal exposure to thimerosal-containing vaccines might induce excitotoxic brain injuries, leading to neurodevelopmental disorders. DHEAS may partially protect against mercurials-induced neurotoxicity. PMID:22015977

  9. In vivo and in vitro pharmacological characterization of SVT-40776, a novel M3 muscarinic receptor antagonist, for the treatment of overactive bladder

    PubMed Central

    Salcedo, C; Davalillo, S; Cabellos, J; Lagunas, C; Balsa, D; Pérez-del-Pulgar, S; Ballarín, M; Fernández, AG

    2009-01-01

    Background and purpose: Highly selective M3 muscarinic receptor antagonists may represent a better treatment for overactive bladder syndrome, diminishing side effects. Cardiac side effects of non-selective antimuscarinics have been associated with activity at M2 receptors as these receptors are mainly responsible for muscarinic receptor-dependent bradycardia. We have investigated a novel antimuscarinic, SVT-40776, highly selective for M3 over M2 receptors (Ki = 0.19 nmol·L−1 for M3 receptor affinity). This study reports the functional activity of SVT-40776 in the bladder, relative to its activity in atria. Experimental approach: In vitro and ex vivo (oral dosing) inhibition of mouse detrusor and atrial contractile responses to carbachol were used to study the functional activity of SVT-40776. The in vivo efficacy of SVT-40776 was characterized by suppression of isovolumetric spontaneous bladder contractions in anaesthetized guinea pigs after intravenous administration. Key results: SVT-40776 was the most potent in inhibiting carbachol-induced bladder contractions of the anti-cholinergic agents tested, without affecting atrial contractions over the same range of concentrations. SVT-40776 exhibited the highest urinary versus cardiac selectivity (199-fold). In the guinea pig in vivo model, SVT-40776 inhibited 25% of spontaneous bladder contractions at a very low dose (6.97 µg·kg−1 i.v), without affecting arterial blood pressure. Conclusions and implications: SVT-40776 is a potent inhibitor of M3 receptor-related detrusor contractile activity. The absence of effects on isolated atria preparations represents an interesting characteristic and suggests that SVT-40776 may lack unwanted cardiac effects; a feature especially relevant in a compound intended to treat mainly elderly patients. British Journal of Pharmacology (2009) doi:10.1111/j.1476-5381.2008.00082.x PMID:19222482

  10. Diphenyl diselenide administration enhances cortical mitochondrial number and activity by increasing hemeoxygenase type 1 content in a methylmercury-induced neurotoxicity mouse model.

    PubMed

    Glaser, Viviane; Martins, Roberta de Paula; Vieira, Ana Julia Hoffmann; Oliveira, Eliana de Medeiros; Straliotto, Marcos Raniel; Mukdsi, Jorge Humberto; Torres, Alicia Inés; de Bem, Andreza Fabro; Farina, Marcelo; da Rocha, João Batista Teixeira; De Paul, Ana Lucia; Latini, Alexandra

    2014-05-01

    Interest in biochemistry of organoselenium compound has increased in the last decades, mainly due to their chemical and biological activities. Here, we investigated the protective effect of diphenyl diselenide (PhSe)2 (5 μmol/kg), in a mouse model of methylmercury (MeHg)-induced brain toxicity. Swiss male mice were divided into four experimental groups: control, (PhSe)2 (5 μmol/kg, subcutaneous administration), MeHg (40 mg/L, in tap water), and MeHg + (PhSe)2. After the treatment (21 days), the animals were killed and the cerebral cortex was analyzed. Electron microscopy indicated an enlarged and fused mitochondria leading to a reduced number of organelles, in the MeHg-exposed mice. Furthermore, cortical creatine kinase activity, a sensitive mitochondrial oxidative stress sensor, was almost abolished by MeHg. Subcutaneous (PhSe)2 co-treatment rescued from MeHg-induced mitochondrial alterations. (PhSe)2 also behaved as an enhancer of mitochondrial biogenesis, by increasing cortical mitochondria content in mouse-receiving (PhSe)2 alone. Mechanistically, (PhSe)2 (1 μM; 24 h) would trigger the cytoprotective Nrf-2 pathway for activating target genes, since astroglial cells exposed to the chalcogen showed increased content of hemeoxygenase type 1, a sensitive marker of the activation of this via. Thus, it is proposed that the (PhSe)2-neuroprotective effect might be linked to its mitoprotective activity. PMID:24623265

  11. Oral administration of copper to rats leads to increased lymphocyte cellular DNA degradation by dietary polyphenols: implications for a cancer preventive mechanism.

    PubMed

    Khan, Husain Y; Zubair, Haseeb; Ullah, Mohd F; Ahmad, Aamir; Hadi, Sheikh M

    2011-12-01

    To account for the observed anticancer properties of plant polyphenols, we have earlier proposed a mechanism which involves the mobilization of endogenous copper ions by polyphenols leading to the generation of reactive oxygen species (ROS) that serve as proximal DNA cleaving agents and lead to cell death. Over the last decade we have proceeded to validate our hypothesis with considerable success. As a further confirmation of our hypothesis, in this paper we first show that oral administration of copper to rats leads to elevated copper levels in lymphocytes. When such lymphocytes with a copper overload were isolated and treated with polyphenols EGCG, genistein and resveratrol, an increased level of DNA breakage was observed. Further, preincubation of lymphocytes having elevated copper levels with the membrane permeable copper chelator neocuproine, resulted in inhibition of polyphenol induced DNA degradation. However, membrane impermeable chelator of copper bathocuproine, as well as iron and zinc chelators were ineffective in causing such inhibition in DNA breakage, confirming the involvement of endogenous copper in polyphenol induced cellular DNA degradation. It is well established that serum and tissue concentrations of copper are greatly increased in various malignancies. In view of this fact, the present results further confirm our earlier findings and strengthen our hypothesis that an important anticancer mechanism of plant polyphenols could be the mobilization of intracellular copper leading to ROS-mediated cellular DNA breakage. In this context, it may be noted that cancer cells are under considerable oxidative stress and increasing such stress to cytotoxic levels could be a successful anticancer approach.

  12. Effects of dopamine D1-like and D2-like antagonists on cocaine discrimination in muscarinic receptor knockout mice.

    PubMed

    Thomsen, Morgane; Caine, Simon Barak

    2016-04-01

    Muscarinic and dopamine brain systems interact intimately, and muscarinic receptor ligands, like dopamine ligands, can modulate the reinforcing and discriminative stimulus (S(D)) effects of cocaine. To enlighten the dopamine/muscarinic interactions as they pertain to the S(D) effects of cocaine, we evaluated whether muscarinic M1, M2 or M4 receptors are necessary for dopamine D1 and/or D2 antagonist mediated modulation of the S(D) effects of cocaine. Knockout mice lacking M1, M2, or M4 receptors, as well as control wild-type mice and outbred Swiss-Webster mice, were trained to discriminate 10mg/kg cocaine from saline in a food-reinforced drug discrimination procedure. Effects of pretreatments with the dopamine D1 antagonist SCH 23390 and the dopamine D2 antagonist eticlopride were evaluated. In intact mice, both SCH 23390 and eticlopride attenuated the cocaine discriminative stimulus effect, as expected. SCH 23390 similarly attenuated the cocaine discriminative stimulus effect in M1 knockout mice, but not in mice lacking M2 or M4 receptors. The effects of eticlopride were comparable in each knockout strain. These findings demonstrate differences in the way that D1 and D2 antagonists modulate the S(D) effects of cocaine, D1 modulation being at least partially dependent upon activity at the inhibitory M2/M4 muscarinic subtypes, while D2 modulation appeared independent of these systems. PMID:26874213

  13. Lack of increased signal intensity in the dentate nucleus after repeated administration of a macrocyclic contrast agent in multiple sclerosis: An observational study.

    PubMed

    Eisele, Philipp; Alonso, Angelika; Szabo, Kristina; Ebert, Anne; Ong, Melissa; Schoenberg, Stefan O; Gass, Achim

    2016-09-01

    Recently, several studies reported increased signal intensity (SI) in the dentate nucleus (DN) after repeated application of gadolinium-based contrast agents (GBCAs), suggesting a deposition of gadolinium in this location. Patients with relapsing-remitting multiple sclerosis (RRMS) frequently show increased permeability of the blood-brain barrier as part of the inflammatory process in the brain parenchyma, which theoretically might increase the risk of gadolinium deposition. In this retrospective study, we investigated a possible increasing SI in the DN after repeated administrations of the macrocyclic contrast agent gadoterate meglumine.Forty-one RRMS patients (33 women, mean age 38 years) with at least 6 prior gadolinium-enhanced examinations (single dose gadoterate meglumine) were identified. A total of 279 unenhanced T1-weighted examinations were analyzed.SI ratio differences did not differ between the first and last MRI examination, neither for the DN-to-pons ratio (P = 0.594) nor for the DN-to-cerebellum ratio (P = 0.847). There was no correlation between the mean DN-to-pons, or between the mean DN-to-cerebellum SI ratio and the number of MRI examinations (P = 0.848 and 0.891), disease duration (P = 0.676 and 0.985), and expanded disability status scale (EDSS) (P = 0.639 and 0.945).We found no signal increases in the DN after a minimum of 6 injections of the macrocyclic GBCA gadoterate meglumine in RRMS patients. This warrants further investigations in regard to the true pathophysiologic basis of intracerebral gadolinium deposition. PMID:27684794

  14. Oral Administration of Faecalibacterium prausnitzii Decreased the Incidence of Severe Diarrhea and Related Mortality Rate and Increased Weight Gain in Preweaned Dairy Heifers.

    PubMed

    Foditsch, Carla; Pereira, Richard Van Vleck; Ganda, Erika Korzune; Gomez, Marilia Souza; Marques, Eduardo Carvalho; Santin, Thiago; Bicalho, Rodrigo Carvalho

    2015-01-01

    Probiotics are a promising alternative to improve food animal productivity and health. However, scientific evidence that specific microbes can be used to benefit animal health and performance is limited. The objective of this study was to evaluate the effects of administering a live culture of Faecalibacterium prausnitzii to newborn dairy calves on subsequent growth, health, and fecal microbiome. Initially, a safety trial was conducted using 30 newborn bull calves to assess potential adverse effects of the oral and rectal administration of F. prausnitzii to neonatal calves. No adverse reactions, such as increased body temperature or heart and respiratory rates, were observed after the administration of the treatments. All calves survived the experimental period, and there was no difference in fecal consistency score, attitude, appetite or dehydration between the treatment groups. The rectal route was not an efficient practice while the oral route ensures that the full dose is administered to the treated calves. Subsequently, a randomized field trial was completed in a commercial farm with preweaned calves. A total of 554 Holstein heifers were assigned to one of two treatment groups: treated calves (FPTRT) and non-treated calves (control). Treated calves received two oral doses of F. prausnitzii, one at treatment assignment (1st week) and another one week later. The FPTRT group presented significantly lower incidence of severe diarrhea (3.1%) compared with the control group (6.8%). Treated calves also had lower mortality rate associated with severe diarrhea (1.5%) compared to control calves (4.4%). Furthermore, FPTRT calves gained significantly more weight, 4.4 kg over the preweaning period, than controls calves. The relative abundance of F. prausnitzii in the fecal microbiota was significantly higher in the 3rd and 5th weeks of life of FPTRT calves than of the control calves, as revealed by sequencing of the 16S rRNA gene. Our findings showed that oral

  15. Oral Administration of Faecalibacterium prausnitzii Decreased the Incidence of Severe Diarrhea and Related Mortality Rate and Increased Weight Gain in Preweaned Dairy Heifers

    PubMed Central

    Foditsch, Carla; Pereira, Richard Van Vleck; Ganda, Erika Korzune; Gomez, Marilia Souza; Marques, Eduardo Carvalho; Santin, Thiago; Bicalho, Rodrigo Carvalho

    2015-01-01

    Probiotics are a promising alternative to improve food animal productivity and health. However, scientific evidence that specific microbes can be used to benefit animal health and performance is limited. The objective of this study was to evaluate the effects of administering a live culture of Faecalibacterium prausnitzii to newborn dairy calves on subsequent growth, health, and fecal microbiome. Initially, a safety trial was conducted using 30 newborn bull calves to assess potential adverse effects of the oral and rectal administration of F. prausnitzii to neonatal calves. No adverse reactions, such as increased body temperature or heart and respiratory rates, were observed after the administration of the treatments. All calves survived the experimental period, and there was no difference in fecal consistency score, attitude, appetite or dehydration between the treatment groups. The rectal route was not an efficient practice while the oral route ensures that the full dose is administered to the treated calves. Subsequently, a randomized field trial was completed in a commercial farm with preweaned calves. A total of 554 Holstein heifers were assigned to one of two treatment groups: treated calves (FPTRT) and non-treated calves (control). Treated calves received two oral doses of F. prausnitzii, one at treatment assignment (1st week) and another one week later. The FPTRT group presented significantly lower incidence of severe diarrhea (3.1%) compared with the control group (6.8%). Treated calves also had lower mortality rate associated with severe diarrhea (1.5%) compared to control calves (4.4%). Furthermore, FPTRT calves gained significantly more weight, 4.4 kg over the preweaning period, than controls calves. The relative abundance of F. prausnitzii in the fecal microbiota was significantly higher in the 3rd and 5th weeks of life of FPTRT calves than of the control calves, as revealed by sequencing of the 16S rRNA gene. Our findings showed that oral

  16. Oral Administration of Faecalibacterium prausnitzii Decreased the Incidence of Severe Diarrhea and Related Mortality Rate and Increased Weight Gain in Preweaned Dairy Heifers.

    PubMed

    Foditsch, Carla; Pereira, Richard Van Vleck; Ganda, Erika Korzune; Gomez, Marilia Souza; Marques, Eduardo Carvalho; Santin, Thiago; Bicalho, Rodrigo Carvalho

    2015-01-01

    Probiotics are a promising alternative to improve food animal productivity and health. However, scientific evidence that specific microbes can be used to benefit animal health and performance is limited. The objective of this study was to evaluate the effects of administering a live culture of Faecalibacterium prausnitzii to newborn dairy calves on subsequent growth, health, and fecal microbiome. Initially, a safety trial was conducted using 30 newborn bull calves to assess potential adverse effects of the oral and rectal administration of F. prausnitzii to neonatal calves. No adverse reactions, such as increased body temperature or heart and respiratory rates, were observed after the administration of the treatments. All calves survived the experimental period, and there was no difference in fecal consistency score, attitude, appetite or dehydration between the treatment groups. The rectal route was not an efficient practice while the oral route ensures that the full dose is administered to the treated calves. Subsequently, a randomized field trial was completed in a commercial farm with preweaned calves. A total of 554 Holstein heifers were assigned to one of two treatment groups: treated calves (FPTRT) and non-treated calves (control). Treated calves received two oral doses of F. prausnitzii, one at treatment assignment (1st week) and another one week later. The FPTRT group presented significantly lower incidence of severe diarrhea (3.1%) compared with the control group (6.8%). Treated calves also had lower mortality rate associated with severe diarrhea (1.5%) compared to control calves (4.4%). Furthermore, FPTRT calves gained significantly more weight, 4.4 kg over the preweaning period, than controls calves. The relative abundance of F. prausnitzii in the fecal microbiota was significantly higher in the 3rd and 5th weeks of life of FPTRT calves than of the control calves, as revealed by sequencing of the 16S rRNA gene. Our findings showed that oral

  17. Non-invasive administration of 17β-estradiol rapidly increases aggressive behavior in non-breeding, but not breeding, male song sparrows.

    PubMed

    Heimovics, Sarah A; Ferris, Jennifer K; Soma, Kiran K

    2015-03-01

    17β-Estradiol (E2) acts in the brain via genomic and non-genomic mechanisms to influence physiology and behavior. There is seasonal plasticity in the mechanisms by which E2 activates aggression, and non-genomic mechanisms appear to predominate during the non-breeding season. Male song sparrows (Melospiza melodia) display E2-dependent territorial aggression throughout the year. Field studies show that song sparrow aggression during a territorial intrusion is similar in the non-breeding and breeding seasons, but aggression after an intrusion ends differs seasonally. Non-breeding males stop behaving aggressively within minutes whereas breeding males remain aggressive for hours. We hypothesize that this seasonal plasticity in the persistence of aggression relates to seasonal plasticity in E2 signaling. We used a non-invasive route of E2 administration to compare the non-genomic (within 20min) effects of E2 on aggressive behavior in captive non-breeding and breeding season males. E2 rapidly increased barrier contacts (attacks) during an intrusion by 173% in non-breeding season males only. Given that these effects were observed within 20min of E2 administration, they likely occurred via a non-genomic mechanism of action. The present data, taken together with past work, suggest that environmental cues associated with the non-breeding season influence the molecular mechanisms through which E2 influences behavior. In song sparrows, transient expression of aggressive behavior during the non-breeding season is highly adaptive: it minimizes energy expenditure and maximizes the amount of time available for foraging. In all, these data suggest the intriguing possibility that aggression in the non-breeding season may be activated by a non-genomic E2 mechanism due to the fitness benefits associated with rapid and transient expression of aggression.

  18. Pharmacological and ionic characterizations of the muscarinic receptors modulating (/sup 3/H)acetylcholine release from rat cortical synaptosomes

    SciTech Connect

    Meyer, E.M.; Otero, D.H.

    1985-05-01

    The muscarinic receptors that modulate acetylcholine release from rat cortical synaptosomes were characterized with respect to sensitivity to drugs that act selectively at M1 or M2 receptor subtypes, as well as to changes in ionic strength and membrane potential. The modulatory receptors appear to be of the M2 type, since they are activated by carbachol, acetylcholine, methacholine, oxotremorine, and bethanechol, but not by pilocarpine, and are blocked by atropine, scopolamine, and gallamine (at high concentrations), but not by pirenzepine or dicyclomine. The ED50S for carbachol, acetylcholine, and oxotremorine are less than 10 microM, suggesting that the high affinity state of the receptor is functional. High ionic strength induced by raising the NaCl concentration has no effect on agonist (oxotremorine) potency, but increases the efficacy of this compound, which disagrees with receptor-binding studies. On the other hand, depolarization with either KCl or with veratridine (20 microM) reduces agonist potencies by approximately an order of magnitude, suggesting a potential mechanism for receptor regulation.

  19. A supersulfated low-molecular-weight heparin (IK-SSH) increases plasma levels of free and total tissue factor pathway inhibitor after intravenous and subcutaneous administration in humans.

    PubMed

    Kaiser, B; Glusa, E; Hoppensteadt, D A; Breddin, H K; Amiral, J; Fareed, J

    1998-09-01

    Unfractionated as well as low-molecular-weight heparins (LMWH) are known to cause an increase in blood levels of tissue factor pathway inhibitor (TFPI). To study the effect of a newly developed supersulfated LMWH (IK-SSH, Iketon Farmaceutici) on TFPI concentrations in human plasma, the compound was injected into volunteers at doses of 0.14, 0.33 and 0.66 mg/kg intravenously or 0.33, 0.66 and 1.0 mg/kg subcutaneously. At certain known times blood was drawn and plasma levels of both total and free TFPI were measured using enzyme-linked immunosorbent assay methodology. Baseline plasma concentrations of TFPI were 72.2+/-3.1 ng/ml for total and 10.8+/-0.8 ng/ml for free TFPI. Intravenous or subcutaneous injection of IK-SSH led to a strong and long-lasting rise in TFPI levels which were increased more than 5-fold for total TFPI and more than 30-fold for free TFPI. Maximum TFPI levels were reached 5-10 min after intravenous and 60 min after subcutaneous administration. IK-SSH caused prolongation of ex-vivo clotting times in the APTT and Heptest assay, whereas thrombin time was not affected. Anticoagulant actions of IK-SSH showed a significant correlation to plasma concentrations of TFPI and they are thought to be based at least partially on the release of TFPI from vascular sites.

  20. An increase in sensitivity of rat cingulate cortical neurones to substance P occurs following withdrawal of chronic administration of antidepressant drugs.

    PubMed Central

    Jones, R. S.; Olpe, H. R.

    1984-01-01

    The sensitivity of neurones in the cingulate cortex of the rat to iontophoretically applied substance P (SP) was tested one hour after a single dose of various antidepressant drugs and also 1 day following the termination of a chronic dosing schedule (14 once daily injections) of the same agents. One hour after a single injection of desipramine (DMI), chlorimipramine (CMI), trimipramine (TMI) or zimelidine ( ZIM ) (all at 10 mg kg-1 i.p.) there was no change in the mean size of excitatory responses to SP compared to those before the injection. There was a tendency towards a decrease in the TMI group. One day following the last of 14 consecutive daily injections (10 mg kg-1 i.p.) of the above agents there was a significant increase in the size of excitatory responses to SP compared to those in rats receiving daily saline injections. However, 14 days of treatment with DMI did not alter the responses to L-glutamate. Similar chronic dosing schedules with either diazepam (5 mg kg-1) or fluphenazine (5 mg kg-1) did not affect the responses to SP. Thus chronic but not acute administration of antidepressant drugs results in an increase in the sensitivity of neurones, in the cingulate cortex of the rat, to SP. PMID:6202353

  1. Regulation of muscarinic acetylcholine receptor-mediated synaptic responses by adenosine receptors in the rat hippocampus.

    PubMed Central

    Morton, R A; Davies, C H

    1997-01-01

    1. Intracellular current clamp recordings were made from CA1 pyramidal neurones in rat hippocampal slices. Experiments were performed in the presence of ionotropic glutamate receptor antagonists and gamma-aminobutyric acid (GABA) receptor antagonists to block all fast excitatory and inhibitory synaptic transmission. A single stimulus, delivered extracellularly in the stratum oriens, caused a reduction in spike frequency adaptation in response to a depolarizing current step delivered 2 s after the stimulus. A 2- to 10-fold increase in stimulus intensity evoked a slow excitatory postsynaptic potential (EPSP) which was associated with a small increase in input resistance. The peak amplitude of the EPSP occurred approximately 2.5 s after the stimulus and its magnitude (up to 30 mV) and duration (10-50 s) increased with increasing stimulus intensity. 2. The slow EPSP was unaffected by the metabotropic glutamate receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine ((+)-MCPG; 1000 microM) but was greatly enhanced by the acetylcholinesterase inhibitor physostigmine (1-5 microM). Both the slow EPSP and the stimulus-evoked reduction in spike frequency adaptation were inhibited by the muscarinic acetylcholine receptor (mAChR) antagonist atropine (1-5 microM). These results are consistent with these effects being mediated by mAChRs. 3. Both the mAChR-mediated EPSP (EPSPm) and the associated reduction in spike frequency adaptation were reversibly depressed (up to 97%) by either adenosine (100 microM) or its non-hydrolysable analogue 2-chloroadenosine (CADO; 0.1-5.0 microM). These effects were often accompanied by postsynaptic hyperpolarization (up to 8 mV) and a reduction in input resistance (up to 11%). The selective adenosine A1 receptor agonists 2-chloro-N6-cyclopentyladenosine (CCPA; 0.1-0.4 microM) and R(-)N6-(2-phenylisopropyl)-adenosine (R-PIA; 1 microM) both depressed the EPSPm. In contrast, the adenosine A2A receptor agonist 2-p-(2-carboxyethyl)-phenethylamino-5

  2. Role of Muscarinic Acetylcholine Receptors in Serial Feature-Positive Discrimination Task during Eyeblink Conditioning in Mice.

    PubMed

    Rahman, Md Ashrafur; Tanaka, Norifumi; Usui, Koji; Kawahara, Shigenori

    2016-01-01

    We investigated the role of muscarinic acetylcholine receptors (mAChRs) in eyeblink serial feature-positive discrimination learning in mice using the mAChR antagonist. A 2-s light cue was delivered 5 or 6 s before the presentation of a 350-ms tone paired with a 100-ms periorbital electrical shock (cued trial) but not before the tone-alone presentation (non-cued trial). Mice received 30 cued and 30 non-cued trials each day in a random order. We found that saline-injected control mice were successfully discriminating between cued and non-cued trials within a few days of conditioning. The mice responded more frequently to the tone in cued trials than in non-cued trials. Analysis of conditioned response (CR) dynamics revealed that the CR onset latency was shorter in cued trials than in non-cued trials, despite the CR peak amplitude not differing significantly between the two conditions. In contrast, scopolamine-injected mice developed an equal number of CRs with similar temporal patterns irrespective of the presence of the cue during the 7 days of conditioning, indicating in a failure to acquire conditional discrimination. In addition, the scopolamine administration to the control mice after they had successfully acquired discrimination did not impair the conditional discrimination and expression of pre-acquired CR. These results suggest that mAChRs may play a pivotal role in memory formation in the conditional brain state associated with the feature cue; however they are unlikely to be involved in the development of discrimination after conditional memory had formed in the serial feature-positive discrimination task during eyeblink conditioning. PMID:26808980

  3. Role of Muscarinic Acetylcholine Receptors in Serial Feature-Positive Discrimination Task during Eyeblink Conditioning in Mice

    PubMed Central

    Rahman, Md. Ashrafur; Tanaka, Norifumi; Usui, Koji; Kawahara, Shigenori

    2016-01-01

    We investigated the role of muscarinic acetylcholine receptors (mAChRs) in eyeblink serial feature-positive discrimination learning in mice using the mAChR antagonist. A 2-s light cue was delivered 5 or 6 s before the presentation of a 350-ms tone paired with a 100-ms periorbital electrical shock (cued trial) but not before the tone-alone presentation (non-cued trial). Mice received 30 cued and 30 non-cued trials each day in a random order. We found that saline-injected control mice were successfully discriminating between cued and non-cued trials within a few days of conditioning. The mice responded more frequently to the tone in cued trials than in non-cued trials. Analysis of conditioned response (CR) dynamics revealed that the CR onset latency was shorter in cued trials than in non-cued trials, despite the CR peak amplitude not differing significantly between the two conditions. In contrast, scopolamine-injected mice developed an equal number of CRs with similar temporal patterns irrespective of the presence of the cue during the 7 days of conditioning, indicating in a failure to acquire conditional discrimination. In addition, the scopolamine administration to the control mice after they had successfully acquired discrimination did not impair the conditional discrimination and expression of pre-acquired CR. These results suggest that mAChRs may play a pivotal role in memory formation in the conditional brain state associated with the feature cue; however they are unlikely to be involved in the development of discrimination after conditional memory had formed in the serial feature-positive discrimination task during eyeblink conditioning. PMID:26808980

  4. Muscarinic signaling influences the patterning and phenotype of cholinergic amacrine cells in the developing chick retina

    PubMed Central

    Stanke, Jennifer J; Lehman, Bret; Fischer, Andy J

    2008-01-01

    Background Many studies in the vertebrate retina have characterized the differentiation of amacrine cells as a homogenous class of neurons, but little is known about the genes and factors that regulate the development of distinct types of amacrine cells. Accordingly, the purpose of this study was to characterize the development of the cholinergic amacrine cells and identify factors that influence their development. Cholinergic amacrine cells in the embryonic chick retina were identified by using antibodies to choline acetyltransferase (ChAT). Results We found that as ChAT-immunoreactive cells differentiate they expressed the homeodomain transcription factors Pax6 and Islet1, and the cell-cycle inhibitor p27kip1. As differentiation proceeds, type-II cholinergic cells, displaced to the ganglion cell layer, transiently expressed high levels of cellular retinoic acid binding protein (CRABP) and neurofilament, while type-I cells in the inner nuclear layer did not. Although there is a 1:1 ratio of type-I to type-II cells in vivo, in dissociated cell cultures the type-I cells (ChAT-positive and CRABP-negative) out-numbered the type-II cells (ChAT and CRABP-positive cells) by 2:1. The relative abundance of type-I to type-II cells was not influenced by Sonic Hedgehog (Shh), but was affected by compounds that act at muscarinic acetylcholine receptors. In addition, the abundance and mosaic patterning of type-II cholinergic amacrine cells is disrupted by interfering with muscarinic signaling. Conclusion We conclude that: (1) during development type-I and type-II cholinergic amacrine cells are not homotypic, (2) the phenotypic differences between these subtypes of cells is controlled by the local microenvironment, and (3) appropriate levels of muscarinic signaling between the cholinergic amacrine cells are required for proper mosaic patterning. PMID:18254959

  5. Selectivity of muscarinic agonists including (+/-)-aceclidine and antimuscarinics on the human intraocular muscles.

    PubMed

    Ishikawa, H; DeSantis, L; Patil, P N

    1998-08-01

    The average EC50 value and the maximum response of carbachol on the human circular ciliary muscle obtained within 24 h of postmortem hypoxia was 517 nmol/l and 135 mg, respectively. These values for carbachol did not differ significantly from that of the longitudinal ciliary muscle. However, when tested at 1 mumol/l of carbachol, the peak response of the longitudinal muscle occurred at 59 sec vs 173 sec for that of the circular muscle of 70 year old donors. The relative potency of the muscarinic agonists on the circular muscle was oxotremorine-M, 1 > carbachol, 1/4 > pilocarpine, 1/19 > aceclidine, 1/132. The relative order of potency of agonists was similar for the longitudinal muscle. Only pilocarpine and aceclidine were partial agonists which produced 80-85% of the maximum response. When compared with the EC50 values of aceclidine on the iris sphincter and the longitudinal ciliary muscles, the agonist potency was only 1/28 for the latter tissue. Implications of these findings in relation to the use of these agonists in glaucoma are discussed. The pKB values of muscarinic antagonists on the circular ciliary muscle were: atropine, 8.8; cyclopentolate, 7.8; tropicamide, 7.4; P.F. HHSiD, 7.0; pirenzepine, 6.4; and methoctramine, 5.7. Nearly equal pKB values of each antagonist were obtained for the longitudinal ciliary muscle and iris sphincter. Based on the affinity constants of various competitive antagonists, the human iris as well as ciliary muscles may contain M3, M2 or M4 subtypes of muscarinic receptors.

  6. Affinity and efficacy of racemic, (+)-, and (-)-methacholine in muscarinic inhibition of [3H]-noradrenaline release.

    PubMed

    Fuder, H; Jung, B

    1985-02-01

    The right postganglionic sympathetic nerves of rat isolated perfused hearts (previously loaded with [3H]-noradrenaline) were stimulated electrically with 10 trains of 10 pulses at 10 Hz. The inhibition by methacholine of stimulation-evoked [3H]-noradrenaline overflow into the perfusate (determined in the presence of corticosterone, desipramine, phentolamine, and propranolol) was taken as a measure for activation of presynaptic muscarinic receptors. The evoked [3H]-noradrenaline overflow was inhibited by (+)-, racemic, and (-)-methacholine in a reversible and concentration-dependent manner. The concentration causing 50% inhibition (IC50) was 0.1, 0.26, and 65 microM, respectively, resulting in an isomeric potency ratio IC50 (+)/IC50(-) of 650. The dissociation constant KA of the (+/-)- or (+)-methacholine-presynaptic receptor complex was determined after fractional receptor inactivation according to Furchgott & Bursztyn (1967) with phenoxybenzamine or propylbenzilylcholine mustard as irreversible antagonists of muscarinic receptors. KA for (-)-methacholine was estimated according to Mackay (1966). KA of (+)-, (+/-)-, and (-)-methacholine were 2.5, 4 and 440 microM, resulting in an isomeric affinity ratio KA (+)/KA(-) of 180. The discrepancy between the isomeric IC50 ratio and the isomeric KA ratio is explained by a higher intrinsic efficacy of the (+)-enantiomer compared to the (-)-enantiomer. Thus, (+)-methacholine has to occupy fewer receptors to induce a given inhibition of release than its antipode as revealed by a plot of fractional receptor occupancy vs response. The results show that, in the effector system of presynaptic muscarinic inhibition, methacholine enantiomers differ greatly not only in affinity for the receptor, but also to some extent in the efficiency of signal transmission, and both parameters contribute to the high isomeric potency ratio. The activity of the racemate is fully accounted for by the activity of the (+)-enantiomer.

  7. Agonists with supraphysiological efficacy at the muscarinic M2 ACh receptor

    PubMed Central

    Schrage, R; Seemann, WK; Klöckner, J; Dallanoce, C; Racké, K; Kostenis, E; De Amici, M; Holzgrabe, U; Mohr, K

    2013-01-01

    Background and Purpose Artificial agonists may have higher efficacy for receptor activation than the physiological agonist. Until now, such ‘superagonism’ has rarely been reported for GPCRs. Iperoxo is an extremely potent muscarinic receptor agonist. We hypothesized that iperoxo is a ‘superagonist’. Experimental Approach Signalling of iperoxo and newly synthesized structural analogues was compared with that of ACh at label-free M2 muscarinic receptors applying whole cell dynamic mass redistribution, measurement of G-protein activation, evaluation of cell surface agonist binding and computation of operational efficacies. Key Results In CHO-hM2 cells, iperoxo significantly exceeds ACh in Gi/Gs signalling competence. In the orthosteric loss-of-function mutant M2-Y1043.33A, the maximum effect of iperoxo is hardly compromised in contrast to ACh. ‘Superagonism’ is preserved in the physiological cellular context of MRC-5 human lung fibroblasts. Structure–signalling relationships including iperoxo derivatives with either modified positively charged head group or altered tail suggest that ‘superagonism’ of iperoxo is mechanistically based on parallel activation of the receptor protein via two orthosteric interaction points. Conclusion and Implications Supraphysiological agonist efficacy at muscarinic M2 ACh receptors is demonstrated for the first time. In addition, a possible underlying molecular mechanism of GPCR ‘superagonism’ is provided. We suggest that iperoxo-like orthosteric GPCR activation is a new avenue towards a novel class of receptor activators. Linked Article This article is commented on by Langmead and Christopoulos, pp. 353–356 of this issue. To view this commentary visit http://dx.doi.org/10.1111/bph.12142 PMID:23062057

  8. Affinities of pirenzepine for muscarinic cholinergic receptors in membranes isolated from bovine tracheal mucosa and smooth muscle

    SciTech Connect

    Madison, J.M.; Jones, C.A.; Tom-Moy, M.; Brown, J.K.

    1987-03-01

    Muscarinic cholinergic receptors have been classified into subtypes based on their high (M-1 subtype) or low (M-2 subtype) affinities for the nonclassic antagonist pirenzepine, and this classification has important experimental and therapeutic implications. Because muscarinic receptors are abundant in the airways where they mediate several different cellular responses, the goal of this study was to characterize the affinities of pirenzepine for the muscarinic receptors in bovine tracheal mucosa and smooth muscle. After isolating membrane particulates from mucosa and smooth muscle, as well as from bovine cerebral cortex (a known source of M-1 receptors), we used /sup 3/H-quinuclidinyl benzilate to label muscarinic receptors in the particulates and performed competition radioligand binding assays in the presence of either atropine or pirenzepine. Receptors from all 3 tissues (mucosa, smooth muscle, and cerebral cortex) were of a relatively uniform affinity for atropine (range of KI values: 0.8 +/- 0.4 X 10(-9) to 2.4 +/- 1.7 X 10(-9) M), as would be predicted for this classic muscarinic antagonist. By contrast, affinities for pirenzepine differed depending on the tissue. In cerebral cortex, the majority of receptors were of high affinity for pirenzepine (KI = 1.8 +/- 1.4 X 10(-8) M). In both mucosa and smooth muscle, receptors were of low affinity for pirenzepine (Kl = 4.8 +/- 0.4 to 6.9 +/- 3.8 X 10(-7) M). We conclude that muscarinic cholinergic receptors in bovine tracheal mucosa and smooth muscle are predominantly of the M-2 subtype.

  9. L-689,660, a novel cholinomimetic with functional selectivity for M1 and M3 muscarinic receptors.

    PubMed Central

    Hargreaves, R. J.; McKnight, A. T.; Scholey, K.; Newberry, N. R.; Street, L. J.; Hutson, P. H.; Semark, J. E.; Harley, E. A.; Patel, S.; Freedman, S. B.

    1992-01-01

    1. L-689,660, 1-azabicyclo[2.2.2]octane, 3-(6-chloropyrazinyl)maleate, a novel cholinomimetic, demonstrated high affinity binding (pKD (apparent) 7.42) at rat cerebral cortex muscarinic receptors. L-689,660 had a low ratio (34) of pKD (apparent) values for the displacement of binding of the antagonist ([3H]-N-methylscopolamine ([3H]-NMS) compared with the displacement of the agonist [3H]-oxotremorine-M ([3H]-Oxo-M), in rat cerebral cortex. Low NMS/Oxo-M ratios have been shown previously to be a characteristic of compounds that are low efficacy partial agonists with respect to stimulation of phosphatidyl inositol turnover in the cerebral cortex. 2. L-689,660 showed no muscarinic receptor subtype selectivity in radioligand binding assays but showed functional selectivity in pharmacological assays. At M1 muscarinic receptors in the rat superior cervical ganglion, L-689,660 was a potent (pEC50 7.3 +/- 0.2) full agonist in comparison with (+/-)-muscarine. At M3 receptors in the guinea-pig ileum myenteric plexus-longitudinal muscle or in trachea, L-689,660 was again a potent agonist (pEC50 7.5 +/- 0.2 and 7.7 +/- 0.3 respectively) but had a lower maximum response than carbachol. In contrast L-689,660 was an antagonist at M2 receptors in guinea-pig atria (pA2 7.2 (95% confidence limits 7, 7.4)) and at muscarinic autoreceptors in rat hippocampal slices. 3. The putative M1-selective muscarinic agonist, AF102B (cis-2-methylspiro-(1,3-oxathiolane 5,3')-quinuclidine hydrochloride) was found to have a profile similar to L-689,660 but had up to 100 times less affinity in binding and functional assays.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1422595

  10. Expression of the human muscarinic receptor gene m2 in Dictyostelium discoideum

    SciTech Connect

    Voith, G.; Dingermann, T.

    1995-11-01

    We have expressed a functional human muscarinic M2 receptor, under the control of the homologous discoidin I{gamma} promoter, in the cellular slime mold Dictyostelium discoideum. The use of a contact site A leader peptide ensured insertion of the newly synthesized receptor protein into the plasma membrane. Due to the characteristics of the discoidin I{gamma} promoter, the M2 receptor is expressed during late growth and early development. The heterologously expressed M2 receptors show binding characteristics similar to authentic receptors. Membranes as well as whole cells can be used in ligand binding assays. 36 refs., 4 figs.

  11. Muscarinic receptors in rat nasal mucosa are predominantly of the low affinity agonist type.

    PubMed

    Rodrigues de Miranda, J F; Scheres, H M; Salden, H J; Beld, A J; Klaassen, A B; Kuijpers, W

    1985-07-31

    Specific [3H]l-quinuclidinyl benzilate binding to rat nasal mucosa homogenates occurs to a homogeneous class of binding sites with Kd = 60 +/- 2 10(-12) M and Bmax = 8.1 +/- 2 pmol/g tissue. Binding is stereoselectively inhibited by benzetimide enantiomers. Pirenzepine inhibits [3H]l-quinuclidinyl benzilate binding with low affinity (Ki = 5.0 10(-7) M), classifying the binding sites as muscarinic M2-receptors. Methylfurtrethonium and methacholine inhibit [3H]l-quinuclidinyl benzilate binding following an almost sigmoid curve at high concentrations pointing to the presence of mainly low affinity agonist binding sites. PMID:3840092

  12. Selective labeling and localization of the M4 (m4) muscarinic receptor subtype.

    PubMed

    Ferrari-Dileo, G; Waelbroeck, M; Mash, D C; Flynn, D D

    1994-12-01

    We report here a novel strategy for the selective labeling and localization of the M4 (m4) muscarinic receptor subtype, based on the distinct kinetics of the muscarinic antagonists dexetimide and N-methylscopolamine (NMS) and on the selectivity profile of guanylpirenzepine and AF-DX 116 for the m1-m5 muscarinic receptor subtypes expressed in CHO-K1 cells. Incubation with 10 nM dexetimide, a nonselective antagonist, resulted in > 90% occupancy of each of the m1-m5 receptor subtypes. The relatively rapid rates of dexetimide dissociation from the m1, m2, and m4 receptor subtypes (t1/2 values of < 12.5 min) and the slower rates of dexetimide dissociation from the m3 and m5 receptor subtypes (t1/2 values of 65 and 75 min, respectively) favored the labeling of the m1, m2, and m4 receptor subtypes with short incubations with [3H]NMS. Inclusion of 200 nM guanylpirenzepine and 250 nM AF-DX 116 prevented the binding of [3H]NMS to the majority of the m1 and m2 receptor subtypes, respectively, resulting in primary labeling of the m4 receptor subtype. Brief dissociation of the radioligand in the presence of 1 microM atropine improved the ratio of m4 to m2 labeling by selectively removing [3H]NMS from the m2 subtype. Under these conditions, the ratio of [3H]NMS binding to the m4 versus m1, m2, m3, and m5 receptor subtypes was 4:1. In vitro autoradiography combined with these m4-selective labeling conditions demonstrated that the M4 (m4) receptor subtype was localized to the primary visual area (V1, area 17, occipital cortex) and the basal ganglia, a distribution distinct from that demonstrated for the M1 (m1), M2 (m2), and M3 (m3) receptor subtypes. These results demonstrate that a combination of the distinct kinetics of dexetimide and NMS and the receptor subtype selectivity of guanylpirenzepine and AF-DX 116 provides a valuable labeling strategy to examine the distribution and localization of the M4 (m4) muscarinic receptor subtype in brain, peripheral tissues, and cell lines

  13. Expression of the human muscarinic receptor gene m2 in Dictyostelium discoideum.

    PubMed

    Voith, G; Dingermann, T

    1995-11-01

    We have expressed a functional human muscarinic M2 receptor, under the control of the homologous discoidin I gamma promoter, in the cellular slime mold Dictyostelium discoideum. The use of a contact site A leader peptide ensured insertion of the newly synthesized receptor protein into the plasma membrane. Due to the characteristics of the discoidin I gamma promoter, the M2 receptor is expressed during late growth and early development. The heterologously expressed M2 receptors show binding characteristics similar to authentic receptors. Membranes as well as whole cells can be used in ligand binding assays. PMID:9636297