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Sample records for cb1 receptor inhibition

  1. Anandamide inhibits Theiler's virus induced VCAM-1 in brain endothelial cells and reduces leukocyte transmigration in a model of blood brain barrier by activation of CB1 receptors

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    Loría Frida

    2011-08-01

    Full Text Available Abstract Background VCAM-1 represents one of the most important adhesion molecule involved in the transmigration of blood leukocytes across the blood-brain barrier (BBB that is an essential step in the pathogenesis of MS. Several evidences have suggested the potential therapeutic value of cannabinoids (CBs in the treatment of MS and their experimental models. However, the effects of endocannabinoids on VCAM-1 regulation are poorly understood. In the present study we investigated the effects of anandamide (AEA in the regulation of VCAM-1 expression induced by Theiler's virus (TMEV infection of brain endothelial cells using in vitro and in vivo approaches. Methods i in vitro: VCAM-1 was measured by ELISA in supernatants of brain endothelial cells infected with TMEV and subjected to AEA and/or cannabinoid receptors antagonist treatment. To evaluate the functional effect of VCAM-1 modulation we developed a blood brain barrier model based on a system of astrocytes and brain endothelial cells co-culture. ii in vivo: CB1 receptor deficient mice (Cnr1-/- infected with TMEV were treated with the AEA uptake inhibitor UCM-707 for three days. VCAM-1 expression and microglial reactivity were evaluated by immunohistochemistry. Results Anandamide-induced inhibition of VCAM-1 expression in brain endothelial cell cultures was mediated by activation of CB1 receptors. The study of leukocyte transmigration confirmed the functional relevance of VCAM-1 inhibition by AEA. In vivo approaches also showed that the inhibition of AEA uptake reduced the expression of brain VCAM-1 in response to TMEV infection. Although a decreased expression of VCAM-1 by UCM-707 was observed in both, wild type and CB1 receptor deficient mice (Cnr1-/-, the magnitude of VCAM-1 inhibition was significantly higher in the wild type mice. Interestingly, Cnr1-/- mice showed enhanced microglial reactivity and VCAM-1 expression following TMEV infection, indicating that the lack of CB1 receptor

  2. The Cannabinoid Receptor CB1 Modulates the Signaling Properties of the Lysophosphatidylinositol Receptor GPR55*

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    Kargl, Julia; Balenga, Nariman; Parzmair, Gerald P.; Brown, Andrew J.; Heinemann, Akos; Waldhoer, Maria

    2012-01-01

    The G protein-coupled receptor (GPCR) 55 (GPR55) and the cannabinoid receptor 1 (CB1R) are co-expressed in many tissues, predominantly in the central nervous system. Seven transmembrane spanning (7TM) receptors/GPCRs can form homo- and heteromers and initiate distinct signaling pathways. Recently, several synthetic CB1 receptor inverse agonists/antagonists, such as SR141716A, AM251, and AM281, were reported to activate GPR55. Of these, SR141716A was marketed as a promising anti-obesity drug, but was withdrawn from the market because of severe side effects. Here, we tested whether GPR55 and CB1 receptors are capable of (i) forming heteromers and (ii) whether such heteromers could exhibit novel signaling patterns. We show that GPR55 and CB1 receptors alter each others signaling properties in human embryonic kidney (HEK293) cells. We demonstrate that the co-expression of FLAG-CB1 receptors in cells stably expressing HA-GPR55 specifically inhibits GPR55-mediated transcription factor activation, such as nuclear factor of activated T-cells and serum response element, as well as extracellular signal-regulated kinases (ERK1/2) activation. GPR55 and CB1 receptors can form heteromers, but the internalization of both receptors is not affected. In addition, we observe that the presence of GPR55 enhances CB1R-mediated ERK1/2 and nuclear factor of activated T-cell activation. Our data provide the first evidence that GPR55 can form heteromers with another 7TM/GPCR and that this interaction with the CB1 receptor has functional consequences in vitro. The GPR55-CB1R heteromer may play an important physiological and/or pathophysiological role in tissues endogenously co-expressing both receptors. PMID:23161546

  3. The role of CB1 receptors in psychostimulant addiction

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    Wiskerke, J.; Pattij, T.; Schoffelmeer, A.N.M.; de Vries, T.J.

    2008-01-01

    Recent studies have implicated the endocannabinoid (eCB) system in the neuronal mechanisms underlying substance dependence. Here, we review results of studies using cannabinoid receptor subtype 1 (CB1) knockout mice as well as CB1 antagonists to elucidate the role of this neurotransmitter system in

  4. Loss of cannabinoid receptor CB1 induces preterm birth.

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    Haibin Wang

    2008-10-01

    Full Text Available Preterm birth accounting approximate 10% of pregnancies in women is a tremendous social, clinical and economic burden. However, its underlying causes remain largely unknown. Emerging evidence suggests that endocannabinoid signaling via cannabinoid receptor CB1 play critical roles in multiple early pregnancy events in both animals and humans. Since our previous studies demonstrated that loss of CB1 defers the normal implantation window in mice, we surmised that CB1 deficiency would influence parturition events.Exploiting mouse models with targeted deletion of Cnr1, Cnr2 and Ptgs1 encoding CB1, CB2 and cyclooxygenase-1, respectively, we examined consequences of CB1 or CB2 silencing on the onset of parturition. We observed that genetic or pharmacological inactivation of CB1, but not CB2, induced preterm labor in mice. Radioimmunoassay analysis of circulating levels of ovarian steroid hormones revealed that premature birth resulting from CB1 inactivation is correlated with altered progesterone/estrogen ratios prior to parturition. More strikingly, the phenotypic defects of prolonged pregnancy length and parturition failure in mice missing Ptgs1 were corrected by introducing CB1 deficiency into Ptgs1 null mice. In addition, loss of CB1 resulted in aberrant secretions of corticotrophin-releasing hormone and corticosterone during late gestation. The pathophysiological significance of this altered corticotrophin-releasing hormone-driven endocrine activity in the absence of CB1 was evident from our subsequent findings that a selective corticotrophin-releasing hormone antagonist was able to restore the normal parturition timing in Cnr1 deficient mice. In contrast, wild-type females receiving excessive levels of corticosterone induced preterm birth.CB1 deficiency altering normal progesterone and estrogen levels induces preterm birth in mice. This defect is independent of prostaglandins produced by cyclooxygenase-1. Moreover, CB1 inactivation resulted in

  5. Risperidone treatment increases CB1 receptor binding in rat brain

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    Secher, Anna; Husum, Henriette; Holst, Birgitte

    2010-01-01

    , the ghrelin receptor, neuropeptide Y, adiponectin and proopiomelanocortin. We investigated whether the expression of these factors was affected in rats chronically treated with the antipsychotic risperidone. METHODS: Male Sprague-Dawley rats were treated with risperidone (1.0 mg/kg/day) or vehicle (20...... showed that risperidone treatment altered CB(1) receptor binding in the rat brain. Risperidone-induced adiposity and metabolic dysfunction in the clinic may be explained by increased CB(1) receptor density in brain regions involved in appetite and regulation of metabolic function....

  6. Elevation of endogenous anandamide impairs LTP, learning, and memory through CB1 receptor signaling in mice.

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    Basavarajappa, Balapal S; Nagre, Nagaraja N; Xie, Shan; Subbanna, Shivakumar

    2014-07-01

    In rodents, many exogenous and endogenous cannabinoids, such as anandamide (AEA) and 2-arachidonyl glycerol (2-AG), have been shown to play an important role in certain hippocampal memory processes. However, the mechanisms by which endogenous AEA regulate this processes are not well understood. Here the effects of AEA on long-term potentiation (LTP), hippocampal-dependent learning and memory tasks, pERK1/2, pCaMKIV, and pCREB signaling events in both cannabinoid receptor type 1 (CB1R) wild-type (WT) and knockout (KO) mice were assessed following administration of URB597, an inhibitor of the fatty acid amide hydrolase (FAAH). Acute administration of URB597 enhanced AEA levels without affecting the levels of 2-AG or CB1R in the hippocampus and neocortex as compared to vehicle. In hippocampal slices, URB597 impaired LTP in CB1R WT but not in KO littermates. URB597 impaired object recognition, spontaneous alternation and spatial memory in the Y-maze test in CB1R WT mice but not in KO mice. Furthermore, URB597 enhanced ERK phosphorylation in WT without affecting total ERK levels in WT or KO mice. URB597 impaired CaMKIV and CREB phosphorylation in WT but not in KO mice. CB1R KO mice have a lower pCaMKIV/CaMKIV ratio and higher pCREB/CREB ratio as compared to WT littermates. Our results indicate that pharmacologically elevated AEA impair LTP, learning and memory and inhibit CaMKIV and CREB phosphorylation, via the activation of CB1Rs. Collectively, these findings also suggest that pharmacological elevation of AEA beyond normal concentrations is also detrimental for the underlying physiological responses. © 2014 Wiley Periodicals, Inc.

  7. CB1 cannabinoid receptor expression in the striatum: Association with corticostriatal circuits and developmental regulation

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    Vincent eVan Waes

    2012-03-01

    Full Text Available Corticostriatal circuits mediate various aspects of goal-directed behavior and are critically important for basal ganglia-related disorders. Activity in these circuits is regulated by the endocannabinoid system via stimulation of CB1 cannabinoid receptors. CB1 receptors are highly expressed in projection neurons and select interneurons of the striatum, but expression levels vary considerably between different striatal regions (functional domains. We investigated CB1 receptor expression within specific corticostriatal circuits by mapping CB1 mRNA levels in striatal sectors defined by their cortical inputs in rats. We also assessed changes in CB1 expression in the striatum during development. Our results show that CB1 expression is highest in juveniles (P25 and then progressively decreases towards adolescent (P40 and adult (P70 levels. At every age, CB1 receptors are predominantly expressed in sensorimotor striatal sectors, with considerably lower expression in associative and limbic sectors. Moreover, for most corticostriatal circuits there is an inverse relationship between cortical and striatal expression levels. Thus, striatal sectors with high CB1 expression (sensorimotor sectors tend to receive inputs from cortical areas with low expression, while striatal sectors with low expression (associative/limbic sectors receive inputs from cortical regions with higher expression (medial prefrontal cortex. In so far as CB1 mRNA levels reflect receptor function, our findings suggest differential CB1 signaling between different developmental stages and between sensorimotor and associative/limbic circuits. The regional distribution of CB1 receptor expression in the striatum further suggests that, in sensorimotor sectors, CB1 receptors mostly regulate GABA inputs from local axon collaterals of projection neurons, whereas in associative/limbic sectors, CB1 regulation of GABA inputs from interneurons and glutamate inputs may be more important.

  8. Cannabinoid CB1 Receptors Are Localized in Striated Muscle Mitochondria and Regulate Mitochondrial Respiration

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    Juan Mendizabal-Zubiaga

    2016-10-01

    Full Text Available The cannabinoid type 1 (CB1 receptor is widely distributed in the brain and peripheral organs where it regulates cellular functions and metabolism. In the brain, CB1 is mainly localized on presynaptic axon terminals but is also found on mitochondria (mtCB1, where it regulates cellular respiration and energy production. Likewise, CB1 is localized on muscle mitochondria, but very little is known about it. The aim of this study was to further investigate in detail the distribution and functional role of mtCB1 in three different striated muscles. Immunoelectron microscopy for CB1 was used in skeletal muscles (gastrocnemius and rectus abdominis and myocardium from wild-type and CB1-KO mice. Functional assessments were performed in mitochondria purified from the heart of the mice and the mitochondrial oxygen consumption upon application of different acute delta-9-tetrahidrocannabinol (Δ9-THC concentrations (100 nM or 200 nM was monitored. About 26% of the mitochondrial profiles in gastrocnemius, 22% in the rectus abdominis and 17% in the myocardium expressed CB1. Furthermore, the proportion of mtCB1 versus total CB1 immunoparticles was about 60% in the gastrocnemius, 55% in the rectus abdominis and 78% in the myocardium. Importantly, the CB1 immunolabeling pattern disappeared in muscles of CB1-KO mice. Functionally, acute 100 nM or 200 nM THC treatment specifically decreased mitochondria coupled respiration between 12% and 15% in wild-type isolated mitochondria of myocardial muscles but no significant difference was noticed between THC treated and vehicle in mitochondria isolated from CB1-KO heart. Furthermore, gene expression of key enzymes involved in pyruvate synthesis, tricarboxylic acid (TCA cycle and mitochondrial respiratory chain was evaluated in the striated muscle of CB1-WT and CB1-KO. CB1-KO showed an increase in the gene expression of Eno3, Pkm2, and Pdha1, suggesting an increased production of pyruvate. In contrast, no significant

  9. Postnatal Development of CB1 Receptor Expression in Rodent Somatosensory Cortex

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    Deshmukh, Suvarna; Onozuka, Kaori; Bender, Kevin J.; Bender, Vanessa A.; Lutz, Beat; Mackie, Ken; Feldman, Daniel E.

    2007-01-01

    Endocannabinoids are powerful modulators of synaptic transmission that act on presynaptic cannabinoid receptors. Cannabinoid receptor type 1 (CB1) is the dominant receptor in the CNS, and is present in many brain regions, including sensory cortex. To investigate the potential role of CB1 receptors in cortical development, we examined the developmental expression of CB1 in rodent primary somatosensory (barrel) cortex, using immunohistochemistry with a CB1-specific antibody. We found that before postnatal day (P) 6, CB1 receptor staining was present exclusively in the cortical white matter, and that CB1 staining appeared in the grey matter between P6 and P20 in a specific laminar pattern. CB1 staining was confined to axons, and was most prominent in cortical layers 2/3, 5a, and 6. CB1 null (−/−) mice showed altered anatomical barrel maps in layer 4, with enlarged inter-barrel septa, but normal barrel size. These results indicate that CB1 receptors are present in early postnatal development and influence development of sensory maps. PMID:17210229

  10. The CB1 Receptor as an Important Mediator of Hedonic Reward Processing

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    Friemel, Chris M; Zimmer, Andreas; Schneider, Miriam

    2014-01-01

    The endocannabinoid (ECB) system has emerged recently as a key mediator for reward processing. It is well known that cannabinoids affect appetitive learning processes and can induce reinforcing and rewarding effects. However, the involvement of the ECB system in hedonic aspects of reward-related behavior is not completely understood. With the present study, we investigated the modulatory role of the ECB system on hedonic perception, measured by the pleasure attenuated startle (PAS) paradigm for a palatable food reward. Here, a conditioned odor is thought to induce a pleasant affective state that attenuates an aversive reflex—the acoustic startle response. Modulatory effects of the CB1 receptor antagonist/inverse agonist SR1411716 and the cannabinoid agonist WIN 55 212-2 on PAS were examined in rats. PAS was also measured in CB1 receptor knockout (KO) and wild-type (WT) mice. Pharmacological inhibition as well as the absence of CB1 receptors was found to reduce PAS, whereas WIN 55 212-2 administration increased PAS. Finally, presentation of a conditioned reward cue was found to induce striatal FosB/ΔFosB expression in WT mice, but not in KO mice, indicating a reduced stimulation of reward-related brain regions in conditioned KO mice by odor presentation. We here show that in addition to our previous studies in rats, PAS may also serve as a valuable and suitable measure to assess hedonic processing in mice. Our data further indicate that the ECB system, and in particular CB1 receptor signaling, appears to be highly important for the mediation of hedonic aspects of reward processing. PMID:24718372

  11. Comparative effects of chlorpyrifos in wild type and cannabinoid Cb1 receptor knockout mice

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    Baireddy, Praveena; Liu, Jing; Hinsdale, Myron; Pope, Carey

    2011-01-01

    Endocannabinoids (eCBs) modulate neurotransmission by inhibiting the release of a variety of neurotransmitters. The cannabinoid receptor agonist WIN 55.212-2 (WIN) can modulate organophosphorus (OP) anticholinesterase toxicity in rats, presumably by inhibiting acetylcholine (ACh) release. Some OP anticholinesterases also inhibit eCB-degrading enzymes. We studied the effects of the OP insecticide chlorpyrifos (CPF) on cholinergic signs of toxicity, cholinesterase activity and ACh release in tissues from wild type (+/+) and cannabinoid CB1 receptor knockout (−/−) mice. Mice of both genotypes (n = 5–6/treatment group) were challenged with CPF (300 mg/kg, 2 ml/kg in peanut oil, sc) and evaluated for functional and neurochemical changes. Both genotypes exhibited similar cholinergic signs and cholinesterase inhibition (82–95% at 48 h after dosing) in cortex, cerebellum and heart. WIN reduced depolarization-induced ACh release in vitro in hippocampal slices from wild type mice, but had no effect in hippocampal slices from knockouts or in striatal slices from either genotype. Chlorpyrifos oxon (CPO, 100 μM) reduced release in hippocampal slices from both genotypes in vitro, but with a greater reduction in tissues from wild types (21% vs 12%). CPO had no significant in vitro effect on ACh release in striatum. CPF reduced ACh release in hippocampus from both genotypes ex vivo, but reduction was again significantly greater in tissues from wild types (52% vs 36%). In striatum, CPF led to a similar reduction (20–23%) in tissues from both genotypes. Thus, while CB1 deletion in mice had little influence on the expression of acute toxicity following CPF, CPF- or CPO-induced changes in ACh release appeared sensitive to modulation by CB1-mediated eCB signaling in a brain-regional manner. -- Highlights: ► C57Bl/6 mice showed dose-related cholinergic toxicity following subcutaneous chlorpyrifos exposure. ► Wild type and cannabinoid CB1 receptor knockout littermates

  12. Comparative effects of chlorpyrifos in wild type and cannabinoid Cb1 receptor knockout mice

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    Baireddy, Praveena; Liu, Jing; Hinsdale, Myron; Pope, Carey, E-mail: carey.pope@okstate.edu

    2011-11-15

    Endocannabinoids (eCBs) modulate neurotransmission by inhibiting the release of a variety of neurotransmitters. The cannabinoid receptor agonist WIN 55.212-2 (WIN) can modulate organophosphorus (OP) anticholinesterase toxicity in rats, presumably by inhibiting acetylcholine (ACh) release. Some OP anticholinesterases also inhibit eCB-degrading enzymes. We studied the effects of the OP insecticide chlorpyrifos (CPF) on cholinergic signs of toxicity, cholinesterase activity and ACh release in tissues from wild type (+/+) and cannabinoid CB1 receptor knockout (-/-) mice. Mice of both genotypes (n = 5-6/treatment group) were challenged with CPF (300 mg/kg, 2 ml/kg in peanut oil, sc) and evaluated for functional and neurochemical changes. Both genotypes exhibited similar cholinergic signs and cholinesterase inhibition (82-95% at 48 h after dosing) in cortex, cerebellum and heart. WIN reduced depolarization-induced ACh release in vitro in hippocampal slices from wild type mice, but had no effect in hippocampal slices from knockouts or in striatal slices from either genotype. Chlorpyrifos oxon (CPO, 100 {mu}M) reduced release in hippocampal slices from both genotypes in vitro, but with a greater reduction in tissues from wild types (21% vs 12%). CPO had no significant in vitro effect on ACh release in striatum. CPF reduced ACh release in hippocampus from both genotypes ex vivo, but reduction was again significantly greater in tissues from wild types (52% vs 36%). In striatum, CPF led to a similar reduction (20-23%) in tissues from both genotypes. Thus, while CB1 deletion in mice had little influence on the expression of acute toxicity following CPF, CPF- or CPO-induced changes in ACh release appeared sensitive to modulation by CB1-mediated eCB signaling in a brain-regional manner. -- Highlights: Black-Right-Pointing-Pointer C57Bl/6 mice showed dose-related cholinergic toxicity following subcutaneous chlorpyrifos exposure. Black-Right-Pointing-Pointer Wild type and

  13. Endocannabinoid CB1 Receptor Mediated Rises in Ca2+ and Depolarization-Induced Suppression of Inhibition within the Laterodorsal Tegmental Nucleus

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    Soni, Neeraj; Kohlmeier, Kristi Anne

    2016-01-01

    -sensitive, inhibitory postsynaptic currents (IPSCs), which was an action sensitive to presence of a Ca(2+) chelator. Finally, actions of DHPG, a mGlu1R agonist, on IPSC activity were examined and found to result in an AM251- and BAPTA-sensitive inhibition of both the frequency and amplitude of sIPSCs. Taken...

  14. Regulation of hippocampal cannabinoid CB1 receptor actions by adenosine A1 receptors and chronic caffeine administration: implications for the effects of Δ9-tetrahydrocannabinol on spatial memory.

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    Sousa, Vasco C; Assaife-Lopes, Natália; Ribeiro, Joaquim A; Pratt, Judith A; Brett, Ros R; Sebastião, Ana M

    2011-01-01

    The cannabinoid CB(1) receptor-mediated modulation of γ-aminobutyric acid (GABA) release from inhibitory interneurons is important for the integrity of hippocampal-dependent spatial memory. Although adenosine A(1) receptors have a central role in fine-tuning excitatory transmission in the hippocampus, A(1) receptors localized in GABAergic cells do not directly influence GABA release. CB(1) and A(1) receptors are the main targets for the effects of two of the most heavily consumed psychoactive substances worldwide: Δ(9)-tetrahydrocannabinol (THC, a CB(1) receptor agonist) and caffeine (an adenosine receptor antagonist). We first tested the hypothesis that an A(1)-CB(1) interaction influences GABA and glutamate release in the hippocampus. We found that A(1) receptor activation attenuated the CB(1)-mediated inhibition of GABA and glutamate release and this interaction was manifested at the level of G-protein activation. Using in vivo and in vitro approaches, we then investigated the functional implications of the adenosine-cannabinoid interplay that may arise following chronic caffeine consumption. Chronic administration of caffeine in mice (intraperitoneally, 3 mg/kg/day, for 15 days, >12 h before trials) led to an A(1)-mediated enhancement of the CB(1)-dependent acute disruptive effects of THC on a short-term spatial memory task, despite inducing a reduction in cortical and hippocampal CB(1) receptor number and an attenuation of CB(1) coupling with G protein. A(1) receptor levels were increased following chronic caffeine administration. This study shows that A(1) receptors exert a negative modulatory effect on CB(1)-mediated inhibition of GABA and glutamate release, and provides the first evidence of chronic caffeine-induced alterations on the cannabinoid system in the cortex and hippocampus, with functional implications in spatial memory.

  15. Effects of cannabinoid CB1 receptor antagonist rimonabant in consolidation and reconsolidation of methamphetamine reward memory in mice.

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    Yu, Lu-lu; Wang, Xue-yi; Zhao, Mei; Liu, Yu; Li, Yan-qin; Li, Fang-qiong; Wang, Xiaoyi; Xue, Yan-xue; Lu, Lin

    2009-06-01

    Previous studies have shown that cannabinoid CB1 receptors play an important role in specific aspects of learning and memory, yet there has been no systematic study focusing on the involvement of cannabinoid CB1 receptors in methamphetamine-related reward memory. The purpose of this study was to examine whether rimonabant, a cannabinoid CB1 receptor antagonist, would disrupt the consolidation and reconsolidation of methamphetamine-related reward memory, using conditioned place preference paradigm (CPP). Separate groups of male Kunming mice were trained to acquire methamphetamine CPP. Vehicle or rimonabant (1 mg/kg or 3 mg/kg, i.p.) was given at different time points: immediately after each CPP training session (consolidation), 30 min before the reactivation of CPP (retrieval), or immediately after the reactivation of CPP (reconsolidation). Methamphetamine CPP was retested 24 h and 1 and 2 weeks after rimonabant administration. Rimonabant at doses of 1 and 3 mg/kg significantly inhibited the consolidation of methamphetamine CPP. Only high-dose rimonabant (3 mg/kg) disrupted the retrieval and reconsolidation of methamphetamine CPP. Rimonabant had no effect on methamphetamine CPP in the absence of methamphetamine CPP reactivation. Our findings suggest that cannabinoid CB1 receptors play a major role in methamphetamine reward memory, and cannabinoid CB1 receptor antagonists may be a potential pharmacotherapy to manage relapse associated with drug-reward-related memory.

  16. Interactions of CB1 and mGlu5 receptor antagonists in food intake, anxiety and memory models in rats.

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    Varga, Balázs; Kassai, Ferenc; Gyertyán, István

    2012-12-01

    CB(1) receptor antagonists proved to be effective anti-obesity drugs, however, their depressive and anxiogenic effects became also evident. Finding solution to overcome these psychiatric side effects is still in focus of research. Based on the available clinical and preclinical results we hypothesized that the combination of CB(1) and mGlu(5) receptor antagonisms may result in a pharmacological intervention, where the anxiolytic mGlu(5) receptor inhibition may counteract the anxiogenic psychiatric side effects of CB(1) antagonism, while CB(1) antagonism may ameliorate the memory impairing effect of mGlu(5) receptor antagonism. Further, the two components will synergistically interact in blocking food-intake and reducing obesity. For testing the interaction of mGlu(5) and CB(1) receptor antagonism MTEP [3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pridine; SIB-1757, 6-methyl-2-(phenylazo)-3-pyridinol)] (mGlu(5) antagonist) and rimonabant [(5-(4-Chlorophenyl)-1-(2,4-dichloro-phenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide)hydrochloride] (CB(1) antagonist) were used. All experiments were carried out in rats. Effects of the compounds on anxiety were tested in two foot shock induced ultrasonic vocalization paradigms, appetite suppression was assessed in the food intake test, while memory effects were tested in a context conditioned ultrasonic vocalization setup. MTEP abolished the anxiogenic effect of rimonabant, while there was an additive cooperation in suppressing appetite. However, rimonabant did not ameliorate the memory impairing effect of MTEP. By combination of CB(1) and mGluR5 antagonism, anxiety related side effects might be attenuated, appetite suppression maintained, nevertheless, the possible emergence of unwanted memory impairments can overshadow its therapeutic success. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Association of cannabis use during adolescence, prefrontal CB1 receptor signaling and schizophrenia

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    Adriana eCaballero

    2012-05-01

    Full Text Available The cannabinoid receptor 1 (CB1R is the G-protein coupled receptor responsible for the majority of the endocannabinoid signaling in the human brain. It is widely distributed in the limbic system, basal ganglia, and cerebellum, which are areas responsible for cognition, memory, and motor control. Because of this widespread distribution, it is not surprising that drugs that co-opt CB1R have expected behavioral outcomes consistent with dysregulated signaling from these areas (e.g. memory loss, cognitive deficits, etc. In the context of this review, we present evidence for the role of CB1R signaling in the prefrontal cortex (PFC, an area involved in executive functions, with emphasis on the developmental regulation of CB1R signaling in the acquisition of mature PFC function. We further hypothesize how alterations of CB1R signaling specifically during adolescent maturation might confer liability to psychiatric disorders.

  18. Reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in chronic daily cannabis smokers

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    Hirvonen, J; Goodwin, RS; Li, C-T; Terry, GE; Zoghbi, SS; Morse, C; Pike, VW; Volkow, ND; Huestis, MA; Innis, RB

    2011-01-01

    Chronic cannabis (marijuana, hashish) smoking can result in dependence. Rodent studies show reversible downregulation of brain cannabinoid CB1 (cannabinoid receptor type 1) receptors after chronic exposure to cannabis. However, whether downregulation occurs in humans who chronically smoke cannabis is unknown. Here we show, using positron emission tomography imaging, reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in human subjects who chronically smoke ca...

  19. CB1 receptor antagonism increases hippocampal acetylcholine release: site and mechanism of action.

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    Degroot, Aldemar; Köfalvi, Attila; Wade, Mark R; Davis, Richard J; Rodrigues, Ricardo J; Rebola, Nelson; Cunha, Rodrigo A; Nomikos, George G

    2006-10-01

    Evidence indicates that blockade of cannabinoid receptors increases acetylcholine (ACh) release in brain cortical regions. Although it is assumed that this type of effect is mediated through CB1 receptor (CB1R) antagonism, several in vitro functional studies recently have suggested non-CB1R involvement. In addition, neither the precise neuroanatomical site nor the exact mechanisms underlying this effect are known. We thoroughly examined these issues using a combination of systemic and local administration of CB1R antagonists, different methods of in vivo microdialysis, CB1R knockout (KO) mice, tissue measurements of ACh, and immunochemistry. First, we showed that systemic injections of the CB1R antagonists N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR-141716A) and N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251) dose-dependently increased hippocampal ACh efflux. Likewise, local hippocampal, but not septal, infusions of SR141716A or AM251 increased hippocampal ACh release. It is noteworthy that the stimulatory effects of systemically administered CB1R antagonists on hippocampal ACh release were completely abolished in CB1R KO mice. CB1R KO mice had similar basal but higher stress-enhanced hippocampal ACh levels compared with wild-type controls. It is interesting that dopamine D1 receptor antagonism counteracted the stimulatory effect of CB1R blockade on hippocampal ACh levels. Finally, immunohistochemical methods revealed that a high proportion of CB1R-positive nerve terminals were found in hippocampus and confirmed the colocalization of CB1 receptors with cholinergic and dopaminergic nerve terminals. In conclusion, hippocampal ACh release may specifically be controlled through CB1Rs located on both cholinergic and dopaminergic neuronal projections, and CB1R antagonism increases hippocampal ACh release, probably through both a direct

  20. Reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in chronic daily cannabis smokers.

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    Hirvonen, J; Goodwin, R S; Li, C-T; Terry, G E; Zoghbi, S S; Morse, C; Pike, V W; Volkow, N D; Huestis, M A; Innis, R B

    2012-06-01

    Chronic cannabis (marijuana, hashish) smoking can result in dependence. Rodent studies show reversible downregulation of brain cannabinoid CB(1) (cannabinoid receptor type 1) receptors after chronic exposure to cannabis. However, whether downregulation occurs in humans who chronically smoke cannabis is unknown. Here we show, using positron emission tomography imaging, reversible and regionally selective downregulation of brain cannabinoid CB(1) receptors in human subjects who chronically smoke cannabis. Downregulation correlated with years of cannabis smoking and was selective to cortical brain regions. After ∼4 weeks of continuously monitored abstinence from cannabis on a secure research unit, CB(1) receptor density returned to normal levels. This is the first direct demonstration of cortical cannabinoid CB(1) receptor downregulation as a neuroadaptation that may promote cannabis dependence in human brain.

  1. Participation of hypothalamic CB1 receptors in reproductive axis disruption during immune challenge.

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    Surkin, P N; Di Rosso, M E; Correa, F; Elverdin, J C; Genaro, A M; De Laurentiis, A; Fernández-Solari, J

    2017-08-01

    Immune challenge inhibits reproductive function and endocannabinoids (eCB) modulate sexual hormones. However, no studies have been performed to assess whether the eCB system mediates the inhibition of hormones that control reproduction as a result of immune system activation during systemic infections. For that reason, we evaluated the participation of the hypothalamic cannabinoid receptor CB1 on the hypothalamic-pituitary-gonadal (HPG) axis activity in rats submitted to immune challenge. Male adult rats were treated i.c.v. administration with a CB1 antagonist/inverse agonist (AM251) (500 ng/5 μL), followed by an i.p. injection of lipopolysaccharide (LPS) (5 mg/kg) 15 minutes later. Plasmatic, hypothalamic and adenohypophyseal pro-inflammatory cytokines, hormones and neuropeptides were assessed 90 or 180 minutes post-LPS. The plasma concentration of tumour necrosis factor α and adenohypophyseal mRNA expression of Tnfα and Il1β increased 90 and 180 minutes post i.p. administration of LPS. However, cytokine mRNA expression in the hypothalamus increased only 180 minutes post-LPS, suggesting an inflammatory delay in this organ. CB1 receptor blockade with AM251 increased LPS inflammatory effects, particularly in the hypothalamus. LPS also inhibited the HPG axis by decreasing gonadotrophin-releasing hormone hypothalamic content and plasma levels of luteinising hormone and testosterone. These disruptor effects were accompanied by decreased hypothalamic Kiss1 mRNA expression and prostaglandin E2 content, as well as by increased gonadotrophin-inhibitory hormone (Rfrp3) mRNA expression. All these disruptive effects were prevented by the presence of AM251. In summary, our results suggest that, in male rats, eCB mediate immune challenge-inhibitory effects on reproductive axis at least partially via hypothalamic CB1 activation. In addition, this receptor also participates in homeostasis recovery by modulating the inflammatory process taking place after LPS

  2. Peripheral and central CB1 cannabinoid receptors control stress-induced impairment of memory consolidation.

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    Busquets-Garcia, Arnau; Gomis-González, Maria; Srivastava, Raj Kamal; Cutando, Laura; Ortega-Alvaro, Antonio; Ruehle, Sabine; Remmers, Floortje; Bindila, Laura; Bellocchio, Luigi; Marsicano, Giovanni; Lutz, Beat; Maldonado, Rafael; Ozaita, Andrés

    2016-08-30

    Stressful events can generate emotional memories linked to the traumatic incident, but they also can impair the formation of nonemotional memories. Although the impact of stress on emotional memories is well studied, much less is known about the influence of the emotional state on the formation of nonemotional memories. We used the novel object-recognition task as a model of nonemotional memory in mice to investigate the underlying mechanism of the deleterious effect of stress on memory consolidation. Systemic, hippocampal, and peripheral blockade of cannabinoid type-1 (CB1) receptors abolished the stress-induced memory impairment. Genetic deletion and rescue of CB1 receptors in specific cell types revealed that the CB1 receptor population specifically in dopamine β-hydroxylase (DBH)-expressing cells is both necessary and sufficient for stress-induced impairment of memory consolidation, but CB1 receptors present in other neuronal populations are not involved. Strikingly, pharmacological manipulations in mice expressing CB1 receptors exclusively in DBH(+) cells revealed that both hippocampal and peripheral receptors mediate the impact of stress on memory consolidation. Thus, CB1 receptors on adrenergic and noradrenergic cells provide previously unrecognized cross-talk between central and peripheral mechanisms in the stress-dependent regulation of nonemotional memory consolidation, suggesting new potential avenues for the treatment of cognitive aspects on stress-related disorders.

  3. Localization and function of the cannabinoid CB1 receptor in the anterolateral bed nucleus of the stria terminalis.

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    Nagore Puente

    Full Text Available BACKGROUND: The bed nucleus of the stria terminalis (BNST is involved in behaviors related to natural reward, drug addiction and stress. In spite of the emerging role of the endogenous cannabinoid (eCB system in these behaviors, little is known about the anatomy and function of this system in the anterolateral BNST (alBNST. The aim of this study was to provide a detailed morphological characterization of the localization of the cannabinoid 1 (CB1 receptor a necessary step toward a better understanding of the physiological roles of the eCB system in this region of the brain. METHODOLOGY/PRINCIPAL FINDINGS: We have combined anatomical approaches at the confocal and electron microscopy level to ex-vivo electrophysiological techniques. Here, we report that CB1 is localized on presynaptic membranes of about 55% of immunopositive synaptic terminals for the vesicular glutamate transporter 1 (vGluT1, which contain abundant spherical, clear synaptic vesicles and make asymmetrical synapses with alBNST neurons. About 64% of vGluT1 immunonegative synaptic terminals show CB1 immunolabeling. Furthermore, 30% and 35% of presynaptic boutons localize CB1 in alBNST of conditional mutant mice lacking CB1 mainly from GABAergic neurons (GABA-CB1-KO mice and mainly from cortical glutamatergic neurons (Glu-CB1-KO mice, respectively. Extracellular field recordings and whole cell patch clamp in the alBNST rat brain slice preparation revealed that activation of CB1 strongly inhibits excitatory and inhibitory synaptic transmission. CONCLUSIONS/SIGNIFICANCE: This study supports the anterolateral BNST as a potential neuronal substrate of the effects of cannabinoids on stress-related behaviors.

  4. GABAergic and cortical and subcortical glutamatergic axon terminals contain CB1 cannabinoid receptors in the ventromedial nucleus of the hypothalamus.

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    Leire Reguero

    Full Text Available BACKGROUND: Type-1 cannabinoid receptors (CB(1R are enriched in the hypothalamus, particularly in the ventromedial hypothalamic nucleus (VMH that participates in homeostatic and behavioral functions including food intake. Although CB(1R activation modulates excitatory and inhibitory synaptic transmission in the brain, CB(1R contribution to the molecular architecture of the excitatory and inhibitory synaptic terminals in the VMH is not known. Therefore, the aim of this study was to investigate the precise subcellular distribution of CB(1R in the VMH to better understand the modulation exerted by the endocannabinoid system on the complex brain circuitries converging into this nucleus. METHODOLOGY/PRINCIPAL FINDINGS: Light and electron microscopy techniques were used to analyze CB(1R distribution in the VMH of CB(1R-WT, CB(1R-KO and conditional mutant mice bearing a selective deletion of CB(1R in cortical glutamatergic (Glu-CB(1R-KO or GABAergic neurons (GABA-CB(1R-KO. At light microscopy, CB(1R immunolabeling was observed in the VMH of CB(1R-WT and Glu-CB(1R-KO animals, being remarkably reduced in GABA-CB(1R-KO mice. In the electron microscope, CB(1R appeared in membranes of both glutamatergic and GABAergic terminals/preterminals. There was no significant difference in the percentage of CB(1R immunopositive profiles and CB(1R density in terminals making asymmetric or symmetric synapses in CB(1R-WT mice. Furthermore, the proportion of CB(1R immunopositive terminals/preterminals in CB(1R-WT and Glu-CB(1R-KO mice was reduced in GABA-CB(1R-KO mutants. CB(1R density was similar in all animal conditions. Finally, the percentage of CB(1R labeled boutons making asymmetric synapses slightly decreased in Glu-CB(1R-KO mutants relative to CB(1R-WT mice, indicating that CB(1R was distributed in cortical and subcortical excitatory synaptic terminals. CONCLUSIONS/SIGNIFICANCE: Our anatomical results support the idea that the VMH is a relevant hub candidate in

  5. Cannabinoid-1 receptor (CB1R) blockers as medicines: beyond obesity and cardiometabolic disorders to substance abuse/drug addiction with CB1R neutral antagonists.

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    Janero, David R

    2012-03-01

    Addiction to chemical substances with abuse potential presents medical needs largely unsolved by extant therapeutic strategies. Signal transmission through the cannabinoid-1 receptor (CB1R) in the central nervous system (CNS) modulates neurotransmitters/neuronal pathways contributing to the rewarding properties and hedonic effects of certain nondrug stimuli (e.g., food) and many prototypical addictive drugs, promoting excessive intake and its pathological consequences. Typical CB1R antagonists/inverse agonists reduce the rewarding effects and normalize behavioral phenotypes associated with food and abused drugs, but carry an unacceptable adverse-event profile that may reflect, at least partly, their intrinsic ability to alter basal homeostatic CB1R signaling in the CNS and elicit a negative efficacy response. Alternatively, peripherally biased CB1R inverse agonists with limited CNS permeability and putative CB1R neutral antagonists expressing modest (if any) inverse-agonist efficacy are garnering attention for treating obesity and related cardiometabolic complications with a potentially enhanced benefit-to-risk profile. This mini-review calls attention to the proposition that CB1R neutral antagonists offer attractive opportunities for pharmacotherapeutic exploitation in the substance abuse/drug addiction space, whereas the restricted CNS accessibility of peripherally biased CB1R inverse agonists circumscribes their therapeutic utility for this indication. The unique preclinical pharmacology, efficacy profiles, and reduced adverse-event risk of CB1R neutral antagonists make them worthy of translational study for their potential therapeutic application beyond obesity/cardiometabolic disease to include substance-abuse/drug-addiction disorders.

  6. Controlled-Deactivation CB1 Receptor Ligands as a Novel Strategy to Lower Intraocular Pressure

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    Sally Miller

    2018-05-01

    Full Text Available Nearly half a century has passed since the demonstration that cannabis and its chief psychoactive component Δ9-THC lowers intraocular pressure (IOP. Elevated IOP remains the chief hallmark and therapeutic target for glaucoma, a condition that places millions at risk of blindness. It is likely that Δ9-THC exerts much of its IOP-lowering effects via the activation of CB1 cannabinoid receptors. However, the initial promise of CB1 as a target for treating glaucoma has not thus far translated into a credible therapeutic strategy. We have recently shown that blocking monoacylglycerol lipase (MAGL, an enzyme that breaks the endocannabinoid 2-arachidonoyl glycerol (2-AG, substantially lowers IOP. Another strategy is to develop cannabinoid CB1 receptor agonists that are optimized for topical application to the eye. Recently we have reported on a controlled-deactivation approach where the “soft” drug concept of enzymatic deactivation was combined with a “depot effect” that is commonly observed with Δ9-THC and other lipophilic cannabinoids. This approach allowed us to develop novel cannabinoids with a predictable duration of action and is particularly attractive for the design of CB1 activators for ophthalmic use with limited or no psychoactive effects. We have tested a novel class of compounds using a combination of electrophysiology in autaptic hippocampal neurons, a well-characterized model of endogenous cannabinoid signaling, and measurements of IOP in a mouse model. We now report that AM7410 is a reasonably potent and efficacious agonist at CB1 in neurons and that it substantially (30% lowers IOP for as long as 5 h after a single topical treatment. This effect is absent in CB1 knockout mice. Our results indicate that the direct targeting of CB1 receptors with controlled-deactivation ligands is a viable approach to lower IOP in a murine model and merits further study in other model systems.

  7. Paranoid schizophrenia is characterized by increased CB1 receptor binding in the dorsolateral prefrontal cortex.

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    Dalton, Victoria S; Long, Leonora E; Weickert, Cyndi Shannon; Zavitsanou, Katerina

    2011-07-01

    A number of studies suggest a dysregulation of the endogenous cannabinoid system in schizophrenia (SCZ). In the present study, we examined cannabinoid CB(1) receptor (CB(1)R) binding and mRNA expression in the dorsolateral prefrontal cortex (DLPFC) (Brodmann's area 46) of SCZ patients and controls, post-mortem. Receptor density was investigated using autoradiography with the CB(1)R ligand [(3)H] CP 55,940 and CB(1)R mRNA expression was measured using quantitative RT-PCR in a cohort of 16 patients with paranoid SCZ, 21 patients with non-paranoid SCZ and 37 controls matched for age, post-mortem interval and pH. All cases were obtained from the University of Sydney Tissue Resource Centre. Results were analyzed using one-way analysis of variance (ANOVA) and post hoc Bonferroni tests and with analysis of covariance (ANCOVA) to control for demographic factors that would potentially influence CB(1)R expression. There was a main effect of diagnosis on [(3)H] CP 55,940 binding quantified across all layers of the DLPFC (F(2,71) = 3.740, p = 0.029). Post hoc tests indicated that this main effect was due to patients with paranoid SCZ having 22% higher levels of CB(1)R binding compared with the control group. When ANCOVA was employed, this effect was strengthened (F(2,67) = 6.048, p = 0.004) with paranoid SCZ patients differing significantly from the control (p = 0.004) and from the non-paranoid group (p = 0.016). In contrast, no significant differences were observed in mRNA expression between the different disease subtypes and the control group. Our findings confirm the existence of a CB(1)R dysregulation in SCZ and underline the need for further investigation of the role of this receptor particularly in those diagnosed with paranoid SCZ.

  8. CB1 receptor-mediated signaling underlies the hippocampal synaptic, learning, and memory deficits following treatment with JWH-081, a new component of spice/K2 preparations.

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    Basavarajappa, Balapal S; Subbanna, Shivakumar

    2014-02-01

    Recently, synthetic cannabinoids have been sprayed onto plant material, which is subsequently packaged and sold as "Spice" or "K2" to mimic the effects of marijuana. A recent report identified several synthetic additives in samples of "Spice/K2", including JWH-081, a synthetic ligand for the cannabinoid receptor 1 (CB1). The deleterious effects of JWH-081 on brain function are not known, particularly on CB1 signaling, synaptic plasticity, learning and memory. Here, we evaluated the effects of JWH-081 on pCaMKIV, pCREB, and pERK1/2 signaling events followed by long-term potentiation (LTP), hippocampal-dependent learning and memory tasks using CB1 receptor wild-type (WT) and knockout (KO) mice. Acute administration of JWH-081 impaired CaMKIV phosphorylation in a dose-dependent manner, whereas inhibition of CREB phosphorylation in CB1 receptor WT mice was observed only at higher dose of JWH-081 (1.25 mg/kg). JWH-081 at higher dose impaired CaMKIV and CREB phosphorylation in a time-dependent manner in CB1 receptor WT mice but not in KO mice and failed to alter ERK1/2 phosphorylation. In addition, SR treated or CB1 receptor KO mice have a lower pCaMKIV/CaMKIV ratio and higher pCREB/CREB ratio compared with vehicle or WT littermates. In hippocampal slices, JWH-081 impaired LTP in CB1 receptor WT but not in KO littermates. Furthermore, JWH-081 at higher dose impaired object recognition, spontaneous alternation and spatial memory on the Y-maze in CB1 receptor WT mice but not in KO mice. Collectively our findings suggest that deleterious effects of JWH-081 on hippocampal function involves CB1 receptor mediated impairments in CaMKIV and CREB phosphorylation, LTP, learning and memory in mice. © 2013 Wiley Periodicals, Inc.

  9. Modulation of Network Oscillatory Activity and GABAergic Synaptic Transmission by CB1 Cannabinoid Receptors in the Rat Medial Entorhinal Cortex

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    Nicola H. Morgan

    2008-01-01

    Full Text Available Cannabinoids modulate inhibitory GABAergic neurotransmission in many brain regions. Within the temporal lobe, cannabinoid receptors are highly expressed, and are located presynaptically at inhibitory terminals. Here, we have explored the role of type-1 cannabinoid receptors (CB1Rs at the level of inhibitory synaptic currents and field-recorded network oscillations. We report that arachidonylcyclopropylamide (ACPA; 10 M, an agonist at CB1R, inhibits GABAergic synaptic transmission onto both superficial and deep medial entorhinal (mEC neurones, but this has little effect on network oscillations in beta/gamma frequency bands. By contrast, the CB1R antagonist/inverse agonist LY320135 (500 nM, increased GABAergic synaptic activity and beta/gamma oscillatory activity in superficial mEC, was suppressed, whilst that in deep mEC was enhanced. These data indicate that cannabinoid-mediated effects on inhibitory synaptic activity may be constitutively active in vitro, and that modulation of CB1R activation using inverse agonists unmasks complex effects of CBR function on network activity.

  10. Developmental and visual input-dependent regulation of the CB1 cannabinoid receptor in the mouse visual cortex.

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    Taisuke Yoneda

    Full Text Available The mammalian visual system exhibits significant experience-induced plasticity in the early postnatal period. While physiological studies have revealed the contribution of the CB1 cannabinoid receptor (CB1 to developmental plasticity in the primary visual cortex (V1, it remains unknown whether the expression and localization of CB1 is regulated during development or by visual experience. To explore a possible role of the endocannabinoid system in visual cortical plasticity, we examined the expression of CB1 in the visual cortex of mice. We found intense CB1 immunoreactivity in layers II/III and VI. CB1 mainly localized at vesicular GABA transporter-positive inhibitory nerve terminals. The amount of CB1 protein increased throughout development, and the specific laminar pattern of CB1 appeared at P20 and remained until adulthood. Dark rearing from birth to P30 decreased the amount of CB1 protein in V1 and altered the synaptic localization of CB1 in the deep layer. Dark rearing until P50, however, did not influence the expression of CB1. Brief monocular deprivation for 2 days upregulated the localization of CB1 at inhibitory nerve terminals in the deep layer. Taken together, the expression and the localization of CB1 are developmentally regulated, and both parameters are influenced by visual experience.

  11. Targeting the endocannabinoid/CB1 receptor system for treating obesity in Prader–Willi syndrome

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    Ibrahim Knani

    2016-12-01

    Full Text Available Objective: Extreme obesity is a core phenotypic feature of Prader–Willi syndrome (PWS. Among numerous metabolic regulators, the endocannabinoid (eCB system is critically involved in controlling feeding, body weight, and energy metabolism, and a globally acting cannabinoid-1 receptor (CB1R blockade reverses obesity both in animals and humans. The first-in-class CB1R antagonist rimonabant proved effective in inducing weight loss in adults with PWS. However, it is no longer available for clinical use because of its centrally mediated, neuropsychiatric, adverse effects. Methods: We studied eCB ‘tone’ in individuals with PWS and in the Magel2-null mouse model that recapitulates the major metabolic phenotypes of PWS and determined the efficacy of a peripherally restricted CB1R antagonist, JD5037 in treating obesity in these mice. Results: Individuals with PWS had elevated circulating levels of 2-arachidonoylglycerol and its endogenous precursor and breakdown ligand, arachidonic acid. Increased hypothalamic eCB ‘tone’, manifested by increased eCBs and upregulated CB1R, was associated with increased fat mass, reduced energy expenditure, and decreased voluntary activity in Magel2-null mice. Daily chronic treatment of obese Magel2-null mice and their littermate wild-type controls with JD5037 (3 mg/kg/d for 28 days reduced body weight, reversed hyperphagia, and improved metabolic parameters related to their obese phenotype. Conclusions: Dysregulation of the eCB/CB1R system may contribute to hyperphagia and obesity in Magel2-null mice and in individuals with PWS. Our results demonstrate that treatment with peripherally restricted CB1R antagonists may be an effective strategy for the management of severe obesity in PWS. Author Video: Author Video Watch what authors say about their articles Keywords: Endocannabinoids, PWS, Magel2, Peripheral CB1 blockade, Metabolic syndrome

  12. Role of CB1 cannabinoid receptors on GABAergic neurons in brain aging.

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    Albayram, Onder; Alferink, Judith; Pitsch, Julika; Piyanova, Anastasia; Neitzert, Kim; Poppensieker, Karola; Mauer, Daniela; Michel, Kerstin; Legler, Anne; Becker, Albert; Monory, Krisztina; Lutz, Beat; Zimmer, Andreas; Bilkei-Gorzo, Andras

    2011-07-05

    Brain aging is associated with cognitive decline that is accompanied by progressive neuroinflammatory changes. The endocannabinoid system (ECS) is involved in the regulation of glial activity and influences the progression of age-related learning and memory deficits. Mice lacking the Cnr1 gene (Cnr1(-/-)), which encodes the cannabinoid receptor 1 (CB1), showed an accelerated age-dependent deficit in spatial learning accompanied by a loss of principal neurons in the hippocampus. The age-dependent decrease in neuronal numbers in Cnr1(-/-) mice was not related to decreased neurogenesis or to epileptic seizures. However, enhanced neuroinflammation characterized by an increased density of astrocytes and activated microglia as well as an enhanced expression of the inflammatory cytokine IL-6 during aging was present in the hippocampus of Cnr1(-/-) mice. The ongoing process of pyramidal cell degeneration and neuroinflammation can exacerbate each other and both contribute to the cognitive deficits. Deletion of CB1 receptors from the forebrain GABAergic, but not from the glutamatergic neurons, led to a similar neuronal loss and increased neuroinflammation in the hippocampus as observed in animals lacking CB1 receptors in all cells. Our results suggest that CB1 receptor activity on hippocampal GABAergic neurons protects against age-dependent cognitive decline by reducing pyramidal cell degeneration and neuroinflammation.

  13. A key agonist-induced conformational change in the cannabinoid receptor CB1 is blocked by the allosteric ligand Org 27569.

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    Fay, Jonathan F; Farrens, David L

    2012-09-28

    Allosteric ligands that modulate how G protein-coupled receptors respond to traditional orthosteric drugs are an exciting and rapidly expanding field of pharmacology. An allosteric ligand for the cannabinoid receptor CB1, Org 27569, exhibits an intriguing effect; it increases agonist binding, yet blocks agonist-induced CB1 signaling. Here we explored the mechanism behind this behavior, using a site-directed fluorescence labeling approach. Our results show that Org 27569 blocks conformational changes in CB1 that accompany G protein binding and/or activation, and thus inhibit formation of a fully active CB1 structure. The underlying mechanism behind this behavior is that simultaneous binding of Org 27569 produces a unique agonist-bound conformation, one that may resemble an intermediate structure formed on the pathway to full receptor activation.

  14. Cannabinoid Receptors CB1 and CB2 Modulate the Electroretinographic Waves in Vervet Monkeys

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    Joseph Bouskila

    2016-01-01

    Full Text Available The expression patterns of the cannabinoid receptor type 1 (CB1R and the cannabinoid receptor type 2 (CB2R are well documented in rodents and primates. In vervet monkeys, CB1R is present in the retinal neurons (photoreceptors, horizontal cells, bipolar cells, amacrine cells, and ganglion cells and CB2R is exclusively found in the retinal glia (Müller cells. However, the role of these cannabinoid receptors in normal primate retinal function remains elusive. Using full-field electroretinography in adult vervet monkeys, we recorded changes in neural activity following the blockade of CB1R and CB2R by the intravitreal administration of their antagonists (AM251 and AM630, resp. in photopic and scotopic conditions. Our results show that AM251 increases the photopic a-wave amplitude at high flash intensities, whereas AM630 increases the amplitude of both the photopic a- and b-waves. In scotopic conditions, both blockers increased the b-wave amplitude but did not change the a-wave amplitude. These findings suggest an important role of CB1R and CB2R in primate retinal function.

  15. Elevated Brain Cannabinoid CB1 Receptor Availability in Posttraumatic Stress Disorder: A Positron Emission Tomography Study

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    Neumeister, Alexander; Normandin, Marc D.; Pietrzak, Robert H.; Piomelli, Daniele; Zheng, Ming-Qiang; Gujarro-Anton, Ana; Potenza, Marc N.; Bailey, Christopher R.; Lin, Shu-fei; Najafzadeh, Soheila; Ropchan, Jim; Henry, Shannan; Corsi-Travali, Stefani; Carson, Richard E.; Huang, Yiyun

    2013-01-01

    Endocannabinoids and their attending cannabinoid type 1 receptor (CB1) have been implicated in animal models of posttraumatic stress disorder (PTSD). However, their specific role has not been studied in people with PTSD. Herein, we present an in vivo imaging study using positron emission tomography (PET) and the CB1-selective radioligand [11C]OMAR in individuals with PTSD, and healthy controls with lifetime histories of trauma (trauma controls [TC]) and those without such histories (healthy controls [HC]). Untreated individuals with PTSD (N=25) with non-combat trauma histories, and TC (N=12) and HC (N=23) participated in a magnetic resonance (MR) imaging scan and a resting PET scan with the CB1 receptor antagonist radiotracer [11C]OMAR, which measures volume of distribution (VT) linearly related to CB1 receptor availability. Peripheral levels of anandamide, 2-arachidonoylglycerol (2-AG), oleoylethanolamide (OEA), palmitoylethanolamide (PEA), and cortisol were also assessed. In the PTSD group, relative to the HC and TC groups, we found elevated brain-wide [11C]OMAR VT values (F(2,53)=7.96, p=.001; 19.5% and 14.5% higher, respectively) which were most pronounced in women (F(1,53)=5.52, p=.023). Anandamide concentrations were reduced in the PTSD relative to the TC (53.1% lower) and HC (58.2% lower) groups. Cortisol levels were lower in the PTSD and TC groups relative to the HC group. Three biomarkers examined collectively—OMAR VT, anandamide, and cortisol—correctly classified nearly 85% of PTSD cases. These results suggest that abnormal CB1 receptor-mediated anandamide signaling is implicated in the etiology of PTSD, and provide a promising neurobiological model to develop novel, evidence-based pharmacotherapies for this disorder. PMID:23670490

  16. Beyond the CB1 Receptor: Is Cannabidiol the Answer for Disorders of Motivation?

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    Zlebnik, Natalie E.; Cheer, Joseph F.

    2016-01-01

    The Cannabis sativa plant has been used to treat various physiological and psychiatric conditions for millennia. Current research is focused on isolating potentially therapeutic chemical constituents from the plant for use in the treatment of many central nervous system disorders. Of particular interest is the primary nonpsychoactive constituent cannabidiol (CBD). Unlike Δ9-tetrahydrocannabinol (THC), CBD does not act through the cannabinoid type 1 (CB1) receptor but has many other receptor t...

  17. Effects of caffeine on striatal neurotransmission: focus on cannabinoid CB1 receptors.

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    Rossi, Silvia; De Chiara, Valentina; Musella, Alessandra; Mataluni, Giorgia; Sacchetti, Lucia; Siracusano, Alberto; Bernardi, Giorgio; Usiello, Alessandro; Centonze, Diego

    2010-04-01

    Caffeine is the most commonly self-administered psychoactive substance worldwide. At usual doses, the effects of caffeine on vigilance, attention, mood and arousal largely depend on the modulation of central adenosine receptors. The present review article describes the action of caffeine within the striatum, to provide a possible molecular mechanism at the basis of the psychomotor and reinforcing properties of this pharmacological agent. The striatum is in fact a subcortical area involved in sensorimotor, cognitive, and emotional processes, and recent experimental findings showed that chronic caffeine consumption enhances the sensitivity of striatal GABAergic synapses to the stimulation of cannabinoid CB1 receptors. The endocannabinoid system is involved in the psychoactive effects of many compounds, and adenosine A2A receptors (the main receptor target of caffeine) elicit a permissive effect towards CB1 receptors, thus suggesting that A2A-CB1 receptor interaction plays a major role in the generation and maintenance of caffeine reinforcing behavior. Aim of this review is to describe the effects of caffeine on striatal neurotransmission with special reference to the modulation of the endocannabinoid system.

  18. Activation of the sympathetic nervous system mediates hypophagic and anxiety-like effects of CB1 receptor blockade

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    Bellocchio, Luigi; Soria-Gómez, Edgar; Quarta, Carmelo; Metna-Laurent, Mathilde; Cardinal, Pierre; Binder, Elke; Cannich, Astrid; Delamarre, Anna; Häring, Martin; Martín-Fontecha, Mar; Vega, David; Leste-Lasserre, Thierry; Bartsch, Dusan; Monory, Krisztina; Lutz, Beat; Chaouloff, Francis; Pagotto, Uberto; Guzman, Manuel; Cota, Daniela; Marsicano, Giovanni

    2013-01-01

    Complex interactions between periphery and the brain regulate food intake in mammals. Cannabinoid type-1 (CB1) receptor antagonists are potent hypophagic agents, but the sites where this acute action is exerted and the underlying mechanisms are not fully elucidated. To dissect the mechanisms underlying the hypophagic effect of CB1 receptor blockade, we combined the acute injection of the CB1 receptor antagonist rimonabant with the use of conditional CB1-knockout mice, as well as with pharmacological modulation of different central and peripheral circuits. Fasting/refeeding experiments revealed that CB1 receptor signaling in many specific brain neurons is dispensable for the acute hypophagic effects of rimonabant. CB1 receptor antagonist-induced hypophagia was fully abolished by peripheral blockade of β-adrenergic transmission, suggesting that this effect is mediated by increased activity of the sympathetic nervous system. Consistently, we found that rimonabant increases gastrointestinal metabolism via increased peripheral β-adrenergic receptor signaling in peripheral organs, including the gastrointestinal tract. Blockade of both visceral afferents and glutamatergic transmission in the nucleus tractus solitarii abolished rimonabant-induced hypophagia. Importantly, these mechanisms were specifically triggered by lipid-deprivation, revealing a nutrient-specific component acutely regulated by CB1 receptor blockade. Finally, peripheral blockade of sympathetic neurotransmission also blunted central effects of CB1 receptor blockade, such as fear responses and anxiety-like behaviors. These data demonstrate that, independently of their site of origin, important effects of CB1 receptor blockade are expressed via activation of peripheral sympathetic activity. Thus, CB1 receptors modulate bidirectional circuits between the periphery and the brain to regulate feeding and other behaviors. PMID:23487769

  19. Cannabinoid CB1 receptor antagonist rimonabant disrupts nicotine reward-associated memory in rats.

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    Fang, Qin; Li, Fang-Qiong; Li, Yan-Qin; Xue, Yan-Xue; He, Ying-Ying; Liu, Jian-Feng; Lu, Lin; Wang, Ji-Shi

    2011-10-01

    Exposure to cues previously associated with drug intake leads to relapse by activating previously acquired memories. Based on previous findings, in which cannabinoid CB(1) receptors were found to be critically involved in specific aspects of learning and memory, we investigated the role of CB(1) receptors in nicotine reward memory using a rat conditioned place preference (CPP) model. In Experiment 1, rats were trained for CPP with alternating injections of nicotine (0.5mg/kg, s.c.) and saline to acquire the nicotine-conditioned memory. To examine the effects of rimonabant on the reconsolidation of nicotine reward memory, rats were administered rimonabant (0, 0.3, and 3.0mg/kg, i.p.) immediately after reexposure to the drug-paired context. In Experiment 2, rats were trained for CPP similarly to Experiment 1. To examine the effects of rimonabant on the reinstatement of nicotine reward memory, rimonabant (0, 0.3, and 3.0mg/kg, i.p.) was administered before the test of nicotine-induced CPP reinstatement. In Experiment 3, to evaluate whether rimonabant itself produces a reward memory, rats were trained for CPP with alternating injections of different doses of rimonabant (0, 0.3, and 3.0mg/kg) and saline. Rimonabant at a dose of 3.0mg/kg significantly disrupted the reconsolidation of nicotine memory and significantly blocked the reinstatement of nicotine-induced CPP. However, rimonabant itself did not produce CPP. These findings provide clear evidence that CB(1) receptors play a role in nicotine reward memory, suggesting that CB(1) receptor antagonists may be a potential target for managing nicotine addiction. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Acute cannabinoids impair working memory through astroglial CB1 receptor modulation of hippocampal LTD.

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    Han, Jing; Kesner, Philip; Metna-Laurent, Mathilde; Duan, Tingting; Xu, Lin; Georges, Francois; Koehl, Muriel; Abrous, Djoher Nora; Mendizabal-Zubiaga, Juan; Grandes, Pedro; Liu, Qingsong; Bai, Guang; Wang, Wei; Xiong, Lize; Ren, Wei; Marsicano, Giovanni; Zhang, Xia

    2012-03-02

    Impairment of working memory is one of the most important deleterious effects of marijuana intoxication in humans, but its underlying mechanisms are presently unknown. Here, we demonstrate that the impairment of spatial working memory (SWM) and in vivo long-term depression (LTD) of synaptic strength at hippocampal CA3-CA1 synapses, induced by an acute exposure of exogenous cannabinoids, is fully abolished in conditional mutant mice lacking type-1 cannabinoid receptors (CB(1)R) in brain astroglial cells but is conserved in mice lacking CB(1)R in glutamatergic or GABAergic neurons. Blockade of neuronal glutamate N-methyl-D-aspartate receptors (NMDAR) and of synaptic trafficking of glutamate α-amino-3-hydroxy-5-methyl-isoxazole propionic acid receptors (AMPAR) also abolishes cannabinoid effects on SWM and LTD induction and expression. We conclude that the impairment of working memory by marijuana and cannabinoids is due to the activation of astroglial CB(1)R and is associated with astroglia-dependent hippocampal LTD in vivo. Copyright © 2012 Elsevier Inc. All rights reserved.

  1. Anti-inflammatory activity of topical THC in DNFB-mediated mouse allergic contact dermatitis independent of CB1 and CB2 receptors.

    Science.gov (United States)

    Gaffal, E; Cron, M; Glodde, N; Tüting, T

    2013-08-01

    ∆(9) -Tetrahydrocannabinol (THC), the active constituent of Cannabis sativa, exerts its biological effects in part through the G-protein-coupled CB1 and CB2 receptors, which were initially discovered in brain and spleen tissue, respectively. However, THC also has CB1/2 receptor-independent effects. Because of its immune-inhibitory potential, THC and related cannabinoids are being considered for the treatment of inflammatory skin diseases. Here we investigated the mechanism of the anti-inflammatory activity of THC and the role of CB1 and CB2 receptors. We evaluated the impact of topically applied THC on DNFB-mediated allergic contact dermatitis in wild-type and CB1/2 receptor-deficient mice. We performed immunohistochemical analyses for infiltrating immune cells and studied the influence of THC on the interaction between T cells, keratinocytes and myeloid immune cells in vitro. Topical THC application effectively decreased contact allergic ear swelling and myeloid immune cell infiltration not only in wild-type but also in CB1/2 receptor-deficient mice. We found that THC (1) inhibited the production of IFNγ by T cells, (2) decreased the production of CCL2 and of IFNγ-induced CCL8 and CXL10 by epidermal keratinocytes and (3) thereby limited the recruitment of myeloid immune cells in vitro in a CB1/2 receptor-independent manner. Topically applied THC can effectively attenuate contact allergic inflammation by decreasing keratinocyte-derived pro-inflammatory mediators that orchestrate myeloid immune cell infiltration independent of CB1/2 receptors. This has important implications for the future development of strategies to harness cannabinoids for the treatment of inflammatory skin diseases. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  2. Mice expressing a “hyper-sensitive” form of the CB1 cannabinoid receptor (CB1) show modestly enhanced alcohol preference and consumption

    Science.gov (United States)

    Gonek, Maciej; Zee, Michael L.; Farnsworth, Jill C.; Amin, Randa A.; Andrews, Mary-Jeanette; Davis, Brian J.; Mackie, Ken; Morgan, Daniel J.

    2017-01-01

    We recently characterized S426A/S430A mutant mice expressing a desensitization-resistant form of the CB1 receptor. These mice display an enhanced response to endocannabinoids and ∆9-THC. In this study, S426A/S430A mutants were used as a novel model to test whether ethanol consumption, morphine dependence, and reward for these drugs are potentiated in mice with a “hyper-sensitive” form of CB1. Using an unlimited-access, two-bottle choice, voluntary drinking paradigm, S426A/S430A mutants exhibit modestly increased intake and preference for low (6%) but not higher concentrations of ethanol. S426A/S430A mutants and wild-type mice show similar taste preference for sucrose and quinine, exhibit normal sensitivity to the hypothermic and ataxic effects of ethanol, and have normal blood ethanol concentrations following administration of ethanol. S426A/S430A mutants develop robust conditioned place preference for ethanol (2 g/kg), morphine (10 mg/kg), and cocaine (10 mg/kg), demonstrating that drug reward is not changed in S426A/S430A mutants. Precipitated morphine withdrawal is also unchanged in opioid-dependent S426A/S430A mutant mice. Although ethanol consumption is modestly changed by enhanced CB1 signaling, reward, tolerance, and acute sensitivity to ethanol and morphine are normal in this model. PMID:28426670

  3. Mice expressing a "hyper-sensitive" form of the CB1 cannabinoid receptor (CB1 show modestly enhanced alcohol preference and consumption.

    Directory of Open Access Journals (Sweden)

    David J Marcus

    Full Text Available We recently characterized S426A/S430A mutant mice expressing a desensitization-resistant form of the CB1 receptor. These mice display an enhanced response to endocannabinoids and ∆9-THC. In this study, S426A/S430A mutants were used as a novel model to test whether ethanol consumption, morphine dependence, and reward for these drugs are potentiated in mice with a "hyper-sensitive" form of CB1. Using an unlimited-access, two-bottle choice, voluntary drinking paradigm, S426A/S430A mutants exhibit modestly increased intake and preference for low (6% but not higher concentrations of ethanol. S426A/S430A mutants and wild-type mice show similar taste preference for sucrose and quinine, exhibit normal sensitivity to the hypothermic and ataxic effects of ethanol, and have normal blood ethanol concentrations following administration of ethanol. S426A/S430A mutants develop robust conditioned place preference for ethanol (2 g/kg, morphine (10 mg/kg, and cocaine (10 mg/kg, demonstrating that drug reward is not changed in S426A/S430A mutants. Precipitated morphine withdrawal is also unchanged in opioid-dependent S426A/S430A mutant mice. Although ethanol consumption is modestly changed by enhanced CB1 signaling, reward, tolerance, and acute sensitivity to ethanol and morphine are normal in this model.

  4. The central cannabinoid CB1 receptor is required for diet-induced obesity and rimonabant's antiobesity effects in mice.

    Science.gov (United States)

    Pang, Zhen; Wu, Nancy N; Zhao, Weiguang; Chain, David C; Schaffer, Erica; Zhang, Xin; Yamdagni, Preeti; Palejwala, Vaseem A; Fan, Chunpeng; Favara, Sarah G; Dressler, Holly M; Economides, Kyriakos D; Weinstock, Daniel; Cavallo, Jean S; Naimi, Souad; Galzin, Anne-Marie; Guillot, Etienne; Pruniaux, Marie-Pierre; Tocci, Michael J; Polites, H Greg

    2011-10-01

    Cannabinoid receptor CB1 is expressed abundantly in the brain and presumably in the peripheral tissues responsible for energy metabolism. It is unclear if the antiobesity effects of rimonabant, a CB1 antagonist, are mediated through the central or the peripheral CB1 receptors. To address this question, we generated transgenic mice with central nervous system (CNS)-specific knockdown (KD) of CB1, by expressing an artificial microRNA (AMIR) under the control of the neuronal Thy1.2 promoter. In the mutant mice, CB1 expression was reduced in the brain and spinal cord, whereas no change was observed in the superior cervical ganglia (SCG), sympathetic trunk, enteric nervous system, and pancreatic ganglia. In contrast to the neuronal tissues, CB1 was undetectable in the brown adipose tissue (BAT) or the liver. Consistent with the selective loss of central CB1, agonist-induced hypothermia was attenuated in the mutant mice, but the agonist-induced delay of gastrointestinal transit (GIT), a primarily peripheral nervous system-mediated effect, was not. Compared to wild-type (WT) littermates, the mutant mice displayed reduced body weight (BW), adiposity, and feeding efficiency, and when fed a high-fat diet (HFD), showed decreased plasma insulin, leptin, cholesterol, and triglyceride levels, and elevated adiponectin levels. Furthermore, the therapeutic effects of rimonabant on food intake (FI), BW, and serum parameters were markedly reduced and correlated with the degree of CB1 KD. Thus, KD of CB1 in the CNS recapitulates the metabolic phenotype of CB1 knockout (KO) mice and diminishes rimonabant's efficacy, indicating that blockade of central CB1 is required for rimonabant's antiobesity actions.

  5. Molecular basis of cannabinoid CB1 receptor coupling to the G protein heterotrimer Gαiβγ: identification of key CB1 contacts with the C-terminal helix α5 of Gαi.

    Science.gov (United States)

    Shim, Joong-Youn; Ahn, Kwang H; Kendall, Debra A

    2013-11-08

    The cannabinoid (CB1) receptor is a member of the rhodopsin-like G protein-coupled receptor superfamily. The human CB1 receptor, which is among the most expressed receptors in the brain, has been implicated in several disease states, including drug addiction, anxiety, depression, obesity, and chronic pain. Different classes of CB1 agonists evoke signaling pathways through the activation of specific subtypes of G proteins. The molecular basis of CB1 receptor coupling to its cognate G protein is unknown. As a first step toward understanding CB1 receptor-mediated G protein signaling, we have constructed a ternary complex structural model of the CB1 receptor and Gi heterotrimer (CB1-Gi), guided by the x-ray structure of β2-adrenergic receptor (β2AR) in complex with Gs (β2AR-Gs), through 824-ns duration molecular dynamics simulations in a fully hydrated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine bilayer environment. We identified a group of residues at the juxtamembrane regions of the intracellular loops 2 and 3 (IC2 and IC3) of the CB1 receptor, including Ile-218(3.54), Tyr-224(IC2), Asp-338(6.30), Arg-340(6.32), Leu-341(6.33), and Thr-344(6.36), as potential key contacts with the extreme C-terminal helix α5 of Gαi. Ala mutations of these residues at the receptor-Gi interface resulted in little G protein coupling activity, consistent with the present model of the CB1-Gi complex, which suggests tight interactions between CB1 and the extreme C-terminal helix α5 of Gαi. The model also suggests that unique conformational changes in the extreme C-terminal helix α5 of Gα play a crucial role in the receptor-mediated G protein activation.

  6. Cannabinoids Modulate Neuronal Activity and Cancer by CB1 and CB2 Receptor-Independent Mechanisms

    Directory of Open Access Journals (Sweden)

    Ken Soderstrom

    2017-10-01

    Full Text Available Cannabinoids include the active constituents of Cannabis or are molecules that mimic the structure and/or function of these Cannabis-derived molecules. Cannabinoids produce many of their cellular and organ system effects by interacting with the well-characterized CB1 and CB2 receptors. However, it has become clear that not all effects of cannabinoid drugs are attributable to their interaction with CB1 and CB2 receptors. Evidence now demonstrates that cannabinoid agents produce effects by modulating activity of the entire array of cellular macromolecules targeted by other drug classes, including: other receptor types; ion channels; transporters; enzymes, and protein- and non-protein cellular structures. This review summarizes evidence for these interactions in the CNS and in cancer, and is organized according to the cellular targets involved. The CNS represents a well-studied area and cancer is emerging in terms of understanding mechanisms by which cannabinoids modulate their activity. Considering the CNS and cancer together allow identification of non-cannabinoid receptor targets that are shared and divergent in both systems. This comparative approach allows the identified targets to be compared and contrasted, suggesting potential new areas of investigation. It also provides insight into the diverse sources of efficacy employed by this interesting class of drugs. Obtaining a comprehensive understanding of the diverse mechanisms of cannabinoid action may lead to the design and development of therapeutic agents with greater efficacy and specificity for their cellular targets.

  7. Neurophysiological evidence for the presence of cannabinoid CB1 receptors in the laterodorsal tegmental nucleus

    DEFF Research Database (Denmark)

    Soni, Neeraj; Satpathy, Shankha; Kohlmeier, Kristi Anne

    2014-01-01

    Marijuana, which acts within the endocannabinoid (eCB) system as an agonist of the cannabinoid type 1 receptor (CB1R), exhibits addictive properties and has powerful actions on the state of arousal of an organism. The laterodorsal tegmental nucleus (LDT), as a component of the reticular activating...... the firing frequency and synaptic activity of neurons in this nucleus. Therefore, endogenous eCB transmission could play a role in processes involving the LDT, such as cortical activation and motivated behaviours and, further, behavioural actions of marijuana are probably mediated, in part, via cellular...

  8. Cannabinoid receptor CB1 mediates baseline and activity-induced survival of new neurons in adult hippocampal neurogenesis

    Directory of Open Access Journals (Sweden)

    Müller Anke

    2010-06-01

    Full Text Available Abstract Background Adult neurogenesis is a particular example of brain plasticity that is partially modulated by the endocannabinoid system. Whereas the impact of synthetic cannabinoids on the neuronal progenitor cells has been described, there has been lack of information about the action of plant-derived extracts on neurogenesis. Therefore we here focused on the effects of Δ9-tetrahydrocannabinol (THC and Cannabidiol (CBD fed to female C57Bl/6 and Nestin-GFP-reporter mice on proliferation and maturation of neuronal progenitor cells and spatial learning performance. In addition we used cannabinoid receptor 1 (CB1 deficient mice and treatment with CB1 antagonist AM251 in Nestin-GFP-reporter mice to investigate the role of the CB1 receptor in adult neurogenesis in detail. Results THC and CBD differed in their effects on spatial learning and adult neurogenesis. CBD did not impair learning but increased adult neurogenesis, whereas THC reduced learning without affecting adult neurogenesis. We found the neurogenic effect of CBD to be dependent on the CB1 receptor, which is expressed over the whole dentate gyrus. Similarly, the neurogenic effect of environmental enrichment and voluntary wheel running depends on the presence of the CB1 receptor. We found that in the absence of CB1 receptors, cell proliferation was increased and neuronal differentiation reduced, which could be related to CB1 receptor mediated signaling in Doublecortin (DCX-expressing intermediate progenitor cells. Conclusion CB1 affected the stages of adult neurogenesis that involve intermediate highly proliferative progenitor cells and the survival and maturation of new neurons. The pro-neurogenic effects of CBD might explain some of the positive therapeutic features of CBD-based compounds.

  9. Spatial distribution of cannabinoid receptor type 1 (CB1 in normal canine central and peripheral nervous system.

    Directory of Open Access Journals (Sweden)

    Jessica Freundt-Revilla

    Full Text Available The endocannabinoid system is a regulatory pathway consisting of two main types of cannabinoid receptors (CB1 and CB2 and their endogenous ligands, the endocannabinoids. The CB1 receptor is highly expressed in the central and peripheral nervous systems (PNS in mammalians and is involved in neuromodulatory functions. Since endocannabinoids were shown to be elevated in cerebrospinal fluid of epileptic dogs, knowledge about the species specific CB receptor expression in the nervous system is required. Therefore, we assessed the spatial distribution of CB1 receptors in the normal canine CNS and PNS. Immunohistochemistry of several regions of the brain, spinal cord and peripheral nerves from a healthy four-week-old puppy, three six-month-old dogs, and one ten-year-old dog revealed strong dot-like immunoreactivity in the neuropil of the cerebral cortex, Cornu Ammonis (CA and dentate gyrus of the hippocampus, midbrain, cerebellum, medulla oblongata and grey matter of the spinal cord. Dense CB1 expression was found in fibres of the globus pallidus and substantia nigra surrounding immunonegative neurons. Astrocytes were constantly positive in all examined regions. CB1 labelled neurons and satellite cells of the dorsal root ganglia, and myelinating Schwann cells in the PNS. These results demonstrate for the first time the spatial distribution of CB1 receptors in the healthy canine CNS and PNS. These results can be used as a basis for further studies aiming to elucidate the physiological consequences of this particular anatomical and cellular distribution.

  10. Beyond the CB1 Receptor: Is Cannabidiol the Answer for Disorders of Motivation?

    Science.gov (United States)

    Zlebnik, Natalie E; Cheer, Joseph F

    2016-07-08

    The Cannabis sativa plant has been used to treat various physiological and psychiatric conditions for millennia. Current research is focused on isolating potentially therapeutic chemical constituents from the plant for use in the treatment of many central nervous system disorders. Of particular interest is the primary nonpsychoactive constituent cannabidiol (CBD). Unlike Δ(9)-tetrahydrocannabinol (THC), CBD does not act through the cannabinoid type 1 (CB1) receptor but has many other receptor targets that may play a role in psychiatric disorders. Here we review preclinical and clinical data outlining the therapeutic efficacy of CBD for the treatment of motivational disorders such as drug addiction, anxiety, and depression. Across studies, findings suggest promising treatment effects and potentially overlapping mechanisms of action for CBD in these disorders and indicate the need for further systematic investigation of the viability of CBD as a psychiatric pharmacotherapy.

  11. Expression and function of cannabinoid receptors CB1 and CB2 and their cognate cannabinoid ligands in murine embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Shuxian Jiang

    2007-07-01

    Full Text Available Characterization of intrinsic and extrinsic factors regulating the self-renewal/division and differentiation of stem cells is crucial in determining embryonic stem (ES cell fate. ES cells differentiate into multiple hematopoietic lineages during embryoid body (EB formation in vitro, which provides an experimental platform to define the molecular mechanisms controlling germ layer fate determination and tissue formation.The cannabinoid receptor type 1 (CB1 and cannabinoid receptor type 2 (CB2 are members of the G-protein coupled receptor (GPCR family, that are activated by endogenous ligands, the endocannabinoids. CB1 receptor expression is abundant in brain while CB2 receptors are mostly expressed in hematopoietic cells. However, the expression and the precise roles of CB1 and CB2 and their cognate ligands in ES cells are not known. We observed significant induction of CB1 and CB2 cannabinoid receptors during the hematopoietic differentiation of murine ES (mES-derived embryoid bodies. Furthermore, mES cells as well as ES-derived embryoid bodies at days 7 and 14, expressed endocannabinoids, the ligands for both CB1 and CB2. The CB1 and CB2 antagonists (AM251 and AM630, respectively induced mES cell death, strongly suggesting that endocannabinoids are involved in the survival of mES cells. Treatment of mES cells with the exogenous cannabinoid ligand Delta(9-THC resulted in the increased hematopoietic differentiation of mES cells, while addition of AM251 or AM630 blocked embryoid body formation derived from the mES cells. In addition, cannabinoid agonists induced the chemotaxis of ES-derived embryoid bodies, which was specifically inhibited by the CB1 and CB2 antagonists.This work has not been addressed previously and yields new information on the function of cannabinoid receptors, CB1 and CB2, as components of a novel pathway regulating murine ES cell differentiation. This study provides insights into cannabinoid system involvement in ES cell

  12. Integrated cannabinoid CB1 receptor transmission within the amygdala-prefrontal cortical pathway modulates neuronal plasticity and emotional memory encoding.

    Science.gov (United States)

    Tan, Huibing; Lauzon, Nicole M; Bishop, Stephanie F; Bechard, Melanie A; Laviolette, Steven R

    2010-06-01

    The cannabinoid CB1 receptor system is functionally involved in the processing and encoding of emotionally salient sensory information, learning and memory. The CB1 receptor is found in high concentrations in brain structures that are critical for emotional processing, including the basolateral amygdala (BLA) and the medial prefrontal cortex (mPFC). In addition, synaptic plasticity in the form of long-term potentiation (LTP) within the BLA > mPFC pathway is an established correlate of exposure to emotionally salient events. We performed a series of in vivo LTP studies by applying tetanic stimulation to the BLA combined with recordings of local field potentials within prelimbic cortical (PLC) region of the rat mPFC. Systemic pretreatment with AM-251 dose dependently blocked LTP along the BLA-PLC pathway and also the behavioral acquisition of conditioned fear memories. We next performed a series of microinfusion experiments wherein CB1 receptor transmission within the BLA > PLC circuit was pharmacologically blocked. Asymmetrical, interhemispheric blockade of CB1 receptor transmission along the BLA > PLC pathway prevented the acquisition of emotionally salient associative memory. Our results indicate that coordinated CB1 receptor transmission within the BLA > PLC pathway is critically involved in the encoding of emotional fear memories and modulates neural plasticity related to the encoding of emotionally salient associative learning.

  13. Mapping CB1 cannabinoid receptors with [3H]OMAR in the Flinders rodent model of depression

    DEFF Research Database (Denmark)

    Nahimi, A.; Gjedde, A.; Wong, D. F.

    2012-01-01

    not significantly different. Conclusions: Although changes in CB1 receptor expression have been demonstrated in human suicide victims with depression and in animal models of depression, the present maps of [3H]OMAR binding revealed no difference between FSL and FRL rats. We used a single concentration of [3H......Background: The endocannabinoid system regulates cognitive and emotional processes and pathology of this system is implicated in psychiatric disorders, including depression and schizophrenia. The precise role of the endocannabinoid system in psychiatric disorders remains unclear, but changes...... in expression of CB1 receptors and subsequent altered modulation of monoamines is suggested in depression (Esteban & Garcia-Sevilla, 2011). CB1 receptor agonists, such as WIN55,212-2 and CP55,940 regulate synthesis and release of monoamines and are suggested as a novel therapy in the treatment of depression...

  14. Chronic ethanol exposure decreases CB1 receptor function at GABAergic synapses in the rat central amygdala

    DEFF Research Database (Denmark)

    Varodayan, Florence P.; Soni, Neeraj; Bajo, Michal

    2016-01-01

    release, and GABAergic dysregulation in the central nucleus of the amygdala (CeA) is critical in the transition to alcohol dependence. We investigated possible disruptions in CB1 signaling of rat CeA GABAergic transmission following intermittent ethanol exposure. In the CeA of alcohol-naive rats, CB1...

  15. Phencyclidine-Induced Social Withdrawal Results from Deficient Stimulation of Cannabinoid CB1 Receptors: Implications for Schizophrenia

    Science.gov (United States)

    Seillier, Alexandre; Martinez, Alex A; Giuffrida, Andrea

    2013-01-01

    The neuronal mechanisms underlying social withdrawal, one of the core negative symptoms of schizophrenia, are not well understood. Recent studies suggest an involvement of the endocannabinoid system in the pathophysiology of schizophrenia and, in particular, of negative symptoms. We used biochemical, pharmacological, and behavioral approaches to investigate the role played by the endocannabinoid system in social withdrawal induced by sub-chronic administration of phencyclidine (PCP). Pharmacological enhancement of endocannabinoid levels via systemic administration of URB597, an inhibitor of endocannabinoid degradation, reversed social withdrawal in PCP-treated rats via stimulation of CB1 receptors, but reduced social interaction in control animals through activation of a cannabinoid/vanilloid-sensitive receptor. In addition, the potent CB agonist CP55,940 reversed PCP-induced social withdrawal in a CB1-dependent manner, whereas pharmacological blockade of CB1 receptors by either AM251 or SR141716 reduced the time spent in social interaction in control animals. PCP-induced social withdrawal was accompanied by a decrease of anandamide (AEA) levels in the amygdala and prefrontal cortex, and these deficits were reversed by URB597. As CB1 receptors are predominantly expressed on GABAergic interneurons containing the anxiogenic peptide cholecystokinin (CCK), we also examined whether the PCP-induced social withdrawal resulted from deficient CB1-mediated modulation of CCK transmission. The selective CCK2 antagonist LY225910 blocked both PCP- and AM251-induced social withdrawal, but not URB597 effect in control rats. Taken together, these findings indicate that AEA-mediated activation of CB1 receptors is crucial for social interaction, and that PCP-induced social withdrawal results from deficient endocannabinoid transmission. PMID:23563893

  16. Singular Location and Signaling Profile of Adenosine A2A-Cannabinoid CB1 Receptor Heteromers in the Dorsal Striatum.

    Science.gov (United States)

    Moreno, Estefanía; Chiarlone, Anna; Medrano, Mireia; Puigdellívol, Mar; Bibic, Lucka; Howell, Lesley A; Resel, Eva; Puente, Nagore; Casarejos, María J; Perucho, Juan; Botta, Joaquín; Suelves, Nuria; Ciruela, Francisco; Ginés, Silvia; Galve-Roperh, Ismael; Casadó, Vicent; Grandes, Pedro; Lutz, Beat; Monory, Krisztina; Canela, Enric I; Lluís, Carmen; McCormick, Peter J; Guzmán, Manuel

    2018-04-01

    The dorsal striatum is a key node for many neurobiological processes such as motor activity, cognitive functions, and affective processes. The proper functioning of striatal neurons relies critically on metabotropic receptors. Specifically, the main adenosine and endocannabinoid receptors present in the striatum, ie, adenosine A 2A receptor (A 2A R) and cannabinoid CB 1 receptor (CB 1 R), are of pivotal importance in the control of neuronal excitability. Facilitatory and inhibitory functional interactions between striatal A 2A R and CB 1 R have been reported, and evidence supports that this cross-talk may rely, at least in part, on the formation of A 2A R-CB 1 R heteromeric complexes. However, the specific location and properties of these heteromers have remained largely unknown. Here, by using techniques that allowed a precise visualization of the heteromers in situ in combination with sophisticated genetically modified animal models, together with biochemical and pharmacological approaches, we provide a high-resolution expression map and a detailed functional characterization of A 2A R-CB 1 R heteromers in the dorsal striatum. Specifically, our data unveil that the A 2A R-CB 1 R heteromer (i) is essentially absent from corticostriatal projections and striatonigral neurons, and, instead, is largely present in striatopallidal neurons, (ii) displays a striking G protein-coupled signaling profile, where co-stimulation of both receptors leads to strongly reduced downstream signaling, and (iii) undergoes an unprecedented dysfunction in Huntington's disease, an archetypal disease that affects striatal neurons. Altogether, our findings may open a new conceptual framework to understand the role of coordinated adenosine-endocannabinoid signaling in the indirect striatal pathway, which may be relevant in motor function and neurodegenerative diseases.

  17. Regulation of Hippocampal Cannabinoid CB1 Receptor Actions by Adenosine A1 Receptors and Chronic Caffeine Administration: Implications for the Effects of Δ9-Tetrahydrocannabinol on Spatial Memory

    OpenAIRE

    Sousa, Vasco C; Assaife-Lopes, Natália; Ribeiro, Joaquim A; Pratt, Judith A; Brett, Ros R; Sebastião, Ana M

    2010-01-01

    Abstract The cannabinoid CB1 receptor-mediated modulation of ?-aminobutyric acid (GABA) release from inhibitory interneurons is important for the integrity of hippocampal-dependent spatial memory. Although adenosine A1 receptors have a central role in fine-tuning excitatory transmission in the hippocampus, A1 receptors localized in GABAergic cells do not directly influence GABA release. CB1 and A1 receptors are the main targets for the effects of two of the most heavily consumed ps...

  18. Clinical Significance of Cannabinoid Receptors CB1 and CB2 Expression in Human Malignant and Benign Thyroid Lesions

    Directory of Open Access Journals (Sweden)

    Eleftheria Lakiotaki

    2015-01-01

    Full Text Available The endocannabinoid system is comprised of cannabinoid receptors (CB1 and CB2, their endogenous ligands (endocannabinoids, and proteins responsible for their metabolism participate in many different functions indispensable to homeostatic regulation in several tissues, exerting also antitumorigenic effects. The present study aimed to evaluate the clinical significance of CB1 and CB2 expression in human benign and malignant thyroid lesions. CB1 and CB2 proteins’ expression was assessed immunohistochemically on paraffin-embedded thyroid tissues obtained from 87 patients with benign (n=43 and malignant (n=44 lesions and was statistically analyzed with clinicopathological parameters, follicular cells’ proliferative capacity, and risk of recurrence rate estimated according to the American Thyroid Association (ATA staging system. Enhanced CB1 and CB2 expression was significantly more frequently observed in malignant compared to benign thyroid lesions (p=0.0010 and p=0.0005, resp.. Enhanced CB1 and CB2 expression was also significantly more frequently observed in papillary carcinomas compared to hyperplastic nodules (p=0.0097 and p=0.0110, resp.. In malignant thyroid lesions, elevated CB2 expression was significantly associated with the presence of lymph node metastases (p=0.0301. Enhanced CB2 expression was also more frequently observed in malignant thyroid cases with presence of capsular (p=0.1165, lymphatic (p=0.1989, and vascular invasion (p=0.0555, as well as in those with increased risk of recurrence rate (p=0.1165, at a nonsignificant level though, whereas CB1 expression was not associated with any of the clinicopathological parameters examined. Our data suggest that CB receptors may be involved in malignant thyroid transformation and especially CB2 receptor could serve as useful biomarker and potential therapeutic target in thyroid neoplasia.

  19. Cannabinoid Type 1 Receptor (CB1) Ligands with Therapeutic Potential for Withdrawal Syndrome in Chemical Dependents of Cannabis sativa.

    Science.gov (United States)

    Ferreira, Jaderson V; Chaves, Gisele A; Marino, Bianca L B; Sousa, Kessia P A; Souza, Lucilene R; Brito, Maiara F B; Teixeira, Hueldem R C; da Silva, Carlos H T P; Santos, Cleydson B R; Hage-Melim, Lorane I S

    2017-08-22

    Cannabis sativa withdrawal syndrome is characterized mainly by psychological symptoms. By using computational tools, the aim of this study was to propose drug candidates for treating withdrawal syndrome based on the natural ligands of the cannabinoid type 1 receptor (CB1). One compound in particular, 2-n-butyl-5-n-pentylbenzene-1,3-diol (ZINC1730183, also known as stemphol), showed positive predictions as a human CB1 ligand and for facile synthetic accessibility. Therefore, ZINC1730183 is a favorable candidate scaffold for further research into pharmacotherapeutic alternatives to treat C. sativa withdrawal syndrome. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. CB1 and CB2 receptors are novel molecular targets for Tamoxifen and 4OH-Tamoxifen

    Energy Technology Data Exchange (ETDEWEB)

    Prather, Paul L. [Department of Pharmacology and Toxicology, College of Medicine, University of Arkansas for Medical Sciences, 4301 W. Markham, Little Rock, AR 72205 (United States); FrancisDevaraj, FeAna; Dates, Centdrika R.; Greer, Aleksandra K.; Bratton, Stacie M. [Department of Biochemistry and Molecular Biology, College of Medicine, University of Arkansas for Medical Sciences, 4301 W. Markham, Little Rock, AR 72205 (United States); Ford, Benjamin M.; Franks, Lirit N. [Department of Pharmacology and Toxicology, College of Medicine, University of Arkansas for Medical Sciences, 4301 W. Markham, Little Rock, AR 72205 (United States); Radominska-Pandya, Anna, E-mail: RadominskaAnna@uams.edu [Department of Biochemistry and Molecular Biology, College of Medicine, University of Arkansas for Medical Sciences, 4301 W. Markham, Little Rock, AR 72205 (United States)

    2013-11-15

    Highlights: •Tamoxifen produces cytotoxicity via estrogen-receptor (ER) independent mechanisms. •Tamoxifen binds to CB1 and CB2 cannabinoid receptors and acts as an inverse agonist. •CB1 and CB2 receptors are novel molecular targets for Tamoxifen. •ER-independent effects for Tamoxifen may be mediated via CB1 and/or CB2 receptors. -- Abstract: Tamoxifen (Tam) is classified as a selective estrogen receptor modulator (SERM) and is used for treatment of patients with ER-positive breast cancer. However, it has been shown that Tam and its cytochrome P450-generated metabolite 4-hydroxy-Tam (4OH-Tam) also exhibit cytotoxic effects in ER-negative breast cancer cells. These observations suggest that Tam and 4OH-Tam can produce cytotoxicity via estrogen receptor (ER)-independent mechanism(s) of action. The molecular targets responsible for the ER-independent effects of Tam and its derivatives are poorly understood. Interestingly, similar to Tam and 4OH-Tam, cannabinoids have also been shown to exhibit anti-proliferative and apoptotic effects in ER-negative breast cancer cells, and estrogen can regulate expression levels of cannabinoid receptors (CBRs). Therefore, this study investigated whether CBRs might serve as novel molecular targets for Tam and 4OH-Tam. We report that both compounds bind to CB1 and CB2Rs with moderate affinity (0.9–3 μM). Furthermore, Tam and 4OH-Tam exhibit inverse activity at CB1 and CB2Rs in membrane preparations, reducing basal G-protein activity. Tam and 4OH-Tam also act as CB1/CB2R-inverse agonists to regulate the downstream intracellular effector adenylyl cyclase in intact cells, producing concentration-dependent increases in intracellular cAMP. These results suggest that CBRs are molecular targets for Tam and 4OH-Tam and may contribute to the ER-independent cytotoxic effects reported for these drugs. Importantly, these findings also indicate that Tam and 4OH-Tam might be used as structural scaffolds for development of novel

  1. CB1 and CB2 receptors are novel molecular targets for Tamoxifen and 4OH-Tamoxifen

    International Nuclear Information System (INIS)

    Prather, Paul L.; FrancisDevaraj, FeAna; Dates, Centdrika R.; Greer, Aleksandra K.; Bratton, Stacie M.; Ford, Benjamin M.; Franks, Lirit N.; Radominska-Pandya, Anna

    2013-01-01

    Highlights: •Tamoxifen produces cytotoxicity via estrogen-receptor (ER) independent mechanisms. •Tamoxifen binds to CB1 and CB2 cannabinoid receptors and acts as an inverse agonist. •CB1 and CB2 receptors are novel molecular targets for Tamoxifen. •ER-independent effects for Tamoxifen may be mediated via CB1 and/or CB2 receptors. -- Abstract: Tamoxifen (Tam) is classified as a selective estrogen receptor modulator (SERM) and is used for treatment of patients with ER-positive breast cancer. However, it has been shown that Tam and its cytochrome P450-generated metabolite 4-hydroxy-Tam (4OH-Tam) also exhibit cytotoxic effects in ER-negative breast cancer cells. These observations suggest that Tam and 4OH-Tam can produce cytotoxicity via estrogen receptor (ER)-independent mechanism(s) of action. The molecular targets responsible for the ER-independent effects of Tam and its derivatives are poorly understood. Interestingly, similar to Tam and 4OH-Tam, cannabinoids have also been shown to exhibit anti-proliferative and apoptotic effects in ER-negative breast cancer cells, and estrogen can regulate expression levels of cannabinoid receptors (CBRs). Therefore, this study investigated whether CBRs might serve as novel molecular targets for Tam and 4OH-Tam. We report that both compounds bind to CB1 and CB2Rs with moderate affinity (0.9–3 μM). Furthermore, Tam and 4OH-Tam exhibit inverse activity at CB1 and CB2Rs in membrane preparations, reducing basal G-protein activity. Tam and 4OH-Tam also act as CB1/CB2R-inverse agonists to regulate the downstream intracellular effector adenylyl cyclase in intact cells, producing concentration-dependent increases in intracellular cAMP. These results suggest that CBRs are molecular targets for Tam and 4OH-Tam and may contribute to the ER-independent cytotoxic effects reported for these drugs. Importantly, these findings also indicate that Tam and 4OH-Tam might be used as structural scaffolds for development of novel

  2. The cannabinoid receptor CB1 contributes to the development of ectopic lesions in a mouse model of endometriosis.

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    Sanchez, Ana-Maria; Quattrone, Federica; Pannese, Maria; Ulisse, Adele; Candiani, Massimo; Diaz-Alonso, Javier; Velasco, Guillermo; Panina-Bordignon, Paola

    2017-01-01

    Does signaling via the cannabinoid (CB 1 ) receptor play a role in the pathogenesis of endometriosis in a mouse model? Mice treated with a CB 1 agonist developed larger ectopic lesions, while less severe lesions developed in the absence of functional CB 1 expression. The expression of components of the endocannabinoid system has been demonstrated in both mouse and human uteri. CB 1 receptors are expressed in human epithelial and stromal cell lines derived from eutopic endometrium and deep infiltrating endometriosis nodules. This was a randomized study in a mouse model of endometriosis. In a first set of experiments, mice with endometriosis were treated with the CB 1 receptor agonist methanandamide (MET) (5 mg/kg, n = 20) on Days 1-5 and 8-12. In a second set of experiments, endometriosis development was evaluated in CB 1 -/- mice and in their wild-type (WT) littermates. Endometriosis-like lesions were induced in Balb/c and C57/Bl6 mice. Two weeks after disease induction, the lesions were counted, measured and either included for immunohistochemistry analysis or frozen for gene expression profiling by semi-quantitative real-time PCR. To limit the role of chance, the experiments were conducted under standardized laboratory conditions with appropriate controls. The lesion total volume was significantly higher in MET-treated compared with vehicle-treated mice (P endometriosis in a mouse model. However, the relative contribution of the CB 1 -mediated signaling pathways active in inflammatory, uterine and peritoneal cells remains to be ascertained. Since the study was performed in a mouse model, the significance of the findings in the human system warrants further investigation. Clarifying the function and regulation of CB 1 and its molecular interactions with endogenous ligands, and how endocannabinoids levels are regulated in women with endometriosis, represent critical areas of research for the potential development of a novel medical treatment of the disease. A

  3. The CB1 receptor antagonist AM251 impairs reconsolidation of pavlovian fear memory in the rat basolateral amygdala.

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    Ratano, Patrizia; Everitt, Barry J; Milton, Amy L

    2014-10-01

    We have investigated the requirement for signaling at CB1 receptors in the reconsolidation of a previously consolidated auditory fear memory, by infusing the CB1 receptor antagonist AM251, or the FAAH inhibitor URB597, directly into the basolateral amygdala (BLA) in conjunction with memory reactivation. AM251 disrupted memory restabilization, but only when administered after reactivation. URB597 produced a small, transient enhancement of memory restabilization when administered after reactivation. The amnestic effect of AM251 was rescued by coadministration of the GABAA receptor antagonist bicuculline at reactivation, indicating that the disruption of reconsolidation was mediated by altered GABAergic transmission in the BLA. These data show that the endocannabinoid system in the BLA is an important modulator of fear memory reconsolidation and that its effects on memory are mediated by an interaction with the GABAergic system. Thus, targeting the endocannabinoid system may have therapeutic potential to reduce the impact of maladaptive memories in neuropsychiatric disorders such as posttraumatic stress disorder.

  4. Candidate PET radioligands for cannabinoid CB1 receptors: [18F]AM5144 and related pyrazole compounds

    International Nuclear Information System (INIS)

    Li Zizhong; Gifford, Andrew; Liu Qian; Thotapally, Rajesh; Ding Yushin; Makriyannis, Alexandros; Gatley, S. John

    2005-01-01

    Introduction: The mammalian brain contains abundant G protein-coupled cannabinoid CB 1 receptors that respond to Δ 9 -tetrahydrocannabinol, the active ingredient of cannabis. The availability of a positron emission tomography (PET) radioligand would facilitate studies of the addictive and medicinal properties of compounds that bind to this receptor. Among the known classes of ligands for CB 1 receptors, the pyrazoles are attractive targets for radiopharmaceutical development because they are antagonists and are generally less lipophilic than the other classes. Methods: A convenient high-yield synthesis of N-(4-[ 18 F]fluorophenyl)-5-(4-bromophenyl)-1-(2,4-dichlorophenyl)- 1H-pyrazole-3-carboxamide (AM5144) was devised by coupling the appropriate pyrazole-3-carboxyl chloride compound with 4-[ 18 F]fluoroaniline. The labeled precursor was synthesized from 1-[ 18 F]fluoro-4-nitrobenzene in 60% radiochemical yield for 10 min using an improved procedure involving sodium borohydride reduction with cobalt chloride catalysis. The product was purified by HPLC to give a specific activity >400 mCi/μmol and a radiochemical purity >95%, and a PET study was conducted in a baboon. Results: Although the regional uptake of AM5144 in baboon brain was consistent with binding to cannabinoid CB 1 receptors, absolute uptake at 1 receptor ligands is not surprising because of their high lipophilicity as compared with most brain PET radiotracers. However, for nine pyrazole compounds for which rodent data are available, brain uptake and calculated logP values are not correlated. Thus, high logP values should not preclude evaluation of radiotracers for targets such as the CB 1 receptor that may require very lipophilic ligands

  5. Cannabis Users Show Enhanced Expression of CB1-5HT2A Receptor Heteromers in Olfactory Neuroepithelium Cells.

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    Galindo, Liliana; Moreno, Estefanía; López-Armenta, Fernando; Guinart, Daniel; Cuenca-Royo, Aida; Izquierdo-Serra, Mercè; Xicota, Laura; Fernandez, Cristina; Menoyo, Esther; Fernández-Fernández, José M; Benítez-King, Gloria; Canela, Enric I; Casadó, Vicent; Pérez, Víctor; de la Torre, Rafael; Robledo, Patricia

    2018-01-02

    Cannabinoid CB1 receptors (CB 1 R) and serotonergic 2A receptors (5HT 2A R) form heteromers in the brain of mice where they mediate the cognitive deficits produced by delta-9-tetrahydrocannabinol. However, it is still unknown whether the expression of this heterodimer is modulated by chronic cannabis use in humans. In this study, we investigated the expression levels and functionality of CB 1 R-5HT 2A R heteromers in human olfactory neuroepithelium (ON) cells of cannabis users and control subjects, and determined their molecular characteristics through adenylate cyclase and the ERK 1/2 pathway signaling studies. We also assessed whether heteromer expression levels correlated with cannabis consumption and cognitive performance in neuropsychological tests. ON cells from controls and cannabis users expressed neuronal markers such as βIII-tubulin and nestin, displayed similar expression levels of genes related to cellular self-renewal, stem cell differentiation, and generation of neural crest cells, and showed comparable Na + currents in patch clamp recordings. Interestingly, CB 1 R-5HT 2A R heteromer expression was significantly increased in cannabis users and positively correlated with the amount of cannabis consumed, and negatively with age of onset of cannabis use. In addition, a negative correlation was found between heteromer expression levels and attention and working memory performance in cannabis users and control subjects. Our findings suggest that cannabis consumption regulates the formation of CB 1 R-5HT 2A R heteromers, and may have a key role in cognitive processing. These heterodimers could be potential new targets to develop treatment alternatives for cognitive impairments.

  6. Human orexin/hypocretin receptors form constitutive homo- and heteromeric complexes with each other and with human CB1 cannabinoid receptors

    International Nuclear Information System (INIS)

    Jäntti, Maria H.; Mandrika, Ilona; Kukkonen, Jyrki P.

    2014-01-01

    Highlights: • OX 1 and OX 2 orexin and CB 1 cannabinoid receptor dimerization was investigated. • Bioluminescence resonance energy transfer method was used. • All receptors readily formed constitutive homo- and heteromeric complexes. - Abstract: Human OX 1 orexin receptors have been shown to homodimerize and they have also been suggested to heterodimerize with CB 1 cannabinoid receptors. The latter has been suggested to be important for orexin receptor responses and trafficking. In this study, we wanted to assess the ability of the other combinations of receptors to also form similar complexes. Vectors for expression of human OX 1 , OX 2 and CB 1 receptors, C-terminally fused with either Renilla luciferase or GFP 2 green fluorescent protein variant, were generated. The constructs were transiently expressed in Chinese hamster ovary cells, and constitutive dimerization between the receptors was assessed by bioluminescence energy transfer (BRET). Orexin receptor subtypes readily formed homo- and hetero(di)mers, as suggested by significant BRET signals. CB 1 receptors formed homodimers, and they also heterodimerized with both orexin receptors. Interestingly, BRET efficiency was higher for homodimers than for almost all heterodimers. This is likely to be due to the geometry of the interaction; the putatively symmetric dimers may place the C-termini in a more suitable orientation in homomers. Fusion of luciferase to an orexin receptor and GFP 2 to CB 1 produced more effective BRET than the opposite fusions, also suggesting differences in geometry. Similar was seen for the OX 1 –OX 2 interaction. In conclusion, orexin receptors have a significant propensity to make homo- and heterodi-/oligomeric complexes. However, it is unclear whether this affects their signaling. As orexin receptors efficiently signal via endocannabinoid production to CB 1 receptors, dimerization could be an effective way of forming signal complexes with optimal cannabinoid concentrations

  7. Biodistribution and dosimetry in humans of two inverse agonists to image cannabinoid CB1 receptors using positron emission tomography

    International Nuclear Information System (INIS)

    Terry, Garth E.; Hirvonen, Jussi; Liow, Jeih-San; Seneca, Nicholas; Morse, Cheryl L.; Pike, Victor W.; Innis, Robert B.; Tauscher, Johannes T.; Schaus, John M.; Phebus, Lee; Felder, Christian C.; Halldin, Christer

    2010-01-01

    Cannabinoid subtype 1 (CB 1 ) receptors are found in nearly every organ in the body, may be involved in several neuropsychiatric and metabolic disorders, and are therefore an active target for pharmacotherapy and biomarker development. We recently reported brain imaging of CB 1 receptors with two PET radioligands: 11 C-MePPEP and 18 F-FMPEP-d 2 . Here we describe the biodistribution and dosimetry estimates for these two radioligands. Seven healthy subjects (four men and three women) underwent whole-body PET scans for 120 min after injection with 11 C-MePPEP. Another seven healthy subjects (two men and five women) underwent whole-body PET scans for 300 min after injection with 18 F-FMPEP-d 2 . Residence times were acquired from regions of interest drawn on tomographic images of visually identifiable organs for both radioligands and from radioactivity excreted in urine for 18 F-FMPEP-d 2 . The effective doses of 11 C-MePPEP and 18 F-FMPEP-d 2 are 4.6 and 19.7 μSv/MBq, respectively. Both radioligands demonstrated high uptake of radioactivity in liver, lung, and brain shortly after injection and accumulated radioactivity in bone marrow towards the end of the scan. After injection of 11 C-MePPEP, radioactivity apparently underwent hepatobiliary excretion only, while radioactivity from 18 F-FMPEP-d 2 showed both hepatobiliary and urinary excretion. 11 C-MePPEP and 18 F-FMPEP-d 2 yield an effective dose similar to other PET radioligands labeled with either 11 C or 18 F. The high uptake in brain confirms the utility of these two radioligands to image CB 1 receptors in brain, and both may also be useful to image CB 1 receptors in the periphery. (orig.)

  8. The cannabinoid CB1 receptor and mTORC1 signalling pathways interact to modulate glucose homeostasis in mice

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    Francisco J. Bermudez-Silva

    2016-01-01

    Full Text Available The endocannabinoid system (ECS is an intercellular signalling mechanism that is present in the islets of Langerhans and plays a role in the modulation of insulin secretion and expansion of the β-cell mass. The downstream signalling pathways mediating these effects are poorly understood. Mammalian target of rapamycin complex 1 (mTORC1 signalling is a key intracellular pathway involved in energy homeostasis and is known to importantly affect the physiology of pancreatic islets. We investigated the possible relationship between cannabinoid type 1 (CB1 receptor signalling and the mTORC1 pathway in the endocrine pancreas of mice by using pharmacological analysis as well as mice genetically lacking the CB1 receptor or the downstream target of mTORC1, the kinase p70S6K1. In vitro static secretion experiments on islets, western blotting, and in vivo glucose and insulin tolerance tests were performed. The CB1 receptor antagonist rimonabant decreased glucose-stimulated insulin secretion (GSIS at 0.1 µM while increasing phosphorylation of p70S6K1 and ribosomal protein S6 (rpS6 within the islets. Specific pharmacological blockade of mTORC1 by 3 nM rapamycin, as well as genetic deletion of p70S6K1, impaired the CB1-antagonist-mediated decrease in GSIS. In vivo experiments showed that 3 mg/kg body weight rimonabant decreased insulin levels and induced glucose intolerance in lean mice without altering peripheral insulin sensitivity; this effect was prevented by peripheral administration of low doses of rapamycin (0.1 mg/kg body weight, which increased insulin sensitivity. These findings suggest a functional interaction between the ECS and the mTORC1 pathway within the endocrine pancreas and at the whole-organism level, which could have implications for the development of new therapeutic approaches for pancreatic β-cell diseases.

  9. A cell population that strongly expresses the CB1 cannabinoid receptor in the ependyma of the rat spinal cord.

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    Garcia-Ovejero, Daniel; Arevalo-Martin, Angel; Paniagua-Torija, Beatriz; Sierra-Palomares, Yolanda; Molina-Holgado, Eduardo

    2013-01-01

    The cells surrounding the central canal of the spinal cord are a source of stem/precursor cells that may give rise to neurons, astrocytes, or oligodendrocytes. However, they are a heterogeneous population that remains poorly understood. Here we describe a subpopulation characterized by their strong expression of the CB(1) cannabinoid receptor, oval/round soma, apical nucleus, a variable number of cilia (0, 1, or 2), and the presence of a single short and occasionally ramified basal process. These cells are mainly located in the lateral and dorsal central canal throughout the spinal cord. These CB(1)(HIGH) cells are closely related to the basal lamina labyrinths or fractones derived from subependymal microglia. In addition, CB(1)(HIGH) cells express some stem/precursor cell markers, including vimentin, nestin, Sox2, Sox9, and GLAST, but not others such as CD15 or GFAP. In addition, this cell population does not proliferate in the intact adult spinal cord, although up to 50% of these cells express the proliferation marker Ki67 in newly born rats or after a spinal cord contusion. The present findings contribute to our understanding of the spinal cord central canal structure and reveal the targets for endocannabinoids inside this neurogenic niche. Copyright © 2012 Wiley Periodicals, Inc.

  10. Gz mediates the long-lasting desensitization of brain CB1 receptors and is essential for cross-tolerance with morphine

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    Rodríguez-Muñoz María

    2009-03-01

    Full Text Available Abstract Background Although the systemic administration of cannabinoids produces antinociception, their chronic use leads to analgesic tolerance as well as cross-tolerance to morphine. These effects are mediated by cannabinoids binding to peripheral, spinal and supraspinal CB1 and CB2 receptors, making it difficult to determine the relevance of each receptor type to these phenomena. However, in the brain, the CB1 receptors (CB1Rs are expressed at high levels in neurons, whereas the expression of CB2Rs is marginal. Thus, CB1Rs mediate the effects of smoked cannabis and are also implicated in emotional behaviors. We have analyzed the production of supraspinal analgesia and the development of tolerance at CB1Rs by the direct injection of a series of cannabinoids into the brain. The influence of the activation of CB1Rs on supraspinal analgesia evoked by morphine was also evaluated. Results Intracerebroventricular (icv administration of cannabinoid receptor agonists, WIN55,212-2, ACEA or methanandamide, generated a dose-dependent analgesia. Notably, a single administration of these compounds brought about profound analgesic tolerance that lasted for more than 14 days. This decrease in the effect of cannabinoid receptor agonists was not mediated by depletion of CB1Rs or the loss of regulated G proteins, but, nevertheless, it was accompanied by reduced morphine analgesia. On the other hand, acute morphine administration produced tolerance that lasted only 3 days and did not affect the CB1R. We found that both neural mu-opioid receptors (MORs and CB1Rs interact with the HINT1-RGSZ module, thereby regulating pertussis toxin-insensitive Gz proteins. In mice with reduced levels of these Gz proteins, the CB1R agonists produced no such desensitization or morphine cross-tolerance. On the other hand, experimental enhancement of Gz signaling enabled an acute icv administration of morphine to produce a long-lasting tolerance at MORs that persisted for more than

  11. The anabolic steroid nandrolone alters cannabinoid self-administration and brain CB1 receptor density and function.

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    Struik, Dicky; Fadda, Paola; Zara, Tamara; Zamberletti, Erica; Rubino, Tiziana; Parolaro, Daniela; Fratta, Walter; Fattore, Liana

    2017-01-01

    Clinical and pre-clinical observations indicate that anabolic-androgenic steroids can induce neurobiological changes that alter the rewarding effects of drugs of abuse. In this study, we investigated the effect of the anabolic steroid nandrolone on the rewarding properties of the cannabinoid CB 1 receptor agonist WIN55,212-2 (WIN) in rats. Lister Hooded male rats were treated intramuscularly with nandrolone (15mg/kg) or vehicle for 14 consecutive days, and then allowed to self-administer WIN (12.5μg/kg/infusion) intravenously. After reaching stable drug intake, self-administration behavior was extinguished to examine drug- and cue-induced reinstatement of cannabinoid-seeking behavior. Other behavioral parameters presumed to influence drug-taking and drug-seeking behaviors were examined to gain more insight into the behavioral specificity of nandrolone treatment. Finally, animals were sacrificed for analysis of CB 1 receptor density and function in selected brain areas. We found that nandrolone-treated rats self-administered up to 2 times more cannabinoid than vehicle-treated rats, but behaved similarly to control rats when tested for drug- and cue-induced reinstatement of cannabinoid-seeking behavior. Enhanced cannabinoid intake by nandrolone-treated rats was not accompanied by changes in locomotor activity, sensorimotor gating, or memory function. However, our molecular data show that after chronic WIN self-administration nandrolone-treated rats display altered CB 1 receptor density and function in selected brain areas. We hypothesize that increased cannabinoid self-administration in nandrolone-treated rats results from a nandrolone-induced decrease in reward function, which rats seem to compensate by voluntarily increasing their cannabinoid intake. Altogether, our findings corroborate the hypothesis that chronic exposure to anabolic-androgenic steroids induces dysfunction of the reward pathway in rats and might represent a potential risk factor for abuse of

  12. Immunohistochemical characterisation and localisation of cannabinoid CB1 receptor protein in the rat vestibular nucleus complex and the effects of unilateral vestibular deafferentation.

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    Ashton, John C; Zheng, Yiwen; Liu, Ping; Darlington, Cynthia L; Smith, Paul F

    2004-09-24

    CB1 receptor expression has been reported to be low in the brainstem compared with the forebrain, and low in the vestibular nucleus complex (VNC) compared with other regions in the brainstem. However, a frequent effect of cannabis is dizziness and loss of balance. This may be due to the activation of cannabinoid receptors in the central vestibular pathways. We used immunohistochemistry to study the distribution of CB1 receptor protein in the VNC, and Western blotting to measure CB1 receptor expression in the VNC following unilateral vestibular deafferentation (UVD); the hippocampal CA1, CA2/3 and dentate gyrus (DG) regions were also analysed for comparison. This study confirms a previous electrophysiological demonstration that CB1 receptors exist in significant densities in the VNC and are likely to contribute to the neurochemical control of the vestibular reflexes. Nonetheless, CB1 receptor expression did not change significantly in the VNC during vestibular compensation. In addition, despite some small but significant changes in CB1 receptor expression in the CA2/3 and the DG following UVD, in no case were these differences statistically significant in comparison to both control groups.

  13. Cannabinoid CB1 receptors in distinct circuits of the extended amygdala determine fear responsiveness to unpredictable threat.

    Science.gov (United States)

    Lange, M D; Daldrup, T; Remmers, F; Szkudlarek, H J; Lesting, J; Guggenhuber, S; Ruehle, S; Jüngling, K; Seidenbecher, T; Lutz, B; Pape, H C

    2017-10-01

    The brain circuits underlying behavioral fear have been extensively studied over the last decades. Although the vast majority of experimental studies assess fear as a transient state of apprehension in response to a discrete threat, such phasic states of fear can shift to a sustained anxious apprehension, particularly in face of diffuse cues with unpredictable environmental contingencies. Unpredictability, in turn, is considered an important variable contributing to anxiety disorders. The networks of the extended amygdala have been suggested keys to the control of phasic and sustained states of fear, although the underlying synaptic pathways and mechanisms remain poorly understood. Here, we show that the endocannabinoid system acting in synaptic circuits of the extended amygdala can explain the fear response profile during exposure to unpredictable threat. Using fear training with predictable or unpredictable cues in mice, combined with local and cell-type-specific deficiency and rescue of cannabinoid type 1 (CB1) receptors, we found that presynaptic CB1 receptors on distinct amygdala projections to bed nucleus of the stria terminalis (BNST) are both necessary and sufficient for the shift from phasic to sustained fear in response to an unpredictable threat. These results thereby identify the causal role of a defined protein in a distinct brain pathway for the temporal development of a sustained state of anxious apprehension during unpredictability of environmental influences, reminiscent of anxiety symptoms in humans.

  14. Mice Expressing a "Hyper-Sensitive" Form of the Cannabinoid Receptor 1 (CB1 Are Neither Obese Nor Diabetic.

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    David J Marcus

    Full Text Available Multiple lines of evidence implicate the endocannabinoid signaling system in the modulation of metabolic disease. Genetic or pharmacological inactivation of CB1 in rodents leads to reduced body weight, resistance to diet-induced obesity, decreased intake of highly palatable food, and increased energy expenditure. Cannabinoid agonists stimulate feeding in rodents and increased levels of endocannabinoids can disrupt lipid metabolism. Therefore, the hypothesis that sustained endocannabinoid signaling can lead to obesity and diabetes was examined in this study using S426A/S430A mutant mice expressing a desensitization-resistant CB1 receptor. These mice display exaggerated and prolonged responses to acute administration of phytocannabinoids, synthetic cannabinoids, and endocannabinoids. As a consequence these mice represent a novel model for determining the effect of enhanced endocannabinoid signaling on metabolic disease. S426A/S430A mutants consumed equivalent amounts of both high fat (45% and low fat (10% chow control diet compared to wild-type littermate controls. S426A/S430A mutants and wild-type mice fed either high or low fat control diet displayed similar fasting blood glucose levels and normal glucose clearance following a 2 g/kg glucose challenge. Furthermore, S426A/S430A mutants and wild-type mice consumed similar amounts of chow following an overnight fast. While both THC and JZL195 significantly increased food intake two hours after injection, this increase was similar between the S426A/S430A mutant and wildtype control mice Our results indicate that S426A/S430A mutant mice expressing the desensitization-resistant form of CB1 do not exhibit differences in body weight, food intake, glucose homeostasis, or re-feeding following a fast.

  15. Chronic and acute adenosine A2A receptor blockade prevents long-term episodic memory disruption caused by acute cannabinoid CB1 receptor activation.

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    Mouro, Francisco M; Batalha, Vânia L; Ferreira, Diana G; Coelho, Joana E; Baqi, Younis; Müller, Christa E; Lopes, Luísa V; Ribeiro, Joaquim A; Sebastião, Ana M

    2017-05-01

    Cannabinoid-mediated memory impairment is a concern in cannabinoid-based therapies. Caffeine exacerbates cannabinoid CB 1 receptor (CB 1 R)-induced memory deficits through an adenosine A 1 receptor-mediated mechanism. We now evaluated how chronic or acute blockade of adenosine A 2A receptors (A 2A Rs) affects long-term episodic memory deficits induced by a single injection of a selective CB 1 R agonist. Long-term episodic memory was assessed by the novel object recognition (NOR) test. Mice received an intraperitoneal (i.p.) injection of the CB 1 /CB 2 receptor agonist WIN 55,212-2 (1 mg/kg) immediately after the NOR training, being tested for novelty recognition 24 h later. Anxiety levels were assessed by the Elevated Plus Maze test, immediately after the NOR. Mice were also tested for exploratory behaviour at the Open Field. For chronic A 2A R blockade, KW-6002 (istradefylline) (3 mg/kg/day) was administered orally for 30 days; acute blockade of A 2A Rs was assessed by i.p. injection of SCH 58261 (1 mg/kg) administered either together with WIN 55,212-2 or only 30 min before the NOR test phase. The involvement of CB 1 Rs was assessed by using the CB 1 R antagonist, AM251 (3 mg/kg, i.p.). WIN 55,212-2 caused a disruption in NOR, an action absent in mice also receiving AM251, KW-6002 or SCH 58261 during the encoding/consolidation phase; SCH 58251 was ineffective if present during retrieval only. No effects were detected in the Elevated Plus maze or Open Field Test. The finding that CB 1 R-mediated memory disruption is prevented by antagonism of adenosine A 2A Rs, highlights a possibility to prevent cognitive side effects when therapeutic application of CB 1 R drugs is desired. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. AAV-mediated overexpression of the CB1 receptor in the mPFC of adult rats alters cognitive flexibility, social behavior and emotional reactivity

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    Matthias eKlugmann

    2011-07-01

    Full Text Available The endocannabinoid (ECB system is strongly involved in the regulation of cognitive processing and emotional behavior and evidence indicates that ECB signaling might affect these behavioral abilities by modulations of prefrontal cortical functions. The aim of the present study was to examine the role of the CB1 receptor in the medial prefrontal cortex (mPFC on cognitive flexibility and emotional behavior. Therefore, the CB1 receptor was overexpressed by adeno-associated virus (AAV vector-mediated gene transfer specifically in the mPFC of adult Wistar rats. Animals were then tested in different anxiety-related paradigms for emotional reactivity (e.g. elevated plus maze (EPM, light/dark emergence test (EMT, social interaction and the attentional set shift task (ASST - an adaptation of the human Wisconsin card sorting test - for cognitive abilities and behavioral flexibility. A subtle increase in exploratory behavior was found in CB1 receptor overexpressing animals (CB1-R compared to empty vector injected controls (Empty in the EMT and EPM, although general locomotor activity did not differ between the groups. During social interaction testing, social contact behavior towards the unknown conspecific was found to be decreased, whereas social withdrawal was increased in CB1-R animals and they showed an inadequate increase in exploratory behavior compared to control animals. In the ASST, impaired reversal learning abilities were detected in CB1-R animals compared to controls, indicating reduced behavioral flexibility. In conclusion, upregulation of the CB1 receptor specifically in the rat mPFC induces alterations in emotional reactivity, leads to inadequate social behavior and impairs cognitive flexibility. These findings might be relevant for neuropsychiatric disorders, since higher cortical CB1 receptor expression levels as well as similar behavioral impairments as observed in the present study have been described in schizophrenic patients.

  17. Detection of Heteromers Formed by Cannabinoid CB1, Dopamine D2, and Adenosine A2A G-Protein-Coupled Receptors by Combining Bimolecular Fluorescence Complementation and Bioluminescence Energy Transfer

    Science.gov (United States)

    Navarro, Gemma; Carriba, Paulina; Gandí, Jorge; Ciruela, Francisco; Casadó, Vicent; Cortés, Antoni; Mallol, Josefa; Canela, Enric I.; Lluis, Carmen; Franco, Rafael

    2008-01-01

    Functional interactions in signaling occur between dopamine D2 (D2R) and cannabinoid CB1 (CB1R) receptors, between CB1R and adenosine A2A (A2AR) receptors, and between D2R and A2AR. Furthermore, direct molecular interactions have been reported for the pairs CB1R-D2R, A2AR-D2R, and CB1R-A2AR. Here a combination of bimolecular fluorescence complementation and bioluminescence energy transfer techniques was used to identify the occurrence of D2R-CB1R-A2AR hetero-oligomers in living cells. PMID:18956124

  18. Detection of Heteromers Formed by Cannabinoid CB1, Dopamine D2, and Adenosine A2A G-Protein-Coupled Receptors by Combining Bimolecular Fluorescence Complementation and Bioluminescence Energy Transfer

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    Gemma Navarro

    2008-01-01

    Full Text Available Functional interactions in signaling occur between dopamine D2 (D2R and cannabinoid CB1 (CB1R receptors, between CB1R and adenosine A2A (A2AR receptors, and between D2R and A2AR. Furthermore, direct molecular interactions have been reported for the pairs CB1R-D2R, A2AR-D2R, and CB1R-A2AR. Here a combination of bimolecular fluorescence complementation and bioluminescence energy transfer techniques was used to identify the occurrence of D2R-CB1R-A2AR hetero-oligomers in living cells.

  19. Effect of cannabinoids CB1 receptors blockade on hemodynamic parameters and endothelial function at the immobilization stress in the experiment

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    S. V. Gavreliuk

    2017-12-01

    Full Text Available The aim of the study was to evaluate the response of hemodynamic parameters and changes in endothelial function in modeling of CB1 cannabinoid receptors blockade in chronic stress. Materials and мethods. The study was performed on four groups of hundred-day-old rats, which were examined by ultrasonic scanning during the ten-day period of the experiment. The first group consisted of intact animals; the second group – animals, which were exposed to immobilization stress; the third – animals which were given a solution of rimonabant hydrochloride at the rate of 10 mg×kg-1 of animal weight per day daily per os; the fourth group consisted of animals which daily received a solution of rimonabant hydrochloride at the rate of 10 mg×kg-1 of animal weight per day and were exposed to immobilization stress. The intraluminal vessel diameter, the intima-media complex thickness, endothelium-dependent and endothelium-independent dilation were quantified in the ultrasound examination. Quantitative characteristics of the blood flow were studied: peak systolic velocity, end diastolic velocity, resistive index and peak-systolic/end-diastolic ratio, and estimated mean blood flow velocity. Results. It has been found that the effect of chronic immobilization stress in 100-day-old male rats causes intima-media complex structure and thickness change, endothelial dysfunction and increase in the abdominal aorta intraluminal diameter. Hemodynamics changes are characterized by a decrease in the average blood flow velocity and an increase in the values of indices characterizing the vascular wall peripheral resistance. Prolonged blockade of cannabinoids CB1 receptors leads to endothelial dysfunction development, a decrease in the intraluminal diameter of the abdominal aorta and a decrease in the average blood flow velocity while vascular wall elastic properties maintaining. This affects the sensitivity of cardiovascular system to nitrogen oxide, which is manifested by

  20. Contrasting effects of lithium chloride and CB1 receptor blockade on enduring changes in the valuation of reward.

    Directory of Open Access Journals (Sweden)

    Giovanni eHernandez

    2011-09-01

    Full Text Available When an organism has been trained to respond for a reward, its learned behavior can be characterized as goal-directed or habitual based on whether or not it is susceptible to reward devaluation. Here, we evaluated whether instrumental responding for brain stimulation reward (BSR can devalued using a paradigm traditionally used for natural rewards. Rats were trained to lever press for BSR. Subsequently, BSR was paired with either lithium chloride (LiCl, 5 mg/kg, i.p, a pro-emetic, or AM251, a CB1 receptor antagonist (3 mg/kg, i.p.. Pairings of BSR with these two compounds or their respective vehicle were performed in a novel environment so that only unconditional effects of BSR were affected by the pharmacological manipulations. Subsequently, in a probe test, all rats were returned in the drug-free state to the boxes where they had received training instrumental responding was reassessed in the absence of BSR delivery. LiCl produced enduring decreases in the number of responses during the test session, whereas AM251 had no effect. These results show that instrumental responding for BSR is susceptible to devaluation, in accord with the proposal that this behavior is supported at least in part by associations between the response and the rewarding outcome. Furthermore, they suggest that the reward modulation observed in studies involving the use of CB1 receptor antagonists arises from changes in the organism’s motivation rather than due to drug-induced changes in the intrinsic value of reward.

  1. Contrasting Effects of Lithium Chloride and CB1 Receptor Blockade on Enduring Changes in the Valuation of Reward.

    Science.gov (United States)

    Hernandez, Giovanni; Bernstein, David; Schoenbaum, Geoffrey; Cheer, Joseph F

    2011-01-01

    When an organism responds for a reward, its learned behavior can be characterized as goal-directed or habitual based on whether or not it is susceptible to reward devaluation. Here, we evaluated whether instrumental responding for brain stimulation reward (BSR) can be devalued using a paradigm traditionally used for natural rewards. Rats were trained to lever press for BSR; afterward, BSR was paired with either lithium chloride (LiCl, 5 mg/kg, i.p.), a pro-emetic, or AM251, a CB1 receptor antagonist (3 mg/kg, i.p.) or the vehicle of these compounds. Pairings of BSR with these compounds and their vehicles were performed in a novel environment so that only unconditional effects of BSR would be affected by the pharmacological manipulations. Subsequently, in a probe test, all rats were returned in the drug-free state to the boxes where they had received training and instrumental responding was reassessed in the absence of BSR delivery. When compared to control, LiCl produced a significant decrease in the number of responses during the test session, whereas AM251 did not. These results show that instrumental responding for BSR is susceptible to devaluation, in accord with the proposal that this behavior is supported at least in part by associations between the response and the rewarding outcome. Further, they suggest that reward modulation observed in studies involving the use of CB1 receptor antagonists arises from changes in the organism's motivation rather than drug-induced changes in the intrinsic value of reward.

  2. Evidence against a critical role of CB1 receptors in adaptation of the hypothalamic-pituitary-adrenal axis and other consequences of daily repeated stress.

    Science.gov (United States)

    Rabasa, Cristina; Pastor-Ciurana, Jordi; Delgado-Morales, Raúl; Gómez-Román, Almudena; Carrasco, Javier; Gagliano, Humberto; García-Gutiérrez, María S; Manzanares, Jorge; Armario, Antonio

    2015-08-01

    There is evidence that endogenous cannabinoids (eCBs) play a role in the control of the hypothalamic-pituitary-adrenal (HPA) axis, although they appear to have dual, stimulatory and inhibitory, effects. Recent data in rats suggest that eCBs, acting through CB1 receptors (CB1R), may be involved in adaptation of the HPA axis to daily repeated stress. In the present study we analyze this issue in male mice and rats. Using a knock-out mice for the CB1 receptor (CB1-/-) we showed that mutant mice presented similar adrenocorticotropic hormone (ACTH) response to the first IMO as wild-type mice. Daily repeated exposure to 1h of immobilization reduced the ACTH response to the stressor, regardless of the genotype, demonstrating that adaptation occurred to the same extent in absence of CB1R. Prototypical changes observed after repeated stress such as enhanced corticotropin releasing factor (CRH) gene expression in the paraventricular nucleus of the hypothalamus, impaired body weight gain and reduced thymus weight were similarly observed in both genotypes. The lack of effect of CB1R in the expression of HPA adaptation to another similar stressor (restraint) was confirmed in wild-type CD1 mice by the lack of effect of the CB1R antagonist AM251 just before the last exposure to stress. Finally, the latter drug did not blunt the HPA, glucose and behavioral adaptation to daily repeated forced swim in rats. Thus, the present results indicate that CB1R is not critical for overall effects of daily repeated stress or proper adaptation of the HPA axis in mice and rats. Copyright © 2015 Elsevier B.V. and ECNP. All rights reserved.

  3. Involvement of Gaba and Cannabinoid Receptors in Central Food Intake Regulation in Neonatal Layer Chicks: Role of CB1 and Gabaa Receptors

    Directory of Open Access Journals (Sweden)

    M Zendehdel

    Full Text Available ABSTRACT Feeding behavior is regulated via a complex network which interacts via diverse signals from central and peripheral tissues. Endocannabinoids modulate release of GABA in a variety of regions of the central nervous system. Endocannabinoids and GABAergic system have an important role in the central regulation of appetite. Thus, the present study examines the possible interaction of central canabinoidergic and GABAergic systems on food intake in 3-h food-deprived (FD3 neonatal layer-type chicks. The results of this study showed that intracerebroventricular (ICV injection of 2-AG (2-Arachidonoylglycerol, selective CB1 receptors agonist, 2µg significantly increased food intake and this effect of 2-AG was attenuated by Picrotoxin (GABAA antagonist, 0.5µg (P0.05. Also, hyperphagic effect of CB65 (CB2 receptors agonist, 1.25µg was not affected by Picrotoxin or CGP54626 (p>0.05. Moreover, the food intake of chicks was significantly increased by ICV injection of GABAA agonist (Gaboxadol, 0.2 µg and SR141716A (CB1 receptors antagonist, 6.25µg significantly decreased Gaboxadol-induced hyperphagia (P0.05. These data showed there might be an interaction between central cannabinoidergic and GABAergic systems via CB1 and GABAA receptors in control of food intake in neonatal layer chicks.

  4. Temporal changes of CB1 cannabinoid receptor in the basal ganglia as a possible structure-specific plasticity process in 6-OHDA lesioned rats.

    Directory of Open Access Journals (Sweden)

    Gabriela P Chaves-Kirsten

    Full Text Available The endocannabinoid system has been implicated in several neurobiological processes, including neurodegeneration, neuroprotection and neuronal plasticity. The CB1 cannabinoid receptors are abundantly expressed in the basal ganglia, the circuitry that is mostly affected in Parkinson's Disease (PD. Some studies show variation of CB1 expression in basal ganglia in different animal models of PD, however the results are quite controversial, due to the differences in the procedures employed to induce the parkinsonism and the periods analyzed after the lesion. The present study evaluated the CB1 expression in four basal ganglia structures, namely striatum, external globus pallidus (EGP, internal globus pallidus (IGP and substantia nigra pars reticulata (SNpr of rats 1, 5, 10, 20, and 60 days after unilateral intrastriatal 6-hydroxydopamine injections, that causes retrograde dopaminergic degeneration. We also investigated tyrosine hydroxylase (TH, parvalbumin, calbindin and glutamic acid decarboxylase (GAD expression to verify the status of dopaminergic and GABAergic systems. We observed a structure-specific modulation of CB1 expression at different periods after lesions. In general, there were no changes in the striatum, decreased CB1 in IGP and SNpr and increased CB1 in EGP, but this increase was not sustained over time. No changes in GAD and parvalbumin expression were observed in basal ganglia, whereas TH levels were decreased and the calbindin increased in striatum in short periods after lesion. We believe that the structure-specific variation of CB1 in basal ganglia in the 6-hydroxydopamine PD model could be related to a compensatory process involving the GABAergic transmission, which is impaired due to the lack of dopamine. Our data, therefore, suggest that the changes of CB1 and calbindin expression may represent a plasticity process in this PD model.

  5. Cognitive Impairment Induced by Delta9-tetrahydrocannabinol Occurs through Heteromers between Cannabinoid CB1 and Serotonin 5-HT2A Receptors.

    Science.gov (United States)

    Viñals, Xavier; Moreno, Estefanía; Lanfumey, Laurence; Cordomí, Arnau; Pastor, Antoni; de La Torre, Rafael; Gasperini, Paola; Navarro, Gemma; Howell, Lesley A; Pardo, Leonardo; Lluís, Carmen; Canela, Enric I; McCormick, Peter J; Maldonado, Rafael; Robledo, Patricia

    2015-07-01

    Activation of cannabinoid CB1 receptors (CB1R) by delta9-tetrahydrocannabinol (THC) produces a variety of negative effects with major consequences in cannabis users that constitute important drawbacks for the use of cannabinoids as therapeutic agents. For this reason, there is a tremendous medical interest in harnessing the beneficial effects of THC. Behavioral studies carried out in mice lacking 5-HT2A receptors (5-HT2AR) revealed a remarkable 5-HT2AR-dependent dissociation in the beneficial antinociceptive effects of THC and its detrimental amnesic properties. We found that specific effects of THC such as memory deficits, anxiolytic-like effects, and social interaction are under the control of 5-HT2AR, but its acute hypolocomotor, hypothermic, anxiogenic, and antinociceptive effects are not. In biochemical studies, we show that CB1R and 5-HT2AR form heteromers that are expressed and functionally active in specific brain regions involved in memory impairment. Remarkably, our functional data shows that costimulation of both receptors by agonists reduces cell signaling, antagonist binding to one receptor blocks signaling of the interacting receptor, and heteromer formation leads to a switch in G-protein coupling for 5-HT2AR from Gq to Gi proteins. Synthetic peptides with the sequence of transmembrane helices 5 and 6 of CB1R, fused to a cell-penetrating peptide, were able to disrupt receptor heteromerization in vivo, leading to a selective abrogation of memory impairments caused by exposure to THC. These data reveal a novel molecular mechanism for the functional interaction between CB1R and 5-HT2AR mediating cognitive impairment. CB1R-5-HT2AR heteromers are thus good targets to dissociate the cognitive deficits induced by THC from its beneficial antinociceptive properties.

  6. Cognitive Impairment Induced by Delta9-tetrahydrocannabinol Occurs through Heteromers between Cannabinoid CB1 and Serotonin 5-HT2A Receptors.

    Directory of Open Access Journals (Sweden)

    Xavier Viñals

    2015-07-01

    Full Text Available Activation of cannabinoid CB1 receptors (CB1R by delta9-tetrahydrocannabinol (THC produces a variety of negative effects with major consequences in cannabis users that constitute important drawbacks for the use of cannabinoids as therapeutic agents. For this reason, there is a tremendous medical interest in harnessing the beneficial effects of THC. Behavioral studies carried out in mice lacking 5-HT2A receptors (5-HT2AR revealed a remarkable 5-HT2AR-dependent dissociation in the beneficial antinociceptive effects of THC and its detrimental amnesic properties. We found that specific effects of THC such as memory deficits, anxiolytic-like effects, and social interaction are under the control of 5-HT2AR, but its acute hypolocomotor, hypothermic, anxiogenic, and antinociceptive effects are not. In biochemical studies, we show that CB1R and 5-HT2AR form heteromers that are expressed and functionally active in specific brain regions involved in memory impairment. Remarkably, our functional data shows that costimulation of both receptors by agonists reduces cell signaling, antagonist binding to one receptor blocks signaling of the interacting receptor, and heteromer formation leads to a switch in G-protein coupling for 5-HT2AR from Gq to Gi proteins. Synthetic peptides with the sequence of transmembrane helices 5 and 6 of CB1R, fused to a cell-penetrating peptide, were able to disrupt receptor heteromerization in vivo, leading to a selective abrogation of memory impairments caused by exposure to THC. These data reveal a novel molecular mechanism for the functional interaction between CB1R and 5-HT2AR mediating cognitive impairment. CB1R-5-HT2AR heteromers are thus good targets to dissociate the cognitive deficits induced by THC from its beneficial antinociceptive properties.

  7. CB1 cannabinoid receptor-mediated anandamide signaling mechanisms of the inferior colliculus modulate the haloperidol-induced catalepsy.

    Science.gov (United States)

    Medeiros, P; de Freitas, R L; Silva, M O; Coimbra, N C; Melo-Thomas, L

    2016-11-19

    The inferior colliculus (IC), a midbrain structure that processes acoustic information of aversive nature, is distinguished from other auditory nuclei in the brainstem by its connections with structures of the motor system. Previous evidence relating the IC to motor behavior shows that glutamatergic and GABAergic mechanisms in the IC exert influence on systemic haloperidol-induced catalepsy. There is substantial evidence supporting a role played by the endocannabinoid system as a modulator of the glutamatergic neurotransmission, as well as the dopaminergic activity in the basal nuclei and therefore it may be considered as a potential pharmacological target for the treatment of movement disorders. The present study evaluated if the endocannabinoid system in the IC plays a role in the elaboration of systemic haloperidol-induced catalepsy. Male Wistar rats received intracollicular microinjection of either the endogenous cannabinoid anandamide (AEA) at different concentrations (5, 50 or 100pmol/0.2μl), the CB 1 cannabinoid receptor antagonist AM251 at 50, 100 or 200pmol/0.2μl or vehicle, followed by intraperitoneal (IP) administration of either haloperidol at 0.5 or 1mg/kg or physiological saline. Systemic injection of haloperidol at both doses (0.5 or 1mg/kg, IP) produced a cataleptic state, compared to vehicle/physiological saline-treated group, lasting 30 and 50min after systemic administration of the dopaminergic receptors non-selective antagonist. The midbrain microinjection of AEA at 50pmol/0.2μl increased the latency for stepping down from the horizontal bar after systemic administration of haloperidol. Moreover, the intracollicular administration of AEA at 50pmol/0.2μl was able to increase the duration of catalepsy as compared to AEA at 100pmol/0.2-μl-treated group. Intracollicular pretreatment with AM251 at the intermediate concentration (100pmol/0.2μl) was able to decrease the duration of catalepsy after systemic administration of haloperidol. However

  8. CB1 Receptor-Mediated Signaling Underlies the Hippocampal Synaptic, Learning and Memory Deficits Following Treatment with JWH-081, a New Component of Spice/K2 Preparations

    OpenAIRE

    Basavarajappa, Balapal S.; Subbanna, Shivakumar

    2014-01-01

    Recently, synthetic cannabinoids have been sprayed onto plant material, which is subsequently packaged and sold as “Spice” or “K2” to mimic the effects of marijuana. A recent report identified several synthetic additives in samples of “Spice/K2”, including JWH-081, a synthetic ligand for the cannabinoid receptor 1 (CB1). The deleterious effects of JWH-081 on brain function are not known, particularly on CB1 signaling, synaptic plasticity, learning and memory. Here, we evaluated the effects of...

  9. Blockade of Cannabinoid CB1 Receptors in the Dorsal Periaqueductal Gray Unmasks the Antinociceptive Effect of Local Injections of Anandamide in Mice

    Directory of Open Access Journals (Sweden)

    Diego C. Mascarenhas

    2017-10-01

    Full Text Available Divergent results in pain management account for the growing number of studies aiming at elucidating the pharmacology of the endocannabinoid/endovanilloid anandamide (AEA within several pain-related brain structures. For instance, the stimulation of both Transient Receptor Potential Vanilloid type 1 (TRPV1 and Cannabinoid type 1 (CB1 receptors led to paradoxical effects on nociception. Here, we attempted to propose a clear and reproducible methodology to achieve the antinociceptive effect of exogenous AEA within the dorsal periaqueductal gray (dPAG of mice exposed to the tail-flick test. Accordingly, male Swiss mice received intra-dPAG injection of AEA (CB1/TRPV1 agonist, capsaicin (TRPV1 agonist, WIN (CB1 agonist, AM251 (CB1 antagonist, and 6-iodonordihydrocapsaicin (6-IODO (TRPV1 selective antagonist and their nociceptive response was assessed with the tail-flick test. In order to assess AEA effects on nociception specifically at vanilloid or cannabinoid (CB substrates into the dPAG, mice underwent an intrinsically inactive dose of AM251 or 6-IODO followed by local AEA injections and were subjected to the same test. While intra-dPAG AEA did not change acute pain, local injections of capsaicin or WIN induced a marked TRPV1- and CB1-dependent antinociceptive effect, respectively. Regarding the role of AEA specifically at CB/vanilloid substrates, while the blockade of TRPV1 did not change the lack of effects of intra-dPAG AEA on nociception, local pre-treatment of AM251, a CB1 antagonist, led to a clear AEA-induced antinociception. It seems that the exogenous AEA-induced antinociception is unmasked when it selectively binds to vanilloid substrates, which might be useful to address acute pain in basic and perhaps clinical trials.

  10. Cytotoxicity of synthetic cannabinoids on primary neuronal cells of the forebrain: the involvement of cannabinoid CB1 receptors and apoptotic cell death

    International Nuclear Information System (INIS)

    Tomiyama, Ken-ichi; Funada, Masahiko

    2014-01-01

    The abuse of herbal products containing synthetic cannabinoids has become an issue of public concern. The purpose of this paper was to evaluate the acute cytotoxicity of synthetic cannabinoids on mouse brain neuronal cells. Cytotoxicity induced by synthetic cannabinoid (CP-55,940, CP-47,497, CP-47,497-C8, HU-210, JWH-018, JWH-210, AM-2201, and MAM-2201) was examined using forebrain neuronal cultures. These synthetic cannabinoids induced cytotoxicity in the forebrain cultures in a concentration-dependent manner. The cytotoxicity was suppressed by preincubation with the selective CB 1 receptor antagonist AM251, but not with the selective CB 2 receptor antagonist AM630. Furthermore, annexin-V-positive cells were found among the treated forebrain cells. Synthetic cannabinoid treatment induced the activation of caspase-3, and preincubation with a caspase-3 inhibitor significantly suppressed the cytotoxicity. These synthetic cannabinoids induced apoptosis through a caspase-3-dependent mechanism in the forebrain cultures. Our results indicate that the cytotoxicity of synthetic cannabinoids towards primary neuronal cells is mediated by the CB 1 receptor, but not by the CB 2 receptor, and further suggest that caspase cascades may play an important role in the apoptosis induced by these synthetic cannabinoids. In conclusion, excessive synthetic cannabinoid abuse may present a serious acute health concern due to neuronal damage or deficits in the brain. - Highlights: • Synthetic cannabinoids (classical cannabinoids, non-classical cannabinoids, and aminoalkylindole derivatives) induce cytotoxicity in mouse forebrain cultures. • Synthetic cannabinoid-induced cytotoxicity towards forebrain cultures is mediated by the CB 1 receptor, but not by the CB 2 receptor, and involves caspase-dependent apoptosis. • A high concentration of synthetic cannabinoids may be toxic to neuronal cells that express CB 1 receptors

  11. Reinforcing and neurochemical effects of cannabinoid CB1 receptor agonists, but not cocaine, are altered by an adenosine A2A receptor antagonist.

    Science.gov (United States)

    Justinová, Zuzana; Ferré, Sergi; Redhi, Godfrey H; Mascia, Paola; Stroik, Jessica; Quarta, Davide; Yasar, Sevil; Müller, Christa E; Franco, Rafael; Goldberg, Steven R

    2011-07-01

    Several recent studies suggest functional and molecular interactions between striatal adenosine A(2A) and cannabinoid CB(1) receptors. Here, we demonstrate that A(2A) receptors selectively modulate reinforcing effects of cannabinoids. We studied effects of A(2A) receptor blockade on the reinforcing effects of delta-9-tetrahydrocannabinol (THC) and the endogenous CB(1) receptor ligand anandamide under a fixed-ratio schedule of intravenous drug injection in squirrel monkeys. A low dose of the selective adenosine A(2A) receptor antagonist MSX-3 (1 mg/kg) caused downward shifts of THC and anandamide dose-response curves. In contrast, a higher dose of MSX-3 (3 mg/kg) shifted THC and anandamide dose-response curves to the left. MSX-3 did not modify cocaine or food pellet self-administration. Also, MSX-3 neither promoted reinstatement of extinguished drug-seeking behavior nor altered reinstatement of drug-seeking behavior by non-contingent priming injections of THC. Finally, using in vivo microdialysis in freely-moving rats, a behaviorally active dose of MSX-3 significantly counteracted THC-induced, but not cocaine-induced, increases in extracellular dopamine levels in the nucleus accumbens shell. The significant and selective results obtained with the lower dose of MSX-3 suggest that adenosine A(2A) antagonists acting preferentially at presynaptic A(2A) receptors might selectively reduce reinforcing effects of cannabinoids that lead to their abuse. However, the appearance of potentiating rather than suppressing effects on cannabinoid reinforcement at the higher dose of MSX-3 would likely preclude the use of such a compound as a medication for cannabis abuse. Adenosine A(2A) antagonists with more selectivity for presynaptic versus postsynaptic receptors could be potential medications for treatment of cannabis abuse. Addiction Biology © 2010 Society for the Study of Addiction. No claim to original US government works.

  12. The Combined Inhibitory Effect of the Adenosine A1 and Cannabinoid CB1 Receptors on cAMP Accumulation in the Hippocampus Is Additive and Independent of A1 Receptor Desensitization

    OpenAIRE

    Serpa, Andr?; Correia, Sara; Ribeiro, Joaquim A.; Sebasti?o, Ana M.; Cascalheira, Jos? F.

    2015-01-01

    Adenosine A1 and cannabinoid CB1 receptors are highly expressed in hippocampus where they trigger similar transduction pathways. We investigated how the combined acute activation of A1 and CB1 receptors modulates cAMP accumulation in rat hippocampal slices. The CB1 agonist WIN55212-2 (0.3?30??M) decreased forskolin-stimulated cAMP accumulation with an EC50 of 6.6 ? 2.7??M and an E max? of 31% ? 2%, whereas for the A1 agonist, N6-cyclopentyladenosine (CPA, 10?150?nM), an EC50 of 35 ? 19?nM, an...

  13. Consequence of dopamine D2 receptor blockade on the hyperphagic effect induced by cannabinoid CB1 and CB2 receptors in layers.

    Science.gov (United States)

    Khodadadi, M; Zendehdel, M; Baghbanzadeh, A; Babapour, V

    2017-10-01

    1. Endocannabinoids (ECBs) and their receptors play a regulatory function on several physiological processes such as feed-intake behaviour, mainly in the brain. This study was carried out in order to investigate the effects of the dopaminergic D1 and D2 receptors on CB1/CB2 ECB receptor-induced hyperphagia in 3-h feed-deprived neonatal layer chickens. 2. A total of 8 experiments were designed to explore the interplay of these two modulatory systems on feed intake in neonatal chickens. In Experiment 1, chickens were intracerebroventricular (ICV) injected with control solution, l-DOPA (levo-dihydroxyphenylalanine as precursor of dopamine; 125 nmol), 2-AG (2-arachidonoylglycerol as CB 1 receptor agonist; 2 µg) and co-administration of l-DOPA (125 nmol) plus 2-AG (2 µg). Experiments 2-4 were similar to Experiment 1 except birds were injected with either 6-OHDA (6-hydroxydopamine as dopamine synthesis inhibitor; 150 nmol), SCH23390 (D1 receptor antagonist; 5 nmol) and AMI-193 (D2 receptor antagonist; 5 nmol) instead of l-DOPA, respectively. Additionally, Experiments 5-8 followed the previous ones using the same dose of l-DOPA, 6-OHDA and dopamine antagonists except that birds were injected with CB65 (CB2 receptor agonist; 5 µg) instead of 2-AG. Coadministrations were at the same dose for each experiment. Cumulative feed intakes were measured until 120 min after each injection. 3. ICV administration of 6-OHDA and AMI-193 significantly attenuated 2-AG-induced hyperphagia. Interestingly, the hyperphagic effect of CB65 was significantly attenuated by administration of l-DOPA, whereas the administration of 6-OHDA and AMI-193 together amplified the hyperphagic effect of CB65. 4. It was concluded that cannabinoid-induced feeding behaviour is probably modulated by dopamine receptors in neonatal layer-type chickens. It seems that their interaction may be mediated by the D2-dopamine receptor.

  14. Modulación del tono vascular por la sobre-expresión de receptores a canabinoides CB1 y CB2 en arterioesclerosis inducida en ratas

    OpenAIRE

    Espinoza P., María Rosa

    2013-01-01

    La arteriosclerosis, es un proceso degenerativo responsable de la mayor parte de las enfermedades cardiovasculares, es una enfermedad compleja que se produce a partir de múltiples factores de riesgo. Se ha observado que los compuestos derivados de la planta cannabis sativa provocan efectos vasorelajantes por medio de receptores específicos CB1 y CB2. ACPA y JWH 133 son potentes agonistas a estos receptores, se desconoce el papel que desempeñan en la arteriosclerosis provocando una relajación ...

  15. Receptor-heteromer mediated regulation of endocannabinoid signaling in activated microglia. Role of CB1 and CB2 receptors and relevance for Alzheimer's disease and levodopa-induced dyskinesia.

    Science.gov (United States)

    Navarro, Gemma; Borroto-Escuela, Dasiel; Angelats, Edgar; Etayo, Íñigo; Reyes-Resina, Irene; Pulido-Salgado, Marta; Rodríguez-Pérez, Ana I; Canela, Enric I; Saura, Josep; Lanciego, José Luis; Labandeira-García, José Luis; Saura, Carlos A; Fuxe, Kjell; Franco, Rafael

    2018-01-01

    Endocannabinoids are important regulators of neurotransmission and, acting on activated microglia, they are postulated as neuroprotective agents. Endocannabinoid action is mediated by CB 1 and CB 2 receptors, which may form heteromeric complexes (CB 1 -CB 2 Hets) with unknown function in microglia. We aimed at establishing the expression and signaling properties of cannabinoid receptors in resting and LPS/IFN-γ-activated microglia. In activated microglia mRNA transcripts increased (2 fold for CB 1 and circa 20 fold for CB 2 ), whereas receptor levels were similar for CB 1 and markedly upregulated for CB 2 ; CB 1 -CB 2 Hets were also upregulated. Unlike in resting cells, CB 2 receptors became robustly coupled to G i in activated cells, in which CB 1 -CB 2 Hets mediated a potentiation effect. Hence, resting cells were refractory while activated cells were highly responsive to cannabinoids. Interestingly, similar results were obtained in cultures treated with ß-amyloid (Aß 1-42 ). Microglial activation markers were detected in the striatum of a Parkinson's disease (PD) model and, remarkably, in primary microglia cultures from the hippocampus of mutant β-amyloid precursor protein (APP Sw,Ind ) mice, a transgenic Alzheimer's disease (AD) model. Also of note was the similar cannabinoid receptor signaling found in primary cultures of microglia from APP Sw,Ind and in cells from control animals activated using LPS plus IFN-γ. Expression of CB 1 -CB 2 Hets was increased in the striatum from rats rendered dyskinetic by chronic levodopa treatment. In summary, our results showed sensitivity of activated microglial cells to cannabinoids, increased CB 1 -CB 2 Het expression in activated microglia and in microglia from the hippocampus of an AD model, and a correlation between levodopa-induced dyskinesia and striatal microglial activation in a PD model. Cannabinoid receptors and the CB 1 -CB 2 heteroreceptor complex in activated microglia have potential as targets in the

  16. Polymorphisms of the dopamine D4 receptor gene (DRD4 VNTR) and cannabinoid CB1 receptor gene (CNR1) are not strongly related to cue-reactivity after alcohol exposure

    NARCIS (Netherlands)

    Wildenberg, E. van den; Janssen, R.G.J.H.; Hutchison, K.E.; Breukelen, G.J.P. van; Wiers, R.W.H.J.

    2007-01-01

    Polymorphisms in the D4 dopamine receptor gene (DRD4) and the CB1 cannabinoid receptor gene (CNR1) have been associated with a differential response to alcohol after consumption. The goal of the present study was to investigate whether heavy drinkers with these polymorphisms would respond with

  17. Suppressing effect of COR659 on alcohol, sucrose, and chocolate self-administration in rats: involvement of the GABAB and cannabinoid CB1 receptors.

    Science.gov (United States)

    Maccioni, Paola; Colombo, Giancarlo; Lorrai, Irene; Zaru, Alessandro; Carai, Mauro A M; Gessa, Gian Luigi; Brizzi, Antonella; Mugnaini, Claudia; Corelli, Federico

    2017-09-01

    COR659 [methyl2-(4-chlorophenylcarboxamido)-4-ethyl-5-methylthiophene-3-carboxylate] is a new, positive allosteric modulator (PAM) of the GABA B receptor. This study evaluated whether COR659 shared with previously tested GABA B PAMs the capacity to reduce alcohol self-administration in rats. Treatment with non-sedative doses of COR659 (2.5, 5, and 10 mg/kg; i.p.) suppressed lever-responding for alcohol (15% v/v) in Sardinian alcohol-preferring (sP) rats under the fixed ratio (FR) 4 (FR4) and progressive ratio (PR) schedules of reinforcement; COR659 was more potent and effective than the reference GABA B PAM, GS39783. Treatment with COR659, but not GS39783, suppressed (a) lever-responding for a sucrose solution (1-3% w/v) in sP rats under the FR4 and PR schedules, (b) lever-responding for a chocolate solution [5% (w/v) Nesquik®] in Wistar rats under the FR10 and PR schedules, and (c) cue-induced reinstatement of chocolate seeking in Wistar rats. Treatment with COR659 was completely ineffective on lever-responding (FR10) for regular food pellets in food-deprived Wistar rats. Pretreatment with the GABA B receptor antagonist, SCH50911, partially blocked COR659-induced reduction of alcohol self-administration, being ineffective on reduction of chocolate self-administration. Pretreatment with the cannabinoid CB 1 receptor antagonist, AM4113, fully blocked COR659-induced reduction of chocolate self-administration, being ineffective on reduction of alcohol self-administration. COR659 might exert its behavioral effects via a composite mechanism: (i) positive allosteric modulation of the GABA B receptor, responsible for a large proportion of reduction of alcohol self-administration; (ii) an action at other receptor system(s), including the cannabinoid CB 1 receptor, through which COR659 affects seeking and consumption of highly palatable foods.

  18. Effects of cannabinoid CB(1) receptor agonism and antagonism on SKF81297-induced dyskinesia and haloperidol-induced dystonia in Cebus apella monkeys

    DEFF Research Database (Denmark)

    Madsen, Morten V; Peacock, Linda P; Werge, Thomas

    2011-01-01

    81297 (SKF) and acute dystonia induced by the dopamine D(2) receptor antagonist haloperidol in Cebus apella monkeys. The monkeys were sensitised to EPS by prior exposure to D(2) receptor antagonists. SKF (0.3 mg/kg) was administered alone and in combination with the CB(1) agonist CP55,940 (0.......0025-0.01 mg/kg) or the CB(1) antagonist SR141716A (0.25-0.75 mg/kg). Haloperidol (individual doses at 0.01-0.02 mg/kg) was administered alone and in combination with CP55,940 (0.005 or 0.01 mg/kg) or SR141716A (0.5 or 0.75 mg/kg). Subsequently, the monkeys were videotaped, and the recordings were rated...... for oral dyskinesia or dystonia. SKF-induced oral dyskinesia was dose-dependently reduced by CP55,940, with no effect of SR141716A. Haloperidol-induced dystonia was not affected by either CP55,940 or SR141716A....

  19. Pharmacodynamic and pharmacokinetic effects of the intravenous CB1 receptor agonist Org 26828 in healthy male volunteers.

    Science.gov (United States)

    Zuurman, Lineke; Passier, Paul C C M; de Kam, Marieke L; Kleijn, Huub J; Cohen, Adam F; van Gerven, Joop M A

    2010-11-01

    An ideal drug for outpatient treatments under conscious sedation would have both sedative and analgesic properties. CB1/CB2 agonists are expected to have sedative, amnestic, analgesic and anti-emetic properties. The main objective of this first study in humans was to assess the sedative properties of intravenous Org 26828. In addition, pharmacokinetics, amnestic properties, postural stability, and behavioural and cardiovascular effects were studied. Midazolam intravenous 0.1 mg/kg and placebo were used as controls. The pharmacokinetic parameters (Cmax and AUC0-inf) of the main metabolite Org 26761 were proportional to dose. No effects were observed after doses up to 0.3 μg/kg of Org 26828. Dose-related effects were observed at higher doses. Although subjects reported subjective sedation after administration of Org 26828 at 3 and 6 μg/kg, the observed sedation was considerably less than after midazolam. Doses higher than the maximum tolerated dose of 1 μg/kg of Org 26828 caused unpleasant central nervous system effects (anxiety, paranoia, hallucinations). Therefore, Org 26828 is not suitable for providing sedation for outpatient surgical procedures.

  20. Maternal deprivation and adolescent cannabinoid exposure impact hippocampal astrocytes, CB1 receptors and brain-derived neurotrophic factor in a sexually dimorphic fashion.

    Science.gov (United States)

    López-Gallardo, M; López-Rodríguez, A B; Llorente-Berzal, Á; Rotllant, D; Mackie, K; Armario, A; Nadal, R; Viveros, M-P

    2012-03-01

    We have recently reported that early maternal deprivation (MD) for 24 h [postnatal day (PND) 9-10] and/or an adolescent chronic treatment with the cannabinoid agonist CP-55,940 (CP) [0.4 mg/kg, PND 28-42] in Wistar rats induced, in adulthood, diverse sex-dependent long-term behavioral and physiological modifications. Here we show the results obtained from investigating the immunohistochemical analysis of CB1 cannabinoid receptors, glial fibrillary acidic protein (GFAP) positive (+) cells and brain-derived neurotrophic factor (BDNF) expression in the hippocampus of the same animals. MD induced, in males, a significant increase in the number of GFAP+ cells in CA1 and CA3 areas and in the polymorphic layer of the dentate gyrus (DG), an effect that was attenuated by CP in the two latter regions. Adolescent cannabinoid exposure induced, in control non-deprived males, a significant increase in the number of GFAP+ cells in the polymorphic layer of the DG. MD induced a decrease in CB1 expression in both sexes, and this effect was reversed in males by the cannabinoid treatment. In turn, the drug "per se" induced, in males, a general decrease in CB1 immunoreactivity, and the opposite effect was observed in females. Cannabinoid exposure tended to reduce BDNF expression in CA1 and CA3 of females, whereas MD counteracted this trend and induced an increase of BDNF in females. As a whole, the present results show sex-dependent long-term effects of both MD and juvenile cannabinoid exposure as well as functional interactions between the two treatments. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

  1. MATERNAL DEPRIVATION AND ADOLESCENT CANNABINOID EXPOSURE IMPACT HIPPOCAMPAL ASTROCYTES, CB1 RECEPTORS AND BRAIN-DERIVED NEUROTROPHIC FACTOR IN A SEXUALLY DIMORPHIC FASHION

    Science.gov (United States)

    LÓPEZ-GALLARDO, M.; LÓPEZ-RODRÍGUEZ, A. B.; LLORENTE-BERZAL, Á.; ROTLLANT, D.; MACKIE, K.; ARMARIO, A.; NADAL, R.; VIVEROS, M.-P.

    2013-01-01

    We have recently reported that early maternal deprivation (MD) for 24 h [postnatal day (PND) 9–10] and/or an adolescent chronic treatment with the cannabinoid agonist CP-55,940 (CP) [0.4 mg/kg, PND 28–42] in Wistar rats induced, in adulthood, diverse sex-dependent long-term behavioral and physiological modifications. Here we show the results obtained from investigating the immunohistochemical analysis of CB1 cannabinoid receptors, glial fibrillary acidic protein (GFAP) positive (+) cells and brain-derived neurotrophic factor (BDNF) expression in the hippocampus of the same animals. MD induced, in males, a significant increase in the number of GFAP+ cells in CA1 and CA3 areas and in the polymorphic layer of the dentate gyrus (DG), an effect that was attenuated by CP in the two latter regions. Adolescent cannabinoid exposure induced, in control non-deprived males, a significant increase in the number of GFAP+ cells in the polymorphic layer of the DG. MD induced a decrease in CB1 expression in both sexes, and this effect was reversed in males by the cannabinoid treatment. In turn, the drug “per se” induced, in males, a general decrease in CB1 immunoreactivity, and the opposite effect was observed in females. Cannabinoid exposure tended to reduce BDNF expression in CA1 and CA3 of females, whereas MD counteracted this trend and induced an increase of BDNF in females. As a whole, the present results show sex-dependent long-term effects of both MD and juvenile cannabinoid exposure as well as functional interactions between the two treatments. PMID:22001306

  2. Prevention of drug priming- and cue-induced reinstatement of MDMA-seeking behaviors by the CB1 cannabinoid receptor antagonist AM251.

    Science.gov (United States)

    Nawata, Yoko; Kitaichi, Kiyoyuki; Yamamoto, Tsuneyuki

    2016-03-01

    3,4-Methylenedioxymethamphetamine (MDMA), a methamphetamine (METH) derivative, exhibits METH-like actions at monoamine transporters and positive reinforcing effects in rodents and primates. The purposes of the present study were to determine whether cross-reinstatement would be observed between MDMA and METH and if the cannabinoid receptor, a receptor known to play critical roles in the brain reward system, could modulate MDMA craving. Rats were trained to press a lever for intravenous MDMA (0.3mg/infusion) or METH (0.02mg/infusion) infusions under a fixed ratio 1 schedule paired with drug-associated cues (light and tone). Following drug self-administration acquisition training, rats underwent extinction training (an infusion of saline). Reinstatement tests were performed once the extinction criteria were achieved. In MDMA-trained rats, the MDMA-priming injection (3.2mg/kg, i.p.) or re-exposure to MDMA-associated cues reinstated MDMA-seeking behavior. Additionally, a priming injection of METH (1.0mg/kg, i.p.) also reinstated MDMA-seeking behavior. In contrast, none of the MDMA doses reinstated METH-seeking behavior in the METH-trained rats. The CB1 cannabinoid receptor antagonist AM251 markedly attenuated the MDMA-seeking behaviors induced by MDMA-priming injection or re-exposure to MDMA-associated cues in a dose-dependent manner. These findings show that MDMA has obvious addictive potential for reinstating drug-seeking behavior and that METH can be an effective stimulus for reinstating MDMA-seeking behaviors. Furthermore, based on the attenuating effect of AM251 in the reinstatement of MDMA-seeking behaviors, drugs that suppress CB1 receptors may be used in treatment of MDMA dependence. Copyright © 2016. Published by Elsevier Ireland Ltd.

  3. Delta-9-tetrahydrocannabinol differentially suppresses cisplatin-induced emesis and indices of motor function via cannabinoid CB(1) receptors in the least shrew.

    Science.gov (United States)

    Darmani, N A

    2001-01-01

    We have recently shown that the cannabinoid CB(1) receptor antagonist, SR 141716A, produces emesis in the least shrew (Cryptotis parva) in a dose- and route-dependent manner. This effect was blocked by delta-9-tetrahydrocannabinol (Delta(9)-THC). The present study investigates the cannabinoid receptor mechanisms by which Delta(9)-THC produces its antiemetic effects against cisplatin (20 mg/kg, i.p.)-induced emesis as well as its cannabimimetic activity profile (motor reduction) in the least shrew. Intraperitoneal administration of Delta(9)-THC (1, 2.5, 5 and 10 mg/kg) dose-dependently reduced both the percentage of animals vomiting (ID(50)=1.8+/-1.6 mg/kg) and the frequency of vomits (ID(50)=0.36+/-1.18 mg/kg) in a potent manner. The lowest significantly effective antiemetic dose of Delta(9)-THC for the latter emesis parameters was 2.5 mg/kg. Although Delta(9)-THC reduced the frequency of vomits up to 98%, it failed to completely protect all tested shrews from vomiting (80% protection). The cannabinoid CB(1) antagonist (SR 141716A) and not the CB(2) antagonist (SR 144528), reversed the antiemetic effects of Delta(9)-THC in a dose-dependent fashion. Delta(9)-THC (1, 5, 10 and 20 mg/kg, ip) suppressed locomotor parameters (spontaneous locomotor activity, duration of movement and rearing frequency) in a biphasic manner and only the 20-mg/kg dose simultaneously suppressed the triad of locomotor parameters to a significant degree. Subcutaneous (1-10 mg/kg) and intraperitoneal (0.05-40 mg/kg) injection of some doses of SR 141716A caused significant reductions in one or more components of the triad of locomotor parameters but these reductions were not dose dependent. Subcutaneous injection of SR 141716A (0.2, 1, 5 and 10 mg/kg) reversed the motor suppressant effects of a 20-mg/kg dose of Delta(9)-THC (ip) in a dose-dependent manner. Relative to its motor suppressant effects, Delta(9)-THC is a more potent antiemetic agent. Both effects are probably mediated via CB(1

  4. Enhanced self-administration of the CB1 receptor agonist WIN55,212-2 in olfactory bulbectomized rats: evaluation of possible serotonergic and dopaminergic underlying mechanisms

    Directory of Open Access Journals (Sweden)

    Petra eAmchova

    2014-03-01

    Full Text Available Depression has been associated with drug consumption, including heavy or problematic cannabis use. According to an animal model of depression and substance use disorder comorbidity, we combined the olfactory bulbectomy model of depression with intravenous drug self-administration procedure to verify whether depressive-like rats displayed higher voluntary intake of the CB1 receptor agonist WIN55,212-2 (WIN, 12.5 µg/kg/infusion. To this aim, olfactory-bulbectomized (OBX and sham-operated (SHAM Lister Hooded rats were allowed to self-administer WIN by lever-pressing under a continuous (FR-1 schedule of reinforcement in 2h daily sessions. Data showed that both OBX and SHAM rats developed stable WIN intake; yet, responses in OBX were constantly higher than in SHAM rats soon after the first week of training. In addition, OBX rats took significantly longer to extinguish the drug-seeking behaviour after vehicle substitution. Acute pre-treatment with serotonin 5HT1B receptor agonist, CGS-12066B (2.5-10 mg/kg, did not significantly modify WIN intake in OBX and SHAM Lister Hooded rats. Furthermore, acute pre-treatment with CGS-12066B (10 and 15 mg/kg did not alter responses in parallel groups of OBX and SHAM Sprague Dawley rats self-administering methamphetamine under higher (FR-2 reinforcement schedule with nose-poking as operandum. Finally, dopamine levels in the nucleus accumbens of OBX rats did not increase in response to a WIN challenge, as in SHAM rats, indicating a dopaminergic dysfunction in bulbectomized rats. Altogether, our findings suggest that a depressive state may alter cannabinoid CB1 receptor agonist-induced brain reward function and that a dopaminergic rather than a 5-HT1B mechanism is likely to underlie enhanced WIN self-administration in OBX rats.

  5. Enhanced novelty-induced corticosterone spike and upregulated serotonin 5-HT1A and cannabinoid CB1 receptors in adolescent BTBR mice.

    Science.gov (United States)

    Gould, Georgianna G; Burke, Teresa F; Osorio, Miguel D; Smolik, Corey M; Zhang, Wynne Q; Onaivi, Emmanuel S; Gu, Ting-Ting; DeSilva, Mauris N; Hensler, Julie G

    2014-01-01

    Hypothalamic pituitary adrenal (HPA) axis responses to change and social challenges during adolescence can influence mental health and behavior into adulthood. To examine how HPA tone in adolescence may contribute to psychopathology, we challenged male adolescent (5 weeks) and adult (16 weeks) BTBR T(+)tf/J (BTBR) and 129S1/SvImJ (129S) mice with novelty in sociability tests. In prior studies these strains had exaggerated or altered HPA stress responses and low sociability relative to C57BL/6J mice in adulthood. In adolescence these strains already exhibited similar or worse sociability deficits than adults or age-matched C57 mice. Yet BTBR adolescents were less hyperactive and buried fewer marbles than adults. Novelty-induced corticosterone (CORT) spikes in adolescent BTBR were double adult levels, and higher than 129S or C57 mice at either age. Due to their established role in HPA feedback, we hypothesized that hippocampal Gαi/o-coupled serotonin 5-HT1A and cannabinoid CB1 receptor function might be upregulated in BTBR mice. Adolescent BTBR mice had higher hippocampal 5-HT1A density as measured by [(3)H] 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) binding than C57 mice, and adult BTBR 8-OH-DPAT-stimulated GTPγS binding was higher than in either C57 or 129S mice in this region. Further, BTBR hippocampal CB1 density measured by [(3)H]CP55,940 binding was 15-20% higher than in C57. CP55,940-stimulated GTPγS binding in adult BTBR dentate gyrus was 30% higher then 129S (p<0.05), but was not a product of greater neuronal or cell density defined by NeuN and DAPI staining. Hence hyperactive HPA responsiveness during adolescence may underlie 5-HT1A and CB1 receptor up-regulation and behavioral phenotype of BTBR mice. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Cannabinoid CB1 /CB2 receptor agonists attenuate hyperactivity and body weight loss in a rat model of activity-based anorexia.

    Science.gov (United States)

    Scherma, Maria; Satta, Valentina; Collu, Roberto; Boi, Maria Francesca; Usai, Paolo; Fratta, Walter; Fadda, Paola

    2017-08-01

    Anorexia nervosa (AN) is a serious psychiatric condition characterized by excessive body weight loss and disturbed perceptions of body shape and size, often associated with excessive physical activity. There is currently no effective drug-related therapy of this disease and this leads to high relapse rate. Clinical data suggest that a promising therapy to treat and reduce reoccurrence of AN may be based on the use of drugs that target the endocannabinoid (EC) system, which appears dysregulated in AN patients. The activity-based anorexia (ABA) rodent model mimics the severe body weight loss and increased physical activity, as well as the neuroendocrine disturbances (i.e. hypoleptinaemia and hypercortisolaemia) in AN. This study investigated whether cannabinoid agonists can effectively modify anorexic-like behaviours and neuroendocrine changes in rats subjected to a repeated ABA regime that mimics the human condition in which patients repeatedly undergo a recovery and illness cycle. Our data show that subchronic treatment with both the natural CB 1 /CB 2 receptor agonist Δ 9 -tetrahydrocannabinol and the synthetic CB 1 /CB 2 receptor agonist CP-55,940 significantly reduced body weight loss and running wheel activity in ABA rats. These behavioural effects were accompanied by an increase in leptin signalling and a decrease in plasma levels of corticosterone. Taken together, our results further demonstrate the involvement of the EC system in AN pathophysiology and that strategies which modulate EC signalling are useful to treat this disorder, specifically in patients where physical hyperactivity plays a central role in its progression and maintenance. © 2017 The British Pharmacological Society.

  7. Cannabinoid Regulation of Brain Reward Processing with an Emphasis on the Role of CB1 Receptors: A Step Back into the Future.

    Science.gov (United States)

    Panagis, George; Mackey, Brian; Vlachou, Styliani

    2014-01-01

    Over the last decades, the endocannabinoid system has been implicated in a large variety of functions, including a crucial modulation of brain-reward circuits and the regulation of motivational processes. Importantly, behavioral studies have shown that cannabinoid compounds activate brain reward mechanisms and circuits in a similar manner to other drugs of abuse, such as nicotine, alcohol, cocaine, and heroin, although the conditions under which cannabinoids exert their rewarding effects may be more limited. Furthermore, there is evidence on the involvement of the endocannabinoid system in the regulation of cue- and drug-induced relapsing phenomena in animal models. The aim of this review is to briefly present the available data obtained using diverse behavioral experimental approaches in experimental animals, namely, the intracranial self-stimulation paradigm, the self-administration procedure, the conditioned place preference procedure, and the reinstatement of drug-seeking behavior procedure, to provide a comprehensive picture of the current status of what is known about the endocannabinoid system mechanisms that underlie modification of brain-reward processes. Emphasis is placed on the effects of cannabinoid 1 (CB1) receptor agonists, antagonists, and endocannabinoid modulators. Further, the role of CB1 receptors in reward processes is investigated through presentation of respective genetic ablation studies in mice. The vast majority of studies in the existing literature suggest that the endocannabinoid system plays a major role in modulating motivation and reward processes. However, much remains to be done before we fully understand these interactions. Further research in the future will shed more light on these processes and, thus, could lead to the development of potential pharmacotherapies designed to treat reward-dysfunction-related disorders.

  8. Cannabinoid regulation of brain reward processing with an emphasis on the role of CB1 receptors: a step back into the future

    Directory of Open Access Journals (Sweden)

    George ePanagis

    2014-07-01

    Full Text Available Over the last decades the endocannabinoid system has been implicated in a large variety of functions, including a crucial modulation of brain reward circuits and the regulation of motivational processes. Importantly, behavioural studies have shown that cannabinoid compounds activate brain reward mechanisms and circuits in a similar manner to other drugs of abuse, such as nicotine, alcohol, cocaine and heroin, although the conditions under which cannabinoids exert their rewarding effects may be more limited. Furthermore, there is evidence on the involvement of the endocannabinoid system in the regulation of cue- and drug-induced relapsing phenomena in animal models. The aim of this review is to briefly present the available data obtained using diverse behavioural experimental approaches in experimental animals, namely, the intracranial self-stimulation paradigm, the self-administration procedure, the conditioned place preference procedure and the reinstatement of drug-seeking behaviour procedure, to provide a comprehensive picture of the current status of what is known about the endocannabinoid system mechanisms that underlie modification of brain reward processes. Emphasis is placed on the effects of cannabinoid 1 (CB1 receptor agonists, antagonists and endocannabinoid modulators. Further, the role of CB1 receptors in reward processes is investigated through presentation of respective genetic ablation studies in mice. The vast majority of studies in the existing literature suggests that the endocannabinoid system plays a major role in modulating motivation and reward processes. However, much remains to be done before we fully understand these interactions. Further research in the future will shed more light on these processes and, thus, could lead to the development of potential pharmacotherapies designed to treat reward-dysfunction related disorders.

  9. CB1 and CB2 Receptors are Novel Molecular Targets for Tamoxifen and 4OH-Tamoxifen

    OpenAIRE

    Prather, Paul L.; FrancisDevaraj, FeAna; Dates, Centdrika R.; Greer, Aleksandra K.; Bratton, Stacie M.; Ford, Benjamin M.; Franks, Lirit N.; Radominska-Pandya, Anna

    2013-01-01

    Tamoxifen (Tam) is classified as a selective estrogen receptor modulator (SERM) and is used for treatment of patients with ER-positive breast cancer. However, it has been shown that Tam and its cytochrome P450-generated metabolite 4-hydroxy-Tam (4OH-Tam) also exhibit cytotoxic effects in ER-negative breast cancer cells. These observations suggest that Tam and 4OH-Tam can produce cytotoxicity via estrogen receptor (ER)-independent mechanism(s) of action. The molecular targets responsible for t...

  10. Involvement of the cannabinoid CB1 receptor in modulation of dopamine output in the prefrontal cortex associated with food restriction in rats.

    Science.gov (United States)

    Dazzi, Laura; Talani, Giuseppe; Biggio, Francesca; Utzeri, Cinzia; Lallai, Valeria; Licheri, Valentina; Lutzu, Stefano; Mostallino, Maria Cristina; Secci, Pietro Paolo; Biggio, Giovanni; Sanna, Enrico

    2014-01-01

    Increase in dopamine output on corticolimbic structures, such as medial prefrontal cortex (mPFC) and nucleus accumbens, has been related to reward effects associated with palatable food or food presentation after a fasting period. The endocannabinoid system regulates feeding behavior through a modulatory action on different neurotransmitter systems, including the dopaminergic system. To elucidate the involvement of type 1 cannabinoid receptors in the regulation of dopamine output in the mPFC associated with feeding in hungry rats, we restricted the food availability to a 2-h period daily for 3 weeks. In food-restricted rats the extracellular dopamine concentration in the mPFC increased starting 80 min before food presentation and returned to baseline after food removal. These changes were attenuated in animals treated with the CB1 receptor antagonist SR141716. To better understand how food restriction can change the response of mesocortical dopaminergic neurons, we studied several components of the neuronal circuit that regulates dopamine output in the mPFC. Patch-clamp experiments revealed that the inhibitory effect of the CB1 receptor agonist WIN 55,212-2 on GABAergic sIPSC frequency was diminished in mPFC neurons of FR compared to fed ad libitum rats. The basal sIPSC frequency resulted reduced in mPFC neurons of food-restricted rats, suggestive of an altered regulation of presynaptic GABA release; these changes were accompanied by an enhanced excitability of mPFC and ventral tegmental area neurons. Finally, type 1 cannabinoid receptor expression in the mPFC was reduced in food-restricted rats. Together, our data support an involvement of the endocannabinoid system in regulation of dopamine release in the mPFC through changes in GABA inhibitory synapses and suggest that the emphasized feeding-associated increase in dopamine output in the mPFC of food-restricted rats might be correlated with an altered expression and function of type 1 cannabinoid receptor in this

  11. Involvement of the cannabinoid CB1 receptor in modulation of dopamine output in the prefrontal cortex associated with food restriction in rats.

    Directory of Open Access Journals (Sweden)

    Laura Dazzi

    Full Text Available Increase in dopamine output on corticolimbic structures, such as medial prefrontal cortex (mPFC and nucleus accumbens, has been related to reward effects associated with palatable food or food presentation after a fasting period. The endocannabinoid system regulates feeding behavior through a modulatory action on different neurotransmitter systems, including the dopaminergic system. To elucidate the involvement of type 1 cannabinoid receptors in the regulation of dopamine output in the mPFC associated with feeding in hungry rats, we restricted the food availability to a 2-h period daily for 3 weeks. In food-restricted rats the extracellular dopamine concentration in the mPFC increased starting 80 min before food presentation and returned to baseline after food removal. These changes were attenuated in animals treated with the CB1 receptor antagonist SR141716. To better understand how food restriction can change the response of mesocortical dopaminergic neurons, we studied several components of the neuronal circuit that regulates dopamine output in the mPFC. Patch-clamp experiments revealed that the inhibitory effect of the CB1 receptor agonist WIN 55,212-2 on GABAergic sIPSC frequency was diminished in mPFC neurons of FR compared to fed ad libitum rats. The basal sIPSC frequency resulted reduced in mPFC neurons of food-restricted rats, suggestive of an altered regulation of presynaptic GABA release; these changes were accompanied by an enhanced excitability of mPFC and ventral tegmental area neurons. Finally, type 1 cannabinoid receptor expression in the mPFC was reduced in food-restricted rats. Together, our data support an involvement of the endocannabinoid system in regulation of dopamine release in the mPFC through changes in GABA inhibitory synapses and suggest that the emphasized feeding-associated increase in dopamine output in the mPFC of food-restricted rats might be correlated with an altered expression and function of type 1

  12. Preparation of iodine-123 labeled AM251: a potential SPECT radioligand for the brain cannabinoid CB1 receptor

    International Nuclear Information System (INIS)

    Lan, Ruoxi; Makriyannis, Alexandros; Gatley, S.J.

    1996-01-01

    We report the synthesis and labeling with iodine-123 of N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251). This compound is an analog of the recently described cannabinoid receptor antagonist, SR141716A, in which a 4-chlorophenyl group is replaced by 4-iodophenyl. Labeling in good yield (62%) and radiochemical purity (> 95%), and high specific activity (> 2500 Ci/mmol) was achieved by an iododestannylation reaction using the tributyltin precursor, no carrier added I-123 iodide, and chloramine-T. (author)

  13. Preparation of iodine-123 labeled AM251: a potential SPECT radioligand for the brain cannabinoid CB1 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Lan, Ruoxi; Makriyannis, Alexandros [Connecticut Univ., Molecular and Cell Biology Dept., Storrs, CT (United States); Gatley, S.J. [Brookhaven National Lab., Medical Dept., Upton, NY (United States)

    1996-10-01

    We report the synthesis and labeling with iodine-123 of N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251). This compound is an analog of the recently described cannabinoid receptor antagonist, SR141716A, in which a 4-chlorophenyl group is replaced by 4-iodophenyl. Labeling in good yield (62%) and radiochemical purity (> 95%), and high specific activity (> 2500 Ci/mmol) was achieved by an iododestannylation reaction using the tributyltin precursor, no carrier added I-123 iodide, and chloramine-T. (author).

  14. Localización de los Receptores Cannabinoides 1 (CB1) en Glándulas Salivales Submandibulares y Sublinguales de Ratones Durante el Desarrollo Postnatal

    OpenAIRE

    Hipkaeo, Wiphawi; Watanabe, Masahiko; Kondo, Hisatake

    2015-01-01

    In view of the fact that human marijuana users often show dry mouth symptom, the present study was attempted to examine the localization of CB1, which was originally identified in brain, in the submandibular and sublingual salivary glands of postnatal developing male mice by immunohistochemistry. In submandibular gland, CB1-immunoreactivity was positive in a majority of acinar cells in forms of granular appearance in their apical cytoplasm, while it was negative in the ducts at newborn stage....

  15. Native CB1 receptor affinity, intrinsic activity and accumbens shell dopamine stimulant properties of third generation SPICE/K2 cannabinoids: BB-22, 5F-PB-22, 5F-AKB-48 and STS-135.

    Science.gov (United States)

    De Luca, Maria Antonietta; Castelli, M Paola; Loi, Barbara; Porcu, Alessandra; Martorelli, Mariella; Miliano, Cristina; Kellett, Kathryn; Davidson, Colin; Stair, Jacqueline L; Schifano, Fabrizio; Di Chiara, Gaetano

    2016-06-01

    In order to investigate the in vivo dopamine (DA) stimulant properties of selected 3rd generation Spice/K2 cannabinoids, BB-22, 5F-PB-22, 5F-AKB-48 and STS-135, their in vitro affinity and agonist potency at native rat and mice CB1 receptors was studied. The compounds bind with high affinity to CB1 receptors in rat cerebral cortex homogenates and stimulate CB1-induced [(35)S]GTPγS binding with high potency and efficacy. BB-22 and 5F-PB-22 showed the lowest Ki of binding to CB1 receptors (0.11 and 0.13 nM), i.e., 30 and 26 times lower respectively than that of JWH-018 (3.38 nM), and a potency (EC50, 2.9 and 3.7 nM, respectively) and efficacy (Emax, 217% and 203%, respectively) as CB1 agonists higher than JWH-018 (EC50, 20.2 nM; Emax, 163%). 5F-AKB-48 and STS-135 had higher Ki for CB1 binding, higher EC50 and lower Emax as CB1 agonists than BB-22 and 5F-PB-22 but still comparatively more favourable than JWH-018. The agonist properties of all the compounds were abolished or drastically reduced by the CB1 antagonist/inverse agonist AM251 (0.1 μM). No activation of G-protein was observed in CB1-KO mice. BB-22 (0.003-0.01 mg/kg i.v.) increased dialysate DA in the accumbens shell but not in the core or in the medial prefrontal cortex, with a bell shaped dose-response curve and an effect at 0.01 mg/kg and a biphasic time-course. Systemic AM251 (1.0 mg/kg i.p.) completely prevented the stimulant effect of BB-22 on dialysate DA in the NAc shell. All the other compounds increased dialysate DA in the NAc shell at doses consistent with their in vitro affinity for CB1 receptors (5F-PB-22, 0.01 mg/kg; 5F-AKB-48, 0.1 mg/kg; STS-135, 0.15 mg/kg i.v.). 3rd generation cannabinoids can be even more potent and super-high CB1 receptor agonists compared to JWH-018. Future research will try to establish if these properties can explain the high toxicity and lethality associated with these compounds. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. A Feedforward Inhibitory Circuit Mediated by CB1-Expressing Fast-Spiking Interneurons in the Nucleus Accumbens.

    Science.gov (United States)

    Wright, William J; Schlüter, Oliver M; Dong, Yan

    2017-04-01

    The nucleus accumbens (NAc) gates motivated behaviors through the functional output of principle medium spiny neurons (MSNs), whereas dysfunctional output of NAc MSNs contributes to a variety of psychiatric disorders. Fast-spiking interneurons (FSIs) are sparsely distributed throughout the NAc, forming local feedforward inhibitory circuits. It remains elusive how FSI-based feedforward circuits regulate the output of NAc MSNs. Here, we investigated a distinct subpopulation of NAc FSIs that express the cannabinoid receptor type-1 (CB1). Using a combination of paired electrophysiological recordings and pharmacological approaches, we characterized and compared feedforward inhibition of NAc MSNs from CB1 + FSIs and lateral inhibition from recurrent MSN collaterals. We observed that CB1 + FSIs exerted robust inhibitory control over a large percentage of nearby MSNs in contrast to local MSN collaterals that provided only sparse and weak inhibitory input to their neighboring MSNs. Furthermore, CB1 + FSI-mediated feedforward inhibition was preferentially suppressed by endocannabinoid (eCB) signaling, whereas MSN-mediated lateral inhibition was unaffected. Finally, we demonstrated that CB1 + FSI synapses onto MSNs are capable of undergoing experience-dependent long-term depression in a voltage- and eCB-dependent manner. These findings demonstrated that CB1 + FSIs are a major source of local inhibitory control of MSNs and a critical component of the feedforward inhibitory circuits regulating the output of the NAc.

  17. Electroacupuncture Inhibition of Hyperalgesia in Rats with Adjuvant Arthritis: Involvement of Cannabinoid Receptor 1 and Dopamine Receptor Subtypes in Striatum

    Directory of Open Access Journals (Sweden)

    Yin Shou

    2013-01-01

    Full Text Available Electroacupuncture (EA has been regarded as an alternative treatment for inflammatory pain for several decades. However, the molecular mechanisms underlying the antinociceptive effect of EA have not been thoroughly clarified. Previous studies have shown that cannabinoid CB1 receptors are related to pain relief. Accumulating evidence has shown that the CB1 and dopamine systems sometimes interact and may operate synergistically in rat striatum. To our knowledge, dopamine D1/D2 receptors are involved in EA analgesia. In this study, we found that repeated EA at Zusanli (ST36 and Kunlun (BL60 acupoints resulted in marked improvements in thermal hyperalgesia. Both western blot assays and FQ-PCR analysis results showed that the levels of CB1 expression in the repeated-EA group were much higher than those in any other group (P=0.001. The CB1-selective antagonist AM251 inhibited the effects of repeated EA by attenuating the increases in CB1 expression. The two kinds of dopamine receptors imparted different actions on the EA-induced CB1 upregulation in AA rat model. These results suggested that the strong activation of the CB1 receptor after repeated EA resulted in the concomitant phenomenon of the upregulation of D1 and D2 levels of gene expression.

  18. Targeting the Endocannabinoid/CB1 Receptor System For Treating Major Depression Through Antidepressant Activities of Curcumin and Dexanabinol-Loaded Solid Lipid Nanoparticles

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    Xiaolie He

    2017-08-01

    Full Text Available Background/Aims: This study investigated the underlying mechanisms of the antidepressant effects of curcumin and dexanabinol-loaded solid lipid nanoparticles in corticosterone-induced cell and mice depression models. Methods: Curcumin and dexanabinol-loaded solid lipid nanoparticles (Cur/SLNs-HU-211 were synthesized via an emulsifcation and low-temperature solidification method. Antidepressant activities of nanoparticles in a corticosterone-induced major depression model were investigated by MTT assay, cellular uptake by flow cytometry, behaviour by Forced Swimming Test and rotarod test, neurotransmitters by High Performance Liquid Chromatography, Western blotting, qPCR and immunofluorescence. Results: Treatment with Cur/SLNs-HU-211 induced greater dopamine (DA/5-hydroxytryptamine (5-HT release with reduced corticosterone-induced apoptotic cell death in PC12 cells. Additionally, in vivo Cur/SLNs-HU-211 significantly induced recovery from depressive behaviour with increased DA/5-HT levels, CB1 mRNA levels and CB1, p-MEK1 and p-ERK1/2 protein expression levels in the hippocampus and striatum. Cur/SLNs-HU-211 improved CB1 expression and inspired the proliferation of astrocytes in the hippocampus and striatum, exerted neuroprotective effects by preventing corticosterone -induced BDNF/NeuN expression reduction. Conclusion: Our study implies that Cur/SLNs-HU-211 may be a useful approach for treatment of major depression.

  19. Repeated administration of phytocannabinoid Δ(9)-THC or synthetic cannabinoids JWH-018 and JWH-073 induces tolerance to hypothermia but not locomotor suppression in mice, and reduces CB1 receptor expression and function in a brain region-specific manner.

    Science.gov (United States)

    Tai, S; Hyatt, W S; Gu, C; Franks, L N; Vasiljevik, T; Brents, L K; Prather, P L; Fantegrossi, W E

    2015-12-01

    These studies probed the relationship between intrinsic efficacy and tolerance/cross-tolerance between ∆(9)-THC and synthetic cannabinoid drugs of abuse (SCBs) by examining in vivo effects and cellular changes concomitant with their repeated administration in mice. Dose-effect relationships for hypothermic effects were determined in order to confirm that SCBs JWH-018 and JWH-073 are higher efficacy agonists than ∆(9)-THC in mice. Separate groups of mice were treated with saline, sub-maximal hypothermic doses of JWH-018 or JWH-073 (3.0mg/kg or 10.0mg/kg, respectively) or a maximally hypothermic dose of 30.0mg/kg ∆(9)-THC once per day for 5 consecutive days while core temperature and locomotor activity were monitored via biotelemetry. Repeated administration of all drugs resulted in tolerance to hypothermic effects, but not locomotor effects, and this tolerance was still evident 14 days after the last drug administration. Further studies treated mice with 30.0mg/kg ∆(9)-THC once per day for 4 days, then tested with SCBs on day 5. Mice with a ∆(9)-THC history were cross-tolerant to both SCBs, and this cross-tolerance also persisted 14 days after testing. Select brain regions from chronically treated mice were examined for changes in CB1 receptor expression and function. Expression and function of hypothalamic CB1Rs were reduced in mice receiving chronic drugs, but cortical CB1R expression and function were not altered. Collectively, these data demonstrate that repeated ∆(9)-THC, JWH-018 and JWH-073 can induce long-lasting tolerance to some in vivo effects, which is likely mediated by region-specific downregulation and desensitization of CB1Rs. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Beneficial metabolic effects of CB1R anti-sense oligonucleotide treatment in diet-induced obese AKR/J mice.

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    Yuting Tang

    Full Text Available An increasing amount of evidence supports pleiotropic metabolic roles of the cannibinoid-1 receptor (CB1R in peripheral tissues such as adipose, liver, skeletal muscle and pancreas. To further understand the metabolic consequences of specific blockade of CB1R function in peripheral tissues, we performed a 10-week-study with an anti-sense oligonucleotide directed against the CB1R in diet-induced obese (DIO AKR/J mice. DIO AKR/J mice were treated with CB1R ASO Isis-414930 (6.25, 12.5 and 25 mg/kg/week or control ASO Isis-141923 (25 mg/kg/week via intraperitoneal injection for 10 weeks. At the end of the treatment, CB1R mRNA from the 25 mg/kg/week CB1R ASO group in the epididymal fat and kidney was decreased by 81% and 63%, respectively. Body weight gain was decreased in a dose-dependent fashion, significantly different in the 25 mg/kg/week CB1R ASO group (46.1±1.0 g vs veh, 51.2±0.9 g, p<0.05. Body fat mass was reduced in parallel with attenuated body weight gain. CB1R ASO treatment led to decreased fed glucose level (at week 8, 25 mg/kg/week group, 145±4 mg/dL vs veh, 195±10 mg/dL, p<0.05. Moreover, CB1R ASO treatment dose-dependently improved glucose excursion during an oral glucose tolerance test, whereas control ASO exerted no effect. Liver steatosis was also decreased upon CB1R ASO treatment. At the end of the study, plasma insulin and leptin levels were significantly reduced by 25 mg/kg/week CB1R ASO treatment. SREBP1 mRNA expression was decreased in both epididymal fat and liver. G6PC and fatty acid translocase/CD36 mRNA levels were also reduced in the liver. In summary, CB1R ASO treatment in DIO AKR/J mice led to improved insulin sensitivity and glucose homeostasis. The beneficial effects of CB1R ASO treatment strongly support the notion that selective inhibition of the peripheral CB1R, without blockade of central CB1R, may serve as an effective approach for treating type II diabetes, obesity and the metabolic syndrome.

  1. Pharmacological blockade of either cannabinoid CB1 or CB2 receptors prevents both cocaine-induced conditioned locomotion and cocaine-induced reduction of cell proliferation in the hippocampus of adult male rat

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    Blanco-Calvo, Eduardo; Rivera, Patricia; Arrabal, Sergio; Vargas, Antonio; Pavón, Francisco Javier; Serrano, Antonia; Castilla-Ortega, Estela; Galeano, Pablo; Rubio, Leticia; Suárez, Juan; Rodriguez de Fonseca, Fernando

    2014-01-01

    Addiction to major drugs of abuse, such as cocaine, has recently been linked to alterations in adult neurogenesis in the hippocampus. The endogenous cannabinoid system modulates this proliferative response as demonstrated by the finding that pharmacological activation/blockade of cannabinoid CB1 and CB2 receptors not only modulates neurogenesis but also modulates cell death in the brain. In the present study, we evaluated whether the endogenous cannabinoid system affects cocaine-induced alterations in cell proliferation. To this end, we examined whether pharmacological blockade of either CB1 (Rimonabant, 3 mg/kg) or CB2 receptors (AM630, 3 mg/kg) would affect cell proliferation [the cells were labeled with 5-bromo-2′-deoxyuridine (BrdU)] in the subventricular zone (SVZ) of the lateral ventricle and the dentate subgranular zone (SGZ). Additionally, we measured cell apoptosis (as monitored by the expression of cleaved caspase-3) and glial activation [by analyzing the expression of glial fibrillary acidic protein (GFAP) and Iba-1] in the striatum and hippocampus during acute and repeated (4 days) cocaine administration (20 mg/kg). The results showed that acute cocaine exposure decreased the number of BrdU-immunoreactive (ir) cells in the SVZ and SGZ. In contrast, repeated cocaine exposure reduced the number of BrdU-ir cells only in the SVZ. Both acute and repeated cocaine exposure increased the number of cleaved caspase-3-, GFAP- and Iba1-ir cells in the hippocampus, and this effect was counteracted by AM630 or Rimonabant, which increased the number of BrdU-, GFAP-, and Iba1-ir cells in the hippocampus. These results indicate that the changes in neurogenic, apoptotic and gliotic processes that were produced by repeated cocaine administration were normalized by pharmacological blockade of CB1 and CB2. The restorative effects of cannabinoid receptor blockade on hippocampal cell proliferation were associated with the prevention of the induction of conditioned

  2. Molecular Understanding of the Activation of CB1 and Blockade of TRPV1 Receptors: Implications for Novel Treatment Strategies in Osteoarthritis

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    Jakub Mlost

    2018-01-01

    Full Text Available Osteoarthritis (OA is a joint disease in which cartilage degenerates as a result of mechanical and biochemical changes. The main OA symptom is chronic pain involving both peripheral and central mechanisms of nociceptive processing. Our previous studies have implicated the benefits of dual- over single-acting compounds interacting with the endocannabinoid system (ECS in OA treatment. In the present study, we focused on the specific molecular alterations associated with pharmacological treatment. OA was induced in Wistar rats by intra-articular injection of 3 mg of monoiodoacetate (MIA. Single target compounds (URB597, an FAAH inhibitor, and SB366791, a TRPV1 antagonist and a dual-acting compound OMDM198 (FAAH inhibitor/TRPV1 antagonist were used in the present study. At day 21 post-MIA injection, rats were sacrificed 1 h after i.p. treatment, and changes in mRNA expression were evaluated in the lumbar spinal cord by RT-qPCR. Following MIA administration, we observed 2-4-fold increase in mRNA expression of targeted receptors (Cnr1, Cnr2, and Trpv1, endocannabinoid degradation enzymes (Faah, Ptgs2, and Alox12, and TRPV1 sensitizing kinases (Mapk3, Mapk14, Prkcg, and Prkaca. OMDM198 treatment reversed some of the MIA effects on the spinal cord towards intact levels (Alox12, Mapk14, and Prkcg. Apparent regulation of ECS and TRPV1 in response to pharmacological intervention is a strong justification for novel ECS-based multi-target drug treatment in OA.

  3. CB1R-Mediated Activation of Caspase-3 Causes Epigenetic and Neurobehavioral Abnormalities in Postnatal Ethanol-Exposed Mice

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    Shivakumar Subbanna

    2018-02-01

    Full Text Available Alcohol exposure can affect brain development, leading to long-lasting behavioral problems, including cognitive impairment, which together is defined as fetal alcohol spectrum disorder (FASD. However, the fundamental mechanisms through which this occurs are largely unknown. In this study, we report that the exposure of postnatal day 7 (P7 mice to ethanol activates caspase-3 via cannabinoid receptor type-1 (CB1R in neonatal mice and causes a reduction in methylated DNA binding protein (MeCP2 levels. The developmental expression of MeCP2 in mice is closely correlated with synaptogenesis and neuronal maturation. It was shown that ethanol treatment of P7 mice enhanced Mecp2 mRNA levels but reduced protein levels. The genetic deletion of CB1R prevented, and administration of a CB1R antagonist before ethanol treatment of P7 mice inhibited caspase-3 activation. Additionally, it reversed the loss of MeCP2 protein, cAMP response element binding protein (CREB activation, and activity-regulated cytoskeleton-associated protein (Arc expression. The inhibition of caspase-3 activity prior to ethanol administration prevented ethanol-induced loss of MeCP2, CREB activation, epigenetic regulation of Arc expression, long-term potentiation (LTP, spatial memory deficits and activity-dependent impairment of several signaling molecules, including MeCP2, in adult mice. Collectively, these results reveal that the ethanol-induced CB1R-mediated activation of caspase-3 degrades the MeCP2 protein in the P7 mouse brain and causes long-lasting neurobehavioral deficits in adult mice. This CB1R-mediated instability of MeCP2 during active synaptic maturation may disrupt synaptic circuit maturation and lead to neurobehavioral abnormalities, as observed in this animal model of FASD.

  4. Endogenous vs Exogenous Allosteric Modulators in GPCRs: A dispute for shuttling CB1 among different membrane microenvironments

    Science.gov (United States)

    Stornaiuolo, Mariano; Bruno, Agostino; Botta, Lorenzo; Regina, Giuseppe La; Cosconati, Sandro; Silvestri, Romano; Marinelli, Luciana; Novellino, Ettore

    2015-10-01

    A Cannabinoid Receptor 1 (CB1) binding site for the selective allosteric modulator ORG27569 is here identified through an integrate approach of consensus pocket prediction, mutagenesis studies and Mass Spectrometry. This unprecedented ORG27569 pocket presents the structural features of a Cholesterol Consensus Motif, a cholesterol interacting region already found in other GPCRs. ORG27569 and cholesterol affects oppositely CB1 affinity for orthosteric ligands. Moreover, the rise in cholesterol intracellular level results in CB1 trafficking to the axonal region of neuronal cells, while, on the contrary, ORG27568 binding induces CB1 enrichment at the soma. This control of receptor migration among functionally different membrane regions of the cell further contributes to downstream signalling and adds a previously unknown mechanism underpinning CB1 modulation by ORG27569 , that goes beyond a mere control of receptor affinity for orthosteric ligands.

  5. The Structure–Function Relationships of Classical Cannabinoids: CB1/CB2 Modulation

    Science.gov (United States)

    Bow, Eric W.; Rimoldi, John M.

    2016-01-01

    The cannabinoids are members of a deceptively simple class of terpenophenolic secondary metabolites isolated from Cannabis sativa highlighted by (−)-Δ9-tetrahydrocannabinol (THC), eliciting distinct pharmacological effects mediated largely by cannabinoid receptor (CB1 or CB2) signaling. Since the initial discovery of THC and related cannabinoids, synthetic and semisynthetic classical cannabinoid analogs have been evaluated to help define receptor binding modes and structure–CB1/CB2 functional activity relationships. This perspective will examine the classical cannabinoids, with particular emphasis on the structure–activity relationship of five regions: C3 side chain, phenolic hydroxyl, aromatic A-ring, pyran B-ring, and cyclohexenyl C-ring. Cumulative structure–activity relationship studies to date have helped define the critical structural elements required for potency and selectivity toward CB1 and CB2 and, more importantly, ushered the discovery and development of contemporary nonclassical cannabinoid modulators with enhanced physicochemical and pharmacological profiles. PMID:27398024

  6. CB 1/2 dual agonists with 3-carbamoyl 2-pyridone derivatives as antipruritics: reduction of CNS side effects by introducing polar functional groups.

    Science.gov (United States)

    Odan, Masahide; Ishizuka, Natsuki; Hiramatsu, Yoshiharu; Inagaki, Masanao; Hashizume, Hiroshi; Fujii, Yasuhiko; Mitsumori, Susumu; Morioka, Yasuhide; Soga, Masahiko; Deguchi, Masashi; Yasui, Kiyoshi; Arimura, Akinori

    2012-04-15

    Our lead compound 1 showed high affinity for both CB1 and CB2 receptors, suggesting the possibility of inducing psychoactive side effects through the CB1 receptor in the brain. To solve this issue, polar functional groups were introduced at the 3-position of the pyridone core of compound 1 to find CB1/2 dual agonists such as 17 and 20 which did not show any CNS side effects. Copyright © 2012 Elsevier Ltd. All rights reserved.

  7. Het endocannabinoïdsysteem, overgewicht en de CB1-endocannabinoïdreceptorblokker rimonabant

    NARCIS (Netherlands)

    Boekholdt, S. M.; Jukema, J. W.; Peters, R. J. G.

    2007-01-01

    --Obesity is an important healthcare issue. --Recent research has led to insights into the role of the endocannabinoid system in the regulation of body weight. --Rimonabant is a CB1-endocannabinoid-receptor antagonist. --Four trials were published recently on the efficacy and safety of rimonabant in

  8. Leptin Receptor Deficiency is Associated With Upregulation of Cannabinoid 1 Receptors in Limbic Brain Regions

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    THANOS, PANAYOTIS K.; RAMALHETE, ROBERTO C.; MICHAELIDES, MICHAEL; PIYIS, YIANNI K.; WANG, GENE-JACK; VOLKOW, NORA D.

    2009-01-01

    Leptin receptor dysfunction results in overeating and obesity. Leptin regulates hypothalamic signaling that underlies the motivation to hyperphagia, but the interaction between leptin and cannabinoid signaling is poorly understood. We evaluated the role of cannabinoid 1 receptors (CB1R) in overeating and the effects of food deprivation on CB1R in the brain. One-month-old Zucker rats were divided into unrestricted and restricted (fed 70% of unrestricted rats) diet groups and maintained until adulthood (4 months). Levels of relative binding sites of CB1R (CB1R binding levels) were assessed using [3H] SR141716A in vitro autoradiography. These levels were higher (except cerebellum and hypothalamus) at 4 months than at 1 month of age. One month CB1R binding levels for most brain regions did not differ between Ob and Lean (Le) rats (except in frontal and cingulate cortices in Le and in the hypothalamus in Ob). Four month Ob rats had higher CB1R binding levels than Le in most brain regions and food restriction was associated with higher CB1R levels in all brain regions in Ob, but not in Le rats. CB1R binding levels increased between adolescence and young adulthood which we believe was influenced by leptin and food availability. The high levels of CB1R in Ob rats suggest that leptin's inhibition of food-intake is in part mediated by downregulation of CB1R and that leptin interferes with CB1R upregulation under food-deprivation conditions. These results are consistent with prior findings showing increased levels of endogenous cannabinoids in the Ob rats corroborating the regulation of cannabinoid signaling by leptin. PMID:18563836

  9. R+-methanandamide inhibits tracheal response to endogenously released acetylcholine via capsazepine-sensitive receptors.

    Science.gov (United States)

    Nieri, Paola; Martinotti, Enrica; Testai, Lara; Adinolfi, Barbara; Calderone, Vincenzo; Breschi, Maria Cristina

    2003-01-10

    The effects of cannabinoid drugs on the cholinergic response evoked by electrical field stimulation (0.2 ms pulse width, 20 V amplitude, 10 Hz, 7.5 s train duration) in guinea-pig tracheal preparations were investigated. The stable analogue of the endocannabinoid anandamide, R(+)-methanandamide (10(-7)-10(-4) M), produced a dose-dependent inhibition (up to 27+/-5% of control) of electrical field stimulation-mediated atropine-sensitive response. This effect was not blocked by the selective cannabinoid CB(1) receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3 carboxamide hydrochloride (SR 141716A; 10(-6) M), and was not reproduced with the cannabinoid CB(1)/CB(2) receptor agonist R(+)-[2,3-dihydro-5-methyl-[(morpholinyl)methyl]pyrrolo [1,2,3-de]-1,4-benzoxazin-6-yl]-(1-naphthalenyl)methanone mesylate) (WIN 55,212-2; 10(-8)-10(-5) M) or the cannabinoid CB(2) receptor selective agonist 1-propyl-2-methyl-3-(1-naphthoyl)indole (JWH-015; 10(-8)-10(-5) M); it was, on the contrary, antagonized by the vanilloid antagonist 2-[2-(4-chlorophenyl)ethyl-amino-thiocarbonyl]-7,8-dihydroxy-2,3,4,5-tetrahydro-1H-2 benzazepine (capsazepine; 10(-6) M). At the postjunctional level, neither R(+)-methanandamide nor WIN 55,212-2 nor JWH-015 did affect tracheal contractions induced by exogenous acetylcholine (10(-6) M). An inhibitory vanilloid receptor-mediated effect on the cholinergic response evoked by electrical stimulation was confirmed with the vanilloid agonist capsaicin, at doses (3-6 x 10(-8) M) which poorly influenced the basal smooth muscle tone of trachea. In conclusion, our data indicate that in guinea-pig trachea (a) neither CB(1) nor CB(2) cannabinoid receptor-mediated modulation of acetylcholine release occurs; (b) vanilloid VR1-like receptors appear involved in R(+)-methanandamide inhibitory activity on the cholinergic response to electrical field stimulation.

  10. Perspectives of CB1 Antagonist in Treatment of Obesity: Experience of RIO-Asia

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    Changyu Pan

    2011-01-01

    Full Text Available Rimonabant, a selective cannabinoid-1 (CB1 receptor antagonist, has been shown to reduce weight and enhance improvements in cardiometabolic risk parameters in Western populations. This study assessed these effects of rimonabant in Asian population. A total of 643 patients (BMI 25 kg/m2 or greater without diabetes from China, Republic of Korea, and Taiwan were prescribed a hypocaloric diet (600 kcal/day deficit and randomized to rimonabant 20 mg (n=318 or placebo (n=325 for 9months. The primary efficacy variable was weight change from baseline after 9 months of treatment. Results showed that rimonabant group lost more weight than placebo, (LSM ± SEM of −4.7 ± 0.3 kg vs. −1.7 ± 0.3 kg, P<.0001. The 5% and 10% responders were 2 or 3 folds more in the rimonabant group (53.0% vs. 20.0% and 21.5% vs. 5.7%, resp. (P<.0001. Rimonabant also significantly increased HDL-cholesterol, decreased triglycerides and waist circumference,by 7.1%, 10.6%, and 2.8 cm, respectively (P<.0001. This study confirmed the comparable efficacy and safety profile of rimonabant in Asian population to Caucasians. Owing to the recent suspension of all the CB1 antagonists off the pharmaceutical market for weight reduction in Europe and USA, a perspective in drug discovery for intervening peripheral CB1 receptor in the management of obesity is discussed.

  11. Cannabinoid transmission in the prelimbic cortex bidirectionally controls opiate reward and aversion signaling through dissociable kappa versus μ-opiate receptor dependent mechanisms.

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    Ahmad, Tasha; Lauzon, Nicole M; de Jaeger, Xavier; Laviolette, Steven R

    2013-09-25

    Cannabinoid, dopamine (DA), and opiate receptor pathways play integrative roles in emotional learning, associative memory, and sensory perception. Modulation of cannabinoid CB1 receptor transmission within the medial prefrontal cortex (mPFC) regulates the emotional valence of both rewarding and aversive experiences. Furthermore, CB1 receptor substrates functionally interact with opiate-related motivational processing circuits, particularly in the context of reward-related learning and memory. Considerable evidence demonstrates functional interactions between CB1 and DA signaling pathways during the processing of motivationally salient information. However, the role of mPFC CB1 receptor transmission in the modulation of behavioral opiate-reward processing is not currently known. Using an unbiased conditioned place preference paradigm with rats, we examined the role of intra-mPFC CB1 transmission during opiate reward learning. We report that activation or inhibition of CB1 transmission within the prelimbic cortical (PLC) division of the mPFC bidirectionally regulates the motivational valence of opiates; whereas CB1 activation switched morphine reward signaling into an aversive stimulus, blockade of CB1 transmission potentiated the rewarding properties of normally sub-reward threshold conditioning doses of morphine. Both of these effects were dependent upon DA transmission as systemic blockade of DAergic transmission prevented CB1-dependent modulation of morphine reward and aversion behaviors. We further report that CB1-mediated intra-PLC opiate motivational signaling is mediated through a μ-opiate receptor-dependent reward pathway, or a κ-opiate receptor-dependent aversion pathway, directly within the ventral tegmental area. Our results provide evidence for a novel CB1-mediated motivational valence switching mechanism within the PLC, controlling dissociable subcortical reward and aversion pathways.

  12. NESS038C6, a novel selective CB1 antagonist agent with anti-obesity activity and improved molecular profile.

    Science.gov (United States)

    Mastinu, Andrea; Pira, Marilena; Pani, Luca; Pinna, Gérard Aimè; Lazzari, Paolo

    2012-10-01

    The present work aims to study the effects induced by a chronic treatment with a novel CB1 antagonist (NESS038C6) in C57BL/6N diet-induced obesity (DIO) mice. Mice treated with NESS038C6 and fed with a fat diet (NESS038C6 FD) were compared with the following three reference experimental groups: DIO mice fed with the same fat diet used for NESS038C6 and treated with vehicle or the reference CB1 antagonist/inverse agonist rimonabant, "VH FD" and "SR141716 FD", respectively; DIO mice treated with vehicle and switched to a normal diet (VH ND). NESS038C6 chronic treatment (30 mg/kg/day for 31 days) determined a significant reduction in DIO mice weight relative to that of VH FD. The entity of the effect was comparable to that detected in both SR141716 FD and VH ND groups. Moreover, if compared to VH FD, NESS038C6 FD evidenced: (i) improvement of cardiovascular risk factors; (ii) significant decrease in adipose tissue leptin expression; (iii) increase in mRNA expression of hypothalamic orexigenic peptides and a decrease of anorexigenic peptides; (iv) expression increase of metabolic enzymes and peroxisome proliferator-activated receptor-α in the liver; (v) normalization of monoaminergic transporters and neurotrophic expression in mesolimbic area. However, in contrast to the case of rimonabant, the novel CB1 antagonist improved the disrupted expression profile of genes linked to the hunger-satiety circuit, without altering monoaminergic transmission. In conclusion, the novel CB1 antagonist compound NESS038C6 may represent a useful candidate agent for the treatment of obesity and its metabolic complications, without or with reduced side effects relative to those instead observed with rimonabant. Copyright © 2012 Elsevier B.V. All rights reserved.

  13. Cannabinoid Receptor Type 1 Expression in the Developing Avian Retina: Morphological and Functional Correlation With the Dopaminergic System

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    Luzia da Silva Sampaio

    2018-03-01

    Full Text Available The avian retina has been used as a model to study signaling by different neuro- and gliotransmitters. It is unclear how dopaminergic and cannabinoid systems are related in the retina. Here we studied the expression of type 1 and 2 cannabinoid receptors (CB1 and CB2, as well as monoacylglycerol lipase (MAGL, the enzyme that degrades 2-arachidonoylglycerol (2-AG, during retina development. Our data show that CB1 receptor is highly expressed from embryonic day 5 (E5 until post hatched day 7 (PE7, decreasing its levels throughout development. CB1 is densely found in the ganglion cell layer (GCL and inner plexiform layer (IPL. CB2 receptor was also found from E5 until PE7 with a decrease in its contents from E9 afterwards. CB2 was mainly present in the lamination of the IPL at PE7. MAGL is expressed in all retinal layers, mainly in the IPL and OPL from E9 to PE7 retina. CB1 and CB2 were found both in neurons and glia cells, but MAGL was only expressed in Müller glia. Older retinas (PE7 show CB1 positive cells mainly in the INL and co-expression of CB1 and tyrosine hydroxylase (TH are shown in a few cells when both systems are mature. CB1 co-localized with TH and was heavily associated to D1 receptor labeling in primary cell cultures. Finally, cyclic AMP (cAMP was activated by the selective D1 agonist SKF38393, and inhibited when cultures were treated with WIN55, 212–2 (WIN in a CB1 dependent manner. The results suggest a correlation between the endocannabinoid and dopaminergic systems (DSs during the avian retina development. Activation of CB1 limits cAMP accumulation via D1 receptor activation and may influence embryological parameters during avian retina differentiation.

  14. Effect of blockage of the endocannabinoid system by CB(1) antagonism on cardiovascular risk.

    Science.gov (United States)

    Mach, François; Montecucco, Fabrizio; Steffens, Sabine

    2009-01-01

    The endocannabinoid system is a crucial player in the inflammatory processes underlying atherosclerosis. Recently, basic research studies and animal models have strongly supported the role of the endocannabinoid system not only in the regulation of classical cardiovascular risk factors (including lipid profile and glucose homeostasis), but also in the activation of immune cells and inflammatory mediators. Clinical trials investigating treatment with rimonabant (a selective antagonist of the cannabinoid type 1 receptor) have suggested a beneficial effect of this drug in the management of obesity. Further studies are needed to explore a possible use for rimonabant in treating type 2 diabetes and acute and chronic cardiovascular disease. Despite the slight increase in adverse events (mainly psychiatric), which has led to the recent withdrawal of rimonabant from the market, CB(1) receptor antagonism might represent a very promising therapeutic strategy to reduce the cardiovascular risk. In the present review, we focused on the most important experimental investigations into the role of the endocannabinoid system in atherosclerosis and cardiovascular risk.

  15. CB1-Dependent Long-Term Depression in Ventral Tegmental Area GABA Neurons: A Novel Target for Marijuana.

    Science.gov (United States)

    Friend, Lindsey; Weed, Jared; Sandoval, Philip; Nufer, Teresa; Ostlund, Isaac; Edwards, Jeffrey G

    2017-11-08

    The VTA is necessary for reward behavior with dopamine cells critically involved in reward signaling. Dopamine cells in turn are innervated and regulated by neighboring inhibitory GABA cells. Using whole-cell electrophysiology in juvenile-adolescent GAD67-GFP male mice, we examined excitatory plasticity in fluorescent VTA GABA cells. A novel CB1-dependent LTD was induced in GABA cells that was dependent on metabotropic glutamate receptor 5, and cannabinoid receptor 1 (CB1). LTD was absent in CB1 knock-out mice but preserved in heterozygous littermates. Bath applied Δ 9 -tetrahydrocannabinol depressed GABA cell activity, therefore downstream dopamine cells will be disinhibited; and thus, this could potentially result in increased reward. Chronic injections of Δ 9 -tetrahydrocannabinol occluded LTD compared with vehicle injections; however, a single exposure was insufficient to do so. As synaptic modifications by drugs of abuse are often tied to addiction, these data suggest a possible mechanism for the addictive effects of Δ 9 -tetrahydrocannabinol in juvenile-adolescents, by potentially altering reward behavioral outcomes. SIGNIFICANCE STATEMENT The present study identifies a novel form of glutamatergic synaptic plasticity in VTA GABA neurons, a currently understudied cell type that is critical for the brain's reward circuit, and how Δ 9 -tetrahydrocannabinol occludes this plasticity. This study specifically addresses a potential unifying mechanism whereby marijuana could exert rewarding and addictive/withdrawal effects. Marijuana use and legalization are a pressing issue for many states in the United States. Although marijuana is the most commonly abused illicit drug, the implications of legalized, widespread, or continued usage are speculative. This study in juvenile-adolescent aged mice identifies a novel form of synaptic plasticity in VTA GABA cells, and the synaptic remodeling that can occur after Δ 9 -tetrahydrocannabinol use. Copyright © 2017 the

  16. Increase in hypothalamic AMPK phosphorylation induced by prolonged exposure to LPS involves ghrelin and CB1R signaling.

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    Rivas, Priscila M S; Vechiato, Fernanda M V; Borges, Beatriz C; Rorato, Rodrigo; Antunes-Rodrigues, Jose; Elias, Lucila L K

    2017-07-01

    Acute administration of lipopolysaccharide (LPS) from Gram-negative bacteria induces hypophagia. However, the repeated administration of LPS leads to desensitization of hypophagia, which is associated with increased hypothalamic p-AMPK expression. Because ghrelin and endocannabinoids modulate AMPK activity in the hypothalamus, we hypothesized that these neuromodulators play a role in the reversal of tolerance to hypophagia in rats under long-term exposure to LPS. Male Wistar rats were treated with single (1 LPS, 100μg/kg body weight, ip) or repeated injections of LPS over 6days (6 LPS). Food intake was reduced in the 1 LPS, but not in the 6 LPS group. 6 LPS rats showed an increased serum concentration of acylated ghrelin and reduced ghrelin receptor mRNA expression in the hypothalamus. Ghrelin injection (40μg/kg body weight, ip) increased food intake, body weight gain, p-AMPK hypothalamic expression, neuropeptide Y (NPY) and Agouti related peptide (AgRP) mRNA expression in control animals (Saline). However, in 6 LPS rats, ghrelin did not alter these parameters. Central administration of a CB1R antagonist (AM251, 200ng/μl in 5μl/rat) induced hypophagia in 6 LPS animals, suggesting that the endocannabinoid system contributes to preserved food intake during LPS tolerance. In the presence of AM251, the ability of ghrelin to phosphorylate AMPK in the hypothalamus of 6 LPS group was restored, but not its orexigenic effect. Our data highlight that the orexigenic effects of ghrelin require CB1R signaling downstream of AMPK activation. Moreover, CB1R-mediated pathways contribute to the absence of hypophagia during repeated exposure to endotoxin. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Combined Angiotensin Receptor Antagonism and Neprilysin Inhibition

    Science.gov (United States)

    Hubers, Scott A.; Brown, Nancy J.

    2016-01-01

    Heart failure affects approximately 5.7 million people in the United States alone. Angiotensin-converting enzyme inhibitors, angiotensin receptor blockers, beta-blockers, and aldosterone antagonists have improved mortality in patients with heart failure and reduced ejection fraction, but mortality remains high. In July 2015, the FDA approved the first of a new class of drugs for the treatment of heart failure; valsartan/sacubitril (formerly known as LCZ696 and currently marketed by Novartis as Entresto) combines the angiotensin receptor blocker valsartan and the neprilysin inhibitor prodrug sacubitril in a 1:1 ratio in a sodium supramolecular complex. Sacubitril is converted by esterases to LBQ657, which inhibits neprilysin, the enzyme responsible for the degradation of the natriuretic peptides and many other vasoactive peptides. Thus, this combined angiotensin receptor antagonist and neprilysin inhibitor addresses two of the pathophysiologic mechanisms of heart failure - activation of the renin-angiotensin-aldosterone system and decreased sensitivity to natriuretic peptides. In the Prospective comparison of ARNI with ACEI to Determine Impact on Global Mortality and morbidity in Heart Failure (PARADIGM-HF) trial, valsartan/sacubitril significantly reduced mortality and hospitalization for heart failure, as well as blood pressure, compared to enalapril in patients with heart failure, reduced ejection fraction, and an elevated circulating level of brain natriuretic peptide or N-terminal pro-brain natriuretic peptide. Ongoing clinical trials are evaluating the role of valsartan/sacubitril in the treatment of heart failure with preserved ejection fraction and hypertension. We review here the mechanisms of action of valsartan/sacubitril, the pharmacologic properties of the drug, and its efficacy and safety in the treatment of heart failure and hypertension. PMID:26976916

  18. High Concentrations of Tranexamic Acid Inhibit Ionotropic Glutamate Receptors.

    Science.gov (United States)

    Lecker, Irene; Wang, Dian-Shi; Kaneshwaran, Kirusanthy; Mazer, C David; Orser, Beverley A

    2017-07-01

    The antifibrinolytic drug tranexamic acid is structurally similar to the amino acid glycine and may cause seizures and myoclonus by acting as a competitive antagonist of glycine receptors. Glycine is an obligatory co-agonist of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors. Thus, it is plausible that tranexamic acid inhibits NMDA receptors by acting as a competitive antagonist at the glycine binding site. The aim of this study was to determine whether tranexamic acid inhibits NMDA receptors, as well as α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid and kainate subtypes of ionotropic glutamate receptors. Tranexamic acid modulation of NMDA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid, and kainate receptors was studied using whole cell voltage-clamp recordings of current from cultured mouse hippocampal neurons. Tranexamic acid rapidly and reversibly inhibited NMDA receptors (half maximal inhibitory concentration = 241 ± 45 mM, mean ± SD; 95% CI, 200 to 281; n = 5) and shifted the glycine concentration-response curve for NMDA-evoked current to the right. Tranexamic acid also inhibited α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (half maximal inhibitory concentration = 231 ± 91 mM; 95% CI, 148 to 314; n = 5 to 6) and kainate receptors (half maximal inhibitory concentration = 90 ± 24 mM; 95% CI, 68 to 112; n = 5). Tranexamic acid inhibits NMDA receptors likely by reducing the binding of the co-agonist glycine and also inhibits α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid and kainate receptors. Receptor blockade occurs at high millimolar concentrations of tranexamic acid, similar to the concentrations that occur after topical application to peripheral tissues. Glutamate receptors in tissues including bone, heart, and nerves play various physiologic roles, and tranexamic acid inhibition of these receptors may contribute to adverse drug effects.

  19. Platelet-rich plasma loaded in situ-formed hydrogel enhances hyaline cartilage regeneration by CB1 upregulation.

    Science.gov (United States)

    Lee, Hye-Rim; Park, Kyung Min; Joung, Yoon Ki; Park, Ki Dong; Do, Sun Hee

    2012-11-01

    The efficacy of three-dimensional (3D) culture on the proliferation and maturation of chondrocytes seeded into a hydrogel scaffold was assessed. Three types of hydrogel were prepared for the 3D culture of primary isolated chondrocytes. Chondrocyte proliferation was assessed using a live/dead viability/cytotoxicity assay and semiquantitative RT-PCR after 3D culture in hydrogel. Cylindrical defects in the center of rat xyphoids were used for the implantation of platelet-rich plasma (PRP)/hydrogel composites. Rats were killed at day 7 postoperatively and evaluated histochemically and immunohistologically. Xyphoid chondrocytes proliferated well with time in hydrogels. In the PRP-containing hydrogels, xyphoid defects displayed early formation of chondroid matrix with massive peripheral infiltration of spindle cells. These results were consistent with Safranin-O staining for proteoglycans and immunohistochemistry for type II collagen. Gene expression analyses in vitro revealed aggrecan, type II collagen, and ChM-1 and CB1 upregulation by PRP/hydrogel. PRP/hydrogel provided a suitable environment for hyaline cartilaginous regeneration, leading to anti-inflammation by significant increase of CB1 and inhibiting vascular ingrowth via considerable upregulation of ChM-1. The results provide a valuable reference for the clinical application of hydrogel scaffolds for hyaline cartilage regeneration, as well as the use of autologous PRP to improve cellular proliferation and maturation of xyphoid repair. Copyright © 2012 Wiley Periodicals, Inc.

  20. Role of hypothalamic cannabinoid receptors in post-stroke depression in rats.

    Science.gov (United States)

    Wang, Shanshan; Sun, Hong; Liu, Sainan; Wang, Ting; Guan, Jinqun; Jia, Jianjun

    2016-03-01

    One of the most common psychological consequences of stroke is post-stroke depression (PSD). While more than 30 percent of stroke patients eventually develop PSD, the neurobiological mechanisms underlying such a phenomenon have not been well investigated. Given the critical involvement of hypothalamic-pituitary-adrenal axis and endocannabinoid system in response to stressful stimuli, we evaluated the hypothesis that cannabinoid receptors in the hypothalamus are critical for modulation of post-stroke depression-like behaviors in rats. To this end, rats were treated with middle cerebral artery occlusion (MCAO) followed by chronic unpredictable mild stress (CUMS) treatment procedure. We then assessed the expression of CB1 and CB2 receptors in the hypothalamus, and evaluated the effects of pharmacological stimulations of CB1 or CB2 receptors on the expression and development of depression-like behaviors in PSD rats. We found that PSD rats exhibited decreased the expression of CB1 receptor, but not CB2 receptor, in the ventral medial hypothalamus (VMH). Such an effect was not observed in the dorsally adjacent brain regions. Furthermore, intra-VMH injections of CB2 receptor agonist, but not CB1 receptor agonist, attenuated the expression of depression-like behaviors in PSD rats. Finally, repeated intraperitoneal injections of CB1 or CB2 receptor agonists during CUMS treatment inhibited the development of depression-like behaviors in PSD rats. Taken together, these results suggest that decreased CB1 receptor expression is likely associated with the development of post-stroke depression, and CB2 receptor may be a potential therapeutic target for the treatment post-stroke depressive disorders. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Anandamide inhibits adhesion and migration of breast cancer cells

    International Nuclear Information System (INIS)

    Grimaldi, Claudia; Pisanti, Simona; Laezza, Chiara; Malfitano, Anna Maria; Santoro, Antonietta; Vitale, Mario; Caruso, Maria Gabriella; Notarnicola, Maria; Iacuzzo, Irma; Portella, Giuseppe; Di Marzo, Vincenzo; Bifulco, Maurizio

    2006-01-01

    The endocannabinoid system regulates cell proliferation in human breast cancer cells. We reasoned that stimulation of cannabinoid CB 1 receptors could induce a non-invasive phenotype in breast mtastatic cells. In a model of metastatic spreading in vivo, the metabolically stable anandamide analogue, 2-methyl-2'-F-anandamide (Met-F-AEA), significantly reduced the number and dimension of metastatic nodes, this effect being antagonized by the selective CB 1 antagonist SR141716A. In MDA-MB-231 cells, a highly invasive human breast cancer cell line, and in TSA-E1 cells, a murine breast cancer cell line, Met-F-AEA inhibited adhesion and migration on type IV collagen in vitro without modifying integrin expression: both these effects were antagonized by SR141716A. In order to understand the molecular mechanism involved in these processes, we analyzed the phosphorylation of FAK and Src, two tyrosine kinases involved in migration and adhesion. In Met-F-AEA-treated cells, we observed a decreased tyrosine phosphorylation of both FAK and Src, this effect being attenuated by SR141716A. We propose that CB 1 receptor agonists inhibit tumor cell invasion and metastasis by modulating FAK phosphorylation, and that CB 1 receptor activation might represent a novel therapeutic strategy to slow down the growth of breast carcinoma and to inhibit its metastatic diffusion in vivo

  2. Electroacupuncture Potentiates Cannabinoid Receptor-Mediated Descending Inhibitory Control in a Mouse Model of Knee Osteoarthritis

    Directory of Open Access Journals (Sweden)

    Xiao-Cui Yuan

    2018-04-01

    Full Text Available Knee osteoarthritis (KOA is a highly prevalent, chronic joint disorder, which can lead to chronic pain. Although electroacupuncture (EA is effective in relieving chronic pain in the clinic, the involved mechanisms remain unclear. Reduced diffuse noxius inhibitory controls (DNIC function is associated with chronic pain and may be related to the action of endocannabinoids. In the present study, we determined whether EA may potentiate cannabinoid receptor-mediated descending inhibitory control and inhibit chronic pain in a mouse model of KOA. We found that the optimized parameters of EA inhibiting chronic pain were the low frequency and high intensity (2 Hz + 1 mA. EA reversed the reduced expression of CB1 receptors and the 2-arachidonoylglycerol (2-AG level in the midbrain in chronic pain. Microinjection of the CB1 receptor antagonist AM251 into the ventrolateral periaqueductal gray (vlPAG can reversed the EA effect on pain hypersensitivity and DNIC function. In addition, CB1 receptors on GABAergic but not glutamatergic neurons are involved in the EA effect on DNIC function and descending inhibitory control of 5-HT in the medulla, thus inhibiting chronic pain. Our data suggest that endocannabinoid (2-AG-CB1R-GABA-5-HT may be a novel signaling pathway involved in the effect of EA improving DNIC function and inhibiting chronic pain.

  3. Prefrontal activity during working memory is modulated by the interaction of variation in CB1 and COX2 coding genes and correlates with frequency of cannabis use.

    Science.gov (United States)

    Taurisano, Paolo; Antonucci, Linda A; Fazio, Leonardo; Rampino, Antonio; Romano, Raffaella; Porcelli, Annamaria; Masellis, Rita; Colizzi, Marco; Quarto, Tiziana; Torretta, Silvia; Di Giorgio, Annabella; Pergola, Giulio; Bertolino, Alessandro; Blasi, Giuseppe

    2016-08-01

    The CB1 cannabinoid receptor is targeted in the brain by endocannabinoids under physiological conditions as well as by delta9-tetrahydrocannabinol under cannabis use. Furthermore, its signaling appears to affect brain cognitive processing. Recent findings highlight a crucial role of cyclooxygenase-2 (COX-2) in the mechanism of intraneuronal CB1 signaling transduction, while others indicate that two single nucleotide polymorphisms (SNPs) (rs1406977 and rs20417) modulate expression of CB1 (CNR1) and COX-2 (PTGS2) coding genes, respectively. Here, our aim was to use fMRI to investigate in healthy humans whether these SNPs interact in modulating prefrontal activity during working memory processing and if this modulation is linked with cannabis use. We recruited 242 healthy subjects genotyped for CNR1 rs1406977 and PTGS2 rs20417 that performed the N-back working memory task during fMRI and were interviewed using the Cannabis Experience Questionnaire (CEQ). We found that the interaction between CNR1 rs1406977 and PTGS2 rs20417 is associated with dorsolateral prefrontal cortex (DLPFC) activity such that specific genotype configurations (CNR1 C carriers/PTGS2 C carriers and CNR1 TT/PTGS2 GG) predict lower cortical response versus others in spite of similar behavioral accuracy. Furthermore, DLPFC activity in the cluster associated with the CNR1 by PTGS2 interaction was negatively correlated with behavioral efficiency and positively correlated with frequency of cannabis use in cannabis users. These results suggest that a genetically modulated balancing of signaling within the CB1-COX-2 pathway may reflect on more or less efficient patterns of prefrontal activity during working memory. Frequency of cannabis use may be a factor for further modulation of CNR1/PTGS2-mediated cortical processing associated with this cognitive process. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Enzymic Dehalogenation of 4-Chlorobenzoyl Coenzyme A in Acinetobacter sp. Strain 4-CB1

    OpenAIRE

    Copley, Shelley D.; Crooks, Gwen P.

    1992-01-01

    4-Chlorobenzoate degradation in cell extracts of Acinetobacter sp. strain 4-CB1 occurs by initial synthesis of 4-chlorobenzoyl coenzyme A (4-chlorobenzoyl CoA) from 4-chlorobenzoate, CoA, and ATP. 4-Chlorobenzoyl CoA is dehalogenated to 4-hydroxybenzoyl CoA. Following the dehalogenation reaction, 4-hydroxybenzoyl CoA is hydrolyzed to 4-hydroxybenzoate and CoA. Possible roles for the CoA moiety in the dehalogenation reaction are discussed.

  5. Enzymic Dehalogenation of 4-Chlorobenzoyl Coenzyme A in Acinetobacter sp. Strain 4-CB1

    Science.gov (United States)

    Copley, Shelley D.; Crooks, Gwen P.

    1992-01-01

    4-Chlorobenzoate degradation in cell extracts of Acinetobacter sp. strain 4-CB1 occurs by initial synthesis of 4-chlorobenzoyl coenzyme A (4-chlorobenzoyl CoA) from 4-chlorobenzoate, CoA, and ATP. 4-Chlorobenzoyl CoA is dehalogenated to 4-hydroxybenzoyl CoA. Following the dehalogenation reaction, 4-hydroxybenzoyl CoA is hydrolyzed to 4-hydroxybenzoate and CoA. Possible roles for the CoA moiety in the dehalogenation reaction are discussed. PMID:16348702

  6. Role of endocannabinoids and cannabinoid-1 receptors in cerebrocortical blood flow regulation.

    Directory of Open Access Journals (Sweden)

    András Iring

    Full Text Available Endocannabinoids are among the most intensively studied lipid mediators of cardiovascular functions. In the present study the effects of decreased and increased activity of the endocannabinoid system (achieved by cannabinoid-1 (CB1 receptor blockade and inhibition of cannabinoid reuptake, respectively on the systemic and cerebral circulation were analyzed under steady-state physiological conditions and during hypoxia and hypercapnia (H/H.In anesthetized spontaneously ventilating rats the CB1-receptor antagonist/inverse agonist AM-251 (10 mg/kg, i.v. failed to influence blood pressure (BP, cerebrocortical blood flow (CoBF, measured by laser-Doppler flowmetry or arterial blood gas levels. In contrast, the putative cannabinoid reuptake inhibitor AM-404 (10 mg/kg, i.v. induced triphasic responses, some of which could be blocked by AM-251. Hypertension during phase I was resistant to AM-251, whereas the concomitant CoBF-increase was attenuated. In contrast, hypotension during phase III was sensitive to AM-251, whereas the concomitant CoBF-decrease was not. Therefore, CoBF autoregulation appeared to shift towards higher BP levels after CB1-blockade. During phase II H/H developed due to respiratory depression, which could be inhibited by AM-251. Interestingly, however, the concomitant rise in CoBF remained unchanged after AM-251, indicating that CB1-blockade potentially enhanced the reactivity of the CoBF to H/H. In accordance with this hypothesis, AM-251 induced a significant enhancement of the CoBF responses during controlled stepwise H/H.Under resting physiological conditions CB1-receptor mediated mechanisms appear to have limited influence on systemic or cerebral circulation. Enhancement of endocannabinoid levels, however, induces transient CB1-independent hypertension and sustained CB1-mediated hypotension. Furthermore, enhanced endocannabinoid activity results in respiratory depression in a CB1-dependent manner. Finally, our data indicate for the

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

    Directory of Open Access Journals (Sweden)

    Abrar Ashoor

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

  8. Cannabinoid signalling inhibits sarcoplasmic Ca2+ release and regulates excitation–contraction coupling in mammalian skeletal muscle

    Science.gov (United States)

    Oláh, Tamás; Bodnár, Dóra; Tóth, Adrienn; Vincze, János; Fodor, János; Reischl, Barbara; Kovács, Adrienn; Ruzsnavszky, Olga; Dienes, Beatrix; Szentesi, Péter; Friedrich, Oliver

    2016-01-01

    Key points Marijuana was found to cause muscle weakness, although the exact regulatory role of its receptors (CB1 cannabinoid receptor; CB1R) in the excitation–contraction coupling (ECC) of mammalian skeletal muscle remains unknown.We found that CB1R activation or its knockout did not affect muscle force directly, whereas its activation decreased the Ca2+‐sensitivity of the contractile apparatus and made the muscle fibres more prone to fatigue.We demonstrate that CB1Rs are not connected to the inositol 1,4,5‐trisphosphate pathway either in myotubes or in adult muscle fibres.By contrast, CB1Rs constitutively inhibit sarcoplasmic Ca2+ release and sarcoplasmic reticulum Ca2+ ATPase during ECC in a Gi/o protein‐mediated way in adult skeletal muscle fibres but not in myotubes.These results help with our understanding of the physiological effects and pathological consequences of CB1R activation in skeletal muscle and may be useful in the development of new cannabinoid drugs. Abstract Marijuana was found to cause muscle weakness, although it is unknown whether it affects the muscles directly or modulates only the motor control of the central nervous system. Although the presence of CB1 cannabinoid receptors (CB1R), which are responsible for the psychoactive effects of the drug in the brain, have recently been demonstrated in skeletal muscle, it is unclear how CB1R‐mediated signalling affects the contraction and Ca²⁺ homeostasis of mammalian skeletal muscle. In the present study, we demonstrate that in vitro CB1R activation increased muscle fatigability and decreased the Ca2+‐sensitivity of the contractile apparatus, whereas it did not alter the amplitude of single twitch contractions. In myotubes, CB1R agonists neither evoked, nor influenced inositol 1,4,5‐trisphosphate (IP3)‐mediated Ca2+ transients, nor did they alter excitation–contraction coupling. By contrast, in isolated muscle fibres of wild‐type mice, although CB1R agonists did not evoke IP3

  9. Cannabinoid signalling inhibits sarcoplasmic Ca2+ release and regulates excitation-contraction coupling in mammalian skeletal muscle.

    Science.gov (United States)

    Oláh, Tamás; Bodnár, Dóra; Tóth, Adrienn; Vincze, János; Fodor, János; Reischl, Barbara; Kovács, Adrienn; Ruzsnavszky, Olga; Dienes, Beatrix; Szentesi, Péter; Friedrich, Oliver; Csernoch, László

    2016-12-15

    Marijuana was found to cause muscle weakness, although the exact regulatory role of its receptors (CB1 cannabinoid receptor; CB1R) in the excitation-contraction coupling (ECC) of mammalian skeletal muscle remains unknown. We found that CB1R activation or its knockout did not affect muscle force directly, whereas its activation decreased the Ca 2+ -sensitivity of the contractile apparatus and made the muscle fibres more prone to fatigue. We demonstrate that CB1Rs are not connected to the inositol 1,4,5-trisphosphate pathway either in myotubes or in adult muscle fibres. By contrast, CB1Rs constitutively inhibit sarcoplasmic Ca 2+ release and sarcoplasmic reticulum Ca 2+ ATPase during ECC in a G i/o protein-mediated way in adult skeletal muscle fibres but not in myotubes. These results help with our understanding of the physiological effects and pathological consequences of CB1R activation in skeletal muscle and may be useful in the development of new cannabinoid drugs. Marijuana was found to cause muscle weakness, although it is unknown whether it affects the muscles directly or modulates only the motor control of the central nervous system. Although the presence of CB1 cannabinoid receptors (CB1R), which are responsible for the psychoactive effects of the drug in the brain, have recently been demonstrated in skeletal muscle, it is unclear how CB1R-mediated signalling affects the contraction and Ca²⁺ homeostasis of mammalian skeletal muscle. In the present study, we demonstrate that in vitro CB1R activation increased muscle fatigability and decreased the Ca 2+ -sensitivity of the contractile apparatus, whereas it did not alter the amplitude of single twitch contractions. In myotubes, CB1R agonists neither evoked, nor influenced inositol 1,4,5-trisphosphate (IP 3 )-mediated Ca 2+ transients, nor did they alter excitation-contraction coupling. By contrast, in isolated muscle fibres of wild-type mice, although CB1R agonists did not evoke IP 3 -mediated Ca 2

  10. Cocaine Inhibits Dopamine D2 Receptor Signaling via Sigma-1-D2 Receptor Heteromers

    Science.gov (United States)

    Navarro, Gemma; Moreno, Estefania; Bonaventura, Jordi; Brugarolas, Marc; Farré, Daniel; Aguinaga, David; Mallol, Josefa; Cortés, Antoni; Casadó, Vicent; Lluís, Carmen; Ferre, Sergi

    2013-01-01

    Under normal conditions the brain maintains a delicate balance between inputs of reward seeking controlled by neurons containing the D1-like family of dopamine receptors and inputs of aversion coming from neurons containing the D2-like family of dopamine receptors. Cocaine is able to subvert these balanced inputs by altering the cell signaling of these two pathways such that D1 reward seeking pathway dominates. Here, we provide an explanation at the cellular and biochemical level how cocaine may achieve this. Exploring the effect of cocaine on dopamine D2 receptors function, we present evidence of σ1 receptor molecular and functional interaction with dopamine D2 receptors. Using biophysical, biochemical, and cell biology approaches, we discovered that D2 receptors (the long isoform of the D2 receptor) can complex with σ1 receptors, a result that is specific to D2 receptors, as D3 and D4 receptors did not form heteromers. We demonstrate that the σ1-D2 receptor heteromers consist of higher order oligomers, are found in mouse striatum and that cocaine, by binding to σ1 -D2 receptor heteromers, inhibits downstream signaling in both cultured cells and in mouse striatum. In contrast, in striatum from σ1 knockout animals these complexes are not found and this inhibition is not seen. Taken together, these data illuminate the mechanism by which the initial exposure to cocaine can inhibit signaling via D2 receptor containing neurons, destabilizing the delicate signaling balance influencing drug seeking that emanates from the D1 and D2 receptor containing neurons in the brain. PMID:23637801

  11. GABA, its receptors, and GABAergic inhibition in mouse taste buds.

    Science.gov (United States)

    Dvoryanchikov, Gennady; Huang, Yijen A; Barro-Soria, Rene; Chaudhari, Nirupa; Roper, Stephen D

    2011-04-13

    Taste buds consist of at least three principal cell types that have different functions in processing gustatory signals: glial-like (type I) cells, receptor (type II) cells, and presynaptic (type III) cells. Using a combination of Ca2+ imaging, single-cell reverse transcriptase-PCR and immunostaining, we show that GABA is an inhibitory transmitter in mouse taste buds, acting on GABA(A) and GABA(B) receptors to suppress transmitter (ATP) secretion from receptor cells during taste stimulation. Specifically, receptor cells express GABA(A) receptor subunits β2, δ, and π, as well as GABA(B) receptors. In contrast, presynaptic cells express the GABA(A) β3 subunit and only occasionally GABA(B) receptors. In keeping with the distinct expression pattern of GABA receptors in presynaptic cells, we detected no GABAergic suppression of transmitter release from presynaptic cells. We suggest that GABA may serve function(s) in taste buds in addition to synaptic inhibition. Finally, we also defined the source of GABA in taste buds: GABA is synthesized by GAD65 in type I taste cells as well as by GAD67 in presynaptic (type III) taste cells and is stored in both those two cell types. We conclude that GABA is an inhibitory transmitter released during taste stimulation and possibly also during growth and differentiation of taste buds.

  12. Pure Δ9-tetrahydrocannabivarin and a Cannabis sativa extract with high content in Δ9-tetrahydrocannabivarin inhibit nitrite production in murine peritoneal macrophages.

    Science.gov (United States)

    Romano, Barbara; Pagano, Ester; Orlando, Pierangelo; Capasso, Raffaele; Cascio, Maria Grazia; Pertwee, Roger; Marzo, Vincenzo Di; Izzo, Angelo A; Borrelli, Francesca

    2016-11-01

    Historical and scientific evidence suggests that Cannabis use has immunomodulatory and anti-inflammatory effects. We have here investigated the effect of the non-psychotropic phytocannabinoid Δ 9 -tetrahydrocannabivarin (THCV) and of a Cannabis sativa extract with high (64.8%) content in THCV (THCV-BDS) on nitric oxide (NO) production, and on cannabinoid and transient receptor potential (TRP) channel expression in lipopolysaccharide (LPS)-stimulated murine peritoneal macrophages. THCV-BDS and THCV exhibited similar affinity in radioligand binding assays for CB 1 and CB 2 receptors, and inhibited, via CB 2 but not CB 1 cannabinoid receptors, nitrite production evoked by LPS in peritoneal macrophages. THCV down-regulated the over-expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and interleukin 1β (IL-1β) proteins induced by LPS. Furthermore, THCV counteracted LPS-induced up-regulation of CB 1 receptors, without affecting the changes in CB 2 , TRPV2 or TRPV4 mRNA expression caused by LPS. Other TRP channels, namely, TRPA1, TRPV1, TRPV3 and TRPM8 were poorly expressed or undetectable in both unstimulated and LPS-challenged macrophages. It is concluded that THCV - via CB 2 receptor activation - inhibits nitrite production in macrophages. The effect of this phytocannabinoid was associated with a down-regulation of CB 1 , but not CB 2 or TRP channel mRNA expression. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Imidazopyridine CB2 agonists: optimization of CB2/CB1 selectivity and implications for in vivo analgesic efficacy.

    Science.gov (United States)

    Trotter, B Wesley; Nanda, Kausik K; Burgey, Christopher S; Potteiger, Craig M; Deng, James Z; Green, Ahren I; Hartnett, John C; Kett, Nathan R; Wu, Zhicai; Henze, Darrell A; Della Penna, Kimberly; Desai, Reshma; Leitl, Michael D; Lemaire, Wei; White, Rebecca B; Yeh, Suzie; Urban, Mark O; Kane, Stefanie A; Hartman, George D; Bilodeau, Mark T

    2011-04-15

    A new series of imidazopyridine CB2 agonists is described. Structural optimization improved CB2/CB1 selectivity in this series and conferred physical properties that facilitated high in vivo exposure, both centrally and peripherally. Administration of a highly selective CB2 agonist in a rat model of analgesia was ineffective despite substantial CNS exposure, while administration of a moderately selective CB2/CB1 agonist exhibited significant analgesic effects. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Fcγ receptor-mediated inflammation inhibits axon regeneration.

    Directory of Open Access Journals (Sweden)

    Gang Zhang

    Full Text Available Anti-glycan/ganglioside antibodies are the most common immune effectors found in patients with Guillain-Barré Syndrome, which is a peripheral autoimmune neuropathy. We previously reported that disease-relevant anti-glycan autoantibodies inhibited axon regeneration, which echo the clinical association of these antibodies and poor recovery in Guillain-Barré Syndrome. However, the specific molecular and cellular elements involved in this antibody-mediated inhibition of axon regeneration are not previously defined. This study examined the role of Fcγ receptors and macrophages in the antibody-mediated inhibition of axon regeneration. A well characterized antibody passive transfer sciatic nerve crush and transplant models were used to study the anti-ganglioside antibody-mediated inhibition of axon regeneration in wild type and various mutant and transgenic mice with altered expression of specific Fcγ receptors and macrophage/microglia populations. Outcome measures included behavior, electrophysiology, morphometry, immunocytochemistry, quantitative real-time PCR, and western blotting. We demonstrate that the presence of autoantibodies, directed against neuronal/axonal cell surface gangliosides, in the injured mammalian peripheral nerves switch the proregenerative inflammatory environment to growth inhibitory milieu by engaging specific activating Fcγ receptors on recruited monocyte-derived macrophages to cause severe inhibition of axon regeneration. Our data demonstrate that the antibody orchestrated Fcγ receptor-mediated switch in inflammation is one mechanism underlying inhibition of axon regeneration. These findings have clinical implications for nerve repair and recovery in antibody-mediated immune neuropathies. Our results add to the complexity of axon regeneration in injured peripheral and central nervous systems as adverse effects of B cells and autoantibodies on neural injury and repair are increasingly recognized.

  15. Gβ promotes pheromone receptor polarization and yeast chemotropism by inhibiting receptor phosphorylation.

    Science.gov (United States)

    Ismael, Amber; Tian, Wei; Waszczak, Nicholas; Wang, Xin; Cao, Youfang; Suchkov, Dmitry; Bar, Eli; Metodiev, Metodi V; Liang, Jie; Arkowitz, Robert A; Stone, David E

    2016-04-12

    Gradient-directed cell migration (chemotaxis) and growth (chemotropism) are processes that are essential to the development and life cycles of all species. Cells use surface receptors to sense the shallow chemical gradients that elicit chemotaxis and chemotropism. Slight asymmetries in receptor activation are amplified by downstream signaling systems, which ultimately induce dynamic reorganization of the cytoskeleton. During the mating response of budding yeast, a model chemotropic system, the pheromone receptors on the plasma membrane polarize to the side of the cell closest to the stimulus. Although receptor polarization occurs before and independently of actin cable-dependent delivery of vesicles to the plasma membrane (directed secretion), it requires receptor internalization. Phosphorylation of pheromone receptors by yeast casein kinase 1 or 2 (Yck1/2) stimulates their internalization. We showed that the pheromone-responsive Gβγ dimer promotes the polarization of the pheromone receptor by interacting with Yck1/2 and locally inhibiting receptor phosphorylation. We also found that receptor phosphorylation is essential for chemotropism, independently of its role in inducing receptor internalization. A mathematical model supports the idea that the interaction between Gβγ and Yck1/2 results in differential phosphorylation and internalization of the pheromone receptor and accounts for its polarization before the initiation of directed secretion. Copyright © 2016, American Association for the Advancement of Science.

  16. Targeting fibroblast growth factor receptor signaling inhibits prostate cancer progression.

    Science.gov (United States)

    Feng, Shu; Shao, Longjiang; Yu, Wendong; Gavine, Paul; Ittmann, Michael

    2012-07-15

    Extensive correlative studies in human prostate cancer as well as studies in vitro and in mouse models indicate that fibroblast growth factor receptor (FGFR) signaling plays an important role in prostate cancer progression. In this study, we used a probe compound for an FGFR inhibitor, which potently inhibits FGFR-1-3 and significantly inhibits FGFR-4. The purpose of this study is to determine whether targeting FGFR signaling from all four FGFRs will have in vitro activities consistent with inhibition of tumor progression and will inhibit tumor progression in vivo. Effects of AZ8010 on FGFR signaling and invasion were analyzed using immortalized normal prostate epithelial (PNT1a) cells and PNT1a overexpressing FGFR-1 or FGFR-4. The effect of AZ8010 on invasion and proliferation in vitro was also evaluated in prostate cancer cell lines. Finally, the impact of AZ8010 on tumor progression in vivo was evaluated using a VCaP xenograft model. AZ8010 completely inhibits FGFR-1 and significantly inhibits FGFR-4 signaling at 100 nmol/L, which is an achievable in vivo concentration. This results in marked inhibition of extracellular signal-regulated kinase (ERK) phosphorylation and invasion in PNT1a cells expressing FGFR-1 and FGFR-4 and all prostate cancer cell lines tested. Treatment in vivo completely inhibited VCaP tumor growth and significantly inhibited angiogenesis and proliferation and increased cell death in treated tumors. This was associated with marked inhibition of ERK phosphorylation in treated tumors. Targeting FGFR signaling is a promising new approach to treating aggressive prostate cancer.

  17. MDM2 binds and inhibits vitamin D receptor

    OpenAIRE

    Heyne, Kristina; Heil, Tessa-Carina; Bette, Birgit; Reichrath, Jörg; Roemer, Klaus

    2015-01-01

    The E3 ubiquitin ligase and transcriptional repressor MDM2 is a potent inhibitor of the p53 family of transcription factors and tumor suppressors. Herein, we report that vitamin D receptor (VDR), another transcriptional regulator and probably, tumor suppressor, is also bound and inhibited by MDM2. This interaction was not affected by vitamin D ligand. VDR was ubiquitylated in the cell and its steady-state level was controlled by the proteasome. Strikingly, overproduced MDM2 reduced the level ...

  18. Potential upstream regulators of cannabinoid receptor 1 signaling in prostate cancer: a Bayesian network analysis of data from a tissue microarray.

    Science.gov (United States)

    Häggström, Jenny; Cipriano, Mariateresa; Forshell, Linus Plym; Persson, Emma; Hammarsten, Peter; Stella, Nephi; Fowler, Christopher J

    2014-08-01

    The endocannabinoid system regulates cancer cell proliferation, and in prostate cancer a high cannabinoid CB1 receptor expression is associated with a poor prognosis. Down-stream mediators of CB1 receptor signaling in prostate cancer are known, but information on potential upstream regulators is lacking. Data from a well-characterized tumor tissue microarray were used for a Bayesian network analysis using the max-min hill-climbing method. In non-malignant tissue samples, a directionality of pEGFR (the phosphorylated form of the epidermal growth factor receptor) → CB1 receptors were found regardless as to whether the endocannabinoid metabolizing enzyme fatty acid amide hydrolase (FAAH) was included as a parameter. A similar result was found in the tumor tissue, but only when FAAH was included in the analysis. A second regulatory pathway, from the growth factor receptor ErbB2 → FAAH was also identified in the tumor samples. Transfection of AT1 prostate cancer cells with CB1 receptors induced a sensitivity to the growth-inhibiting effects of the CB receptor agonist CP55,940. The sensitivity was not dependent upon the level of receptor expression. Thus a high CB1 receptor expression alone does not drive the cells towards a survival phenotype in the presence of a CB receptor agonist. The data identify two potential regulators of the endocannabinoid system in prostate cancer and allow the construction of a model of a dysregulated endocannabinoid signaling network in this tumor. Further studies should be designed to test the veracity of the predictions of the network analysis in prostate cancer and other solid tumors. © 2014 The Authors. The Prostate published by Wiley Periodicals, Inc.

  19. Resistance to diet-induced adiposity in cannabinoid receptor-1 deficient mice is not due to impaired adipocyte function

    NARCIS (Netherlands)

    Oosterveer, Maaike H.; Koolman, Anniek H.; de Boer, Pieter T.; Bos, Trijnie; Bleeker, Aycha; Bloks, Vincent W.; Kuipers, Folkert; Sauer, Pieter J. J.; van Dijk, Gertjan

    2011-01-01

    Background: Overactivity and/or dysregulation of the endocannabinoid system (ECS) contribute to development of obesity. In vitro studies indicate a regulatory role for the cannabinoid receptor 1 (CB1) in adipocyte function and CB1-receptor deficient (CB1-/-) mice are resistant to high fat

  20. Inhibition of Estrogen Receptor Action by the Orphan Receptors, SHP and DAX-1

    National Research Council Canada - National Science Library

    DiRenzo, James

    2003-01-01

    .... In support of DoD grant # DAMD17-99-1-9163, we present our findings regarding the mechanisms by which two orphan nuclear receptors, SHP and DAX-1 inhibit the actions of ER-alpha and ER-beta action...

  1. IGF-1 receptor inhibition by picropodophyllin in medulloblastoma

    Energy Technology Data Exchange (ETDEWEB)

    Ohshima-Hosoyama, Sachiko; Hosoyama, Tohru; Nelon, Laura D. [Greehey Children' s Cancer Research Institute, University of Texas Health Science Center, San Antonio, TX 78229 (United States); Keller, Charles, E-mail: keller@ohsu.edu [Greehey Children' s Cancer Research Institute, University of Texas Health Science Center, San Antonio, TX 78229 (United States); Department of Pediatrics, University of Texas Health Science Center, San Antonio, TX 78229 (United States); Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78229 (United States)

    2010-09-03

    Research highlights: {yields} Igf1r is overexpressed and activated in a Sonic Hedgehog driven model of medulloblastoma. {yields} Picropodophyllin targets and abrogates IGF signaling in medulloblastoma. {yields} Picropodophyllin inhibits medulloblastoma tumor cell growth by induction of apoptosis. -- Abstract: The insulin-like growth factor-1 receptor (Igf1r) is a multifunctional membrane-associated tyrosine kinase associated with regulation of transformation, proliferation, differentiation and apoptosis. Increased IGF pathway activity has been reported in human and murine medulloblastoma. Tumors from our genetically-engineered medulloblastoma mouse model over-express Igf1r, and thus this mouse model is a good platform with which to study the role of Igf1r in tumor progression. We hypothesize that inhibition of IGF pathway in medulloblastoma can slow or inhibit tumor growth and metastasis. To test our hypothesis, we tested the role of IGF in tumor growth in vitro by treatment with the tyrosine kinase small molecule inhibitor, picropodophyllin (PPP), which strongly inhibits the IGF pathway. Our results demonstrate that PPP-mediated downregulation of the IGF pathway inhibits mouse tumor cell growth and induces apoptotic cell death in vitro in primary medulloblastoma cultures that are most reflective of tumor cell behavior in vivo.

  2. IGF-1 receptor inhibition by picropodophyllin in medulloblastoma

    International Nuclear Information System (INIS)

    Ohshima-Hosoyama, Sachiko; Hosoyama, Tohru; Nelon, Laura D.; Keller, Charles

    2010-01-01

    Research highlights: → Igf1r is overexpressed and activated in a Sonic Hedgehog driven model of medulloblastoma. → Picropodophyllin targets and abrogates IGF signaling in medulloblastoma. → Picropodophyllin inhibits medulloblastoma tumor cell growth by induction of apoptosis. -- Abstract: The insulin-like growth factor-1 receptor (Igf1r) is a multifunctional membrane-associated tyrosine kinase associated with regulation of transformation, proliferation, differentiation and apoptosis. Increased IGF pathway activity has been reported in human and murine medulloblastoma. Tumors from our genetically-engineered medulloblastoma mouse model over-express Igf1r, and thus this mouse model is a good platform with which to study the role of Igf1r in tumor progression. We hypothesize that inhibition of IGF pathway in medulloblastoma can slow or inhibit tumor growth and metastasis. To test our hypothesis, we tested the role of IGF in tumor growth in vitro by treatment with the tyrosine kinase small molecule inhibitor, picropodophyllin (PPP), which strongly inhibits the IGF pathway. Our results demonstrate that PPP-mediated downregulation of the IGF pathway inhibits mouse tumor cell growth and induces apoptotic cell death in vitro in primary medulloblastoma cultures that are most reflective of tumor cell behavior in vivo.

  3. An investigation into ‘two hit’ effects of BDNF deficiency and young-adult cannabinoid receptor stimulation on prepulse inhibition regulation and memory in mice

    Directory of Open Access Journals (Sweden)

    Maren eKlug

    2013-10-01

    Full Text Available Reduced brain-derived neurotrophic factor (BDNF signalling has been shown in the frontal cortex and hippocampus in schizophrenia. The aim of the present study was to investigate whether a BDNF deficit would modulate effects of chronic cannabis intake, a well-described risk factor for schizophrenia development. BDNF heterozygous mice (HET and wild-type controls were chronically treated during weeks 6, 7 and 8 of life with the cannabinoid CB1 receptor agonist, CP55,940 (CP. After a 2-week delay, there were no CP-induced deficits in any of the groups in short-term spatial memory in a Y-maze task or novel object recognition memory. Baseline prepulse inhibition (PPI was lower but average startle was increased in BDNF HET compared to wild-type controls. Acute CP administration before the PPI session caused a marked increase in PPI in male HET mice pre-treated with CP but not in any of the other male groups. In females, there were small increases of PPI in all groups upon acute CP administration. Acute CP administration furthermore reduced startle and this effect was greater in HET mice irrespective of chronic CP pre-treatment. Analysis of the levels of [3H]CP55,940 binding by autoradiography revealed a significant increase in the nucleus accumbens of male BDNF HET mice previously treated with CP but not in any of the other groups or in the caudate nucleus.These results show that BDNF deficiency and chronic young-adult cannabinoid receptor stimulation do not interact in this model on learning and memory later in life. In contrast, male ‘two hit’ mice, but not females, were hypersensitive to the effect of acute CP on sensorimotor gating. These effects may be related to a selective increase of [3H]CP55,940 binding in the nucleus accumbens, reflecting up-regulation of CB1 receptor density in this region. These data could be of relevance to our understanding of differential ‘two hit’ neurodevelopmental mechanisms in schizophrenia.

  4. NMDA receptor antagonists inhibit catalepsy induced by either dopamine D1 or D2 receptor antagonists.

    Science.gov (United States)

    Moore, N A; Blackman, A; Awere, S; Leander, J D

    1993-06-11

    In the present study, we investigated the ability of NMDA receptor antagonists to inhibit catalepsy induced by haloperidol, or SCH23390 and clebopride, selective dopamine D1 and D2 receptor antagonists respectively. Catalepsy was measured by recording the time the animal remained with its forepaws placed over a rod 6 cm above the bench. Pretreatment with either the non-competitive NMDA receptor antagonist, MK-801 (0.25-0.5 mg/kg i.p.) or the competitive antagonist, LY274614 (10-20 mg/kg i.p.) reduced the cataleptic response produced by haloperidol (10 mg/kg), SCH23390 (2.5-10 mg/kp i.p.) or clebopride (5-20 mg/kg i.p.). This demonstrates that NMDA receptor antagonists will reduce both dopamine D1 and D2 receptor antagonist-induced catalepsy. Muscle relaxant doses of chlordiazepoxide (10 mg/kg i.p.) failed to reduce the catalepsy induced by haloperidol, suggesting that the anticataleptic effect of the NMDA receptor antagonists was not due to a non-specific action. These results support the hypothesis that NMDA receptor antagonists may have beneficial effects in disorders involving reduced dopaminergic function, such as Parkinson's disease.

  5. Dose-dependent effects of celecoxib on CB-1 agonist-induced antinociception in the mice

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Zarrindast

    2009-04-01

    Full Text Available "nObjective: Endocannabinoid produce analgesia that is comparable which of opioids. The mechanism of antinociceptive effects of (∆ - 9 tetrahydrocannabinol (THC is suggested to be through cyclooxygenase (COX pathway. In the present work, the effect of two extreme dose ranges of celecoxib (mg/kg and ng/kg, a cyclooxygenase-2 (COX-2 antagonist, on arachidonylcyclopropylamide (ACPA, a selective CB1 agonist induced antinociception in mice was examined. "nMethods: We have investigated the interaction between celecoxib, at the doses of mg/kg (50, 100, 200 and 400 i.p.  and ultra low dose (ULD (25 and 50 ng/kg, i.p., on the antinociceptive effect of intracerebroventricular (i.c.v. administration of ACPA (0.004, 0.0625 and 1 μg/mice, using formalin test in mice. "nResults: I.C.V. administration of ACPA induced antinociception. Intraperitoneal administration of celecoxib (mg/kg and its ULD (ng/kg attenuated and potentiated, ACPA antinociceptive effects, respectively. "nConclusion: It is concluded that the mg/kg doses of COX-2 antagonist showed opposite effects compare to the ultra-low dose of the drug.

  6. Inhibition of colon carcinogenesis by a standardized Cannabis sativa extract with high content of cannabidiol.

    Science.gov (United States)

    Romano, Barbara; Borrelli, Francesca; Pagano, Ester; Cascio, Maria Grazia; Pertwee, Roger G; Izzo, Angelo A

    2014-04-15

    Colon cancer is a major public health problem. Cannabis-based medicines are useful adjunctive treatments in cancer patients. Here, we have investigated the effect of a standardized Cannabis sativa extract with high content of cannabidiol (CBD), here named CBD BDS, i.e. CBD botanical drug substance, on colorectal cancer cell proliferation and in experimental models of colon cancer in vivo. Proliferation was evaluated in colorectal carcinoma (DLD-1 and HCT116) as well as in healthy colonic cells using the MTT assay. CBD BDS binding was evaluated by its ability to displace [(3)H]CP55940 from human cannabinoid CB1 and CB2 receptors. In vivo, the effect of CBD BDS was examined on the preneoplastic lesions (aberrant crypt foci), polyps and tumours induced by the carcinogenic agent azoxymethane (AOM) as well as in a xenograft model of colon cancer in mice. CBD BDS and CBD reduced cell proliferation in tumoral, but not in healthy, cells. The effect of CBD BDS was counteracted by selective CB1 and CB2 receptor antagonists. Pure CBD reduced cell proliferation in a CB1-sensitive antagonist manner only. In binding assays, CBD BDS showed greater affinity than pure CBD for both CB1 and CB2 receptors, with pure CBD having very little affinity. In vivo, CBD BDS reduced AOM-induced preneoplastic lesions and polyps as well as tumour growth in the xenograft model of colon cancer. CBD BDS attenuates colon carcinogenesis and inhibits colorectal cancer cell proliferation via CB1 and CB2 receptor activation. The results may have some clinical relevance for the use of Cannabis-based medicines in cancer patients. Copyright © 2013 Elsevier GmbH. All rights reserved.

  7. Cannabidiol inhibits paclitaxel-induced neuropathic pain through 5-HT1A receptors without diminishing nervous system function or chemotherapy efficacy

    Science.gov (United States)

    Ward, Sara Jane; McAllister, Sean D; Kawamura, Rumi; Murase, Ryuchi; Neelakantan, Harshini; Walker, Ellen A

    2014-01-01

    Background and Purpose Paclitaxel (PAC) is associated with chemotherapy-induced neuropathic pain (CIPN) that can lead to the cessation of treatment in cancer patients even in the absence of alternate therapies. We previously reported that chronic administration of the non-psychoactive cannabinoid cannabidiol (CBD) prevents PAC-induced mechanical and thermal sensitivity in mice. Hence, we sought to determine receptor mechanisms by which CBD inhibits CIPN and whether CBD negatively effects nervous system function or chemotherapy efficacy. Experimental Approach The ability of acute CBD pretreatment to prevent PAC-induced mechanical sensitivity was assessed, as was the effect of CBD on place conditioning and on an operant-conditioned learning and memory task. The potential interaction of CBD and PAC on breast cancer cell viability was determined using the MTT assay. Key Results PAC-induced mechanical sensitivity was prevented by administration of CBD (2.5 – 10 mg·kg−1) in female C57Bl/6 mice. This effect was reversed by co-administration of the 5-HT1A antagonist WAY 100635, but not the CB1 antagonist SR141716 or the CB2 antagonist SR144528. CBD produced no conditioned rewarding effects and did not affect conditioned learning and memory. Also, CBD + PAC combinations produce additive to synergistic inhibition of breast cancer cell viability. Conclusions and Implications Our data suggest that CBD is protective against PAC-induced neurotoxicity mediated in part by the 5-HT1A receptor system. Furthermore, CBD treatment was devoid of conditioned rewarding effects or cognitive impairment and did not attenuate PAC-induced inhibition of breast cancer cell viability. Hence, adjunct treatment with CBD during PAC chemotherapy may be safe and effective in the prevention or attenuation of CIPN. PMID:24117398

  8. Allosteric inhibition of SHP2 phosphatase inhibits cancers driven by receptor tyrosine kinases

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying-Nan P.; LaMarche, Matthew J.; Chan, Ho Man; Fekkes, Peter; Garcia-Fortanet, Jorge; Acker, Michael G.; Antonakos, Brandon; Chen, Christine Hiu-Tung; Chen, Zhouliang; Cooke, Vesselina G.; Dobson, Jason R.; Deng, Zhan; Fei, Feng; Firestone, Brant; Fodor, Michelle; Fridrich, Cary; Gao, Hui; Grunenfelder, Denise; Hao, Huai-Xiang; Jacob, Jaison; Ho, Samuel; Hsiao, Kathy; Kang, Zhao B.; Karki, Rajesh; Kato, Mitsunori; Larrow, Jay; La Bonte, Laura R.; Lenoir, Francois; Liu, Gang; Liu, Shumei; Majumdar, Dyuti; Meyer, Matthew J.; Palermo, Mark; Perez, Lawrence; Pu, Minying; Price, Edmund; Quinn, Christopher; Shakya, Subarna; Shultz, Michael D.; Slisz, Joanna; Venkatesan, Kavitha; Wang, Ping; Warmuth, Markus; Williams, Sarah; Yang, Guizhi; Yuan, Jing; Zhang, Ji-Hu; Zhu, Ping; Ramsey, Timothy; Keen, Nicholas J.; Sellers, William R.; Stams, Travis; Fortin , Pascal D. (Novartis)

    2016-06-29

    The non-receptor protein tyrosine phosphatase SHP2, encoded by PTPN11, has an important role in signal transduction downstream of growth factor receptor signalling and was the first reported oncogenic tyrosine phosphatase1. Activating mutations of SHP2 have been associated with developmental pathologies such as Noonan syndrome and are found in multiple cancer types, including leukaemia, lung and breast cancer and neuroblastoma1, 2, 3, 4, 5. SHP2 is ubiquitously expressed and regulates cell survival and proliferation primarily through activation of the RAS–ERK signalling pathway2, 3. It is also a key mediator of the programmed cell death 1 (PD-1) and B- and T-lymphocyte attenuator (BTLA) immune checkpoint pathways6, 7. Reduction of SHP2 activity suppresses tumour cell growth and is a potential target of cancer therapy8, 9. Here we report the discovery of a highly potent (IC50 = 0.071 μM), selective and orally bioavailable small-molecule SHP2 inhibitor, SHP099, that stabilizes SHP2 in an auto-inhibited conformation. SHP099 concurrently binds to the interface of the N-terminal SH2, C-terminal SH2, and protein tyrosine phosphatase domains, thus inhibiting SHP2 activity through an allosteric mechanism. SHP099 suppresses RAS–ERK signalling to inhibit the proliferation of receptor-tyrosine-kinase-driven human cancer cells in vitro and is efficacious in mouse tumour xenograft models. Together, these data demonstrate that pharmacological inhibition of SHP2 is a valid therapeutic approach for the treatment of cancers.

  9. Type-1 cannabinoid receptors reduce membrane fluidity of capacitated boar sperm by impairing their activation by bicarbonate.

    Directory of Open Access Journals (Sweden)

    Barbara Barboni

    Full Text Available BACKGROUND: Mammalian spermatozoa acquire their full fertilizing ability (so called capacitation within the female genital tract, where they are progressively exposed to inverse gradients of inhibiting and stimulating molecules. METHODOLOGY/PRINCIPAL FINDINGS: In the present research, the effect on this process of anandamide, an endocannabinoid that can either activate or inhibit cannabinoid receptors depending on its concentration, and bicarbonate, an oviductal activatory molecule, was assessed, in order to study the role exerted by the type 1 cannabinoid receptor (CB1R in the process of lipid membrane remodeling crucial to complete capacitation. To this aim, boar sperm were incubated in vitro under capacitating conditions (stimulated by bicarbonate in the presence or in the absence of methanandamide (Met-AEA, a non-hydrolysable analogue of anandamide. The CB1R involvement was studied by using the specific inhibitor (SR141716 or mimicking its activation by adding a permeable cAMP analogue (8Br-cAMP. By an immunocytochemistry approach it was shown that the Met-AEA inhibits the bicarbonate-dependent translocation of CB1R from the post-equatorial to equatorial region of sperm head. In addition it was found that Met-AEA is able to prevent the bicarbonate-induced increase in membrane disorder and the cholesterol extraction, both preliminary to capacitation, acting through a CB1R-cAMP mediated pathway, as indicated by MC540 and filipin staining, EPR spectroscopy and biochemical analysis on whole membranes (CB1R activity and on membrane enriched fraction (C/P content and anisotropy. CONCLUSIONS/SIGNIFICANCE: Altogether, these data demonstrate that the endocannabinoid system strongly inhibits the process of sperm capacitation, acting as membrane stabilizing agent, thus increasing the basic knowledge on capacitation-related signaling and potentially opening new perspectives in diagnostics and therapeutics of male infertility.

  10. Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Feng [Department of Gastroenterology, The Tenth People’s Hospital of Shanghai, Tongji University, Shanghai 200072 (China); Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Yang, Yong, E-mail: yyang@houstonmethodist.org [Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Department of Medicine, Weill Cornell Medical College, New York, NY 10065 (United States)

    2014-10-03

    Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers.

  11. Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

    International Nuclear Information System (INIS)

    Wang, Feng; Yang, Yong

    2014-01-01

    Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers

  12. Targeting fatty acid amide hydrolase and transient receptor potential vanilloid-1 simultaneously to modulate colonic motility and visceral sensation in the mouse: A pharmacological intervention with N-arachidonoyl-serotonin (AA-5-HT).

    Science.gov (United States)

    Bashashati, M; Fichna, J; Piscitelli, F; Capasso, R; Izzo, A A; Sibaev, A; Timmermans, J-P; Cenac, N; Vergnolle, N; Di Marzo, V; Storr, M

    2017-12-01

    Endocannabinoid anandamide (AEA) inhibits intestinal motility and visceral pain, but it may also be proalgesic through transient receptor potential vanilloid-1 (TRPV1). AEA is degraded by fatty acid amide hydrolase (FAAH). This study explored whether dual inhibition of FAAH and TRPV1 reduces diarrhea and abdominal pain. Immunostaining was performed on myenteric plexus of the mouse colon. The effects of the dual FAAH/TRPV1 inhibitor AA-5-HT on electrically induced contractility, excitatory junction potential (EJP) and fast (f) and slow (s) inhibitory junction potentials (IJP) in the mouse colon, colonic propulsion and visceromotor response (VMR) to rectal distension were studied. The colonic levels of endocannabinoids and fatty acid amides were measured. CB1-positive neurons exhibited TRPV1; only some TRPV1 positive neurons did not express CB1. CB1 and FAAH did not colocalize. AA-5-HT (100 nM-10 μM) decreased colonic contractility by ~60%; this effect was abolished by TRPV1 antagonist 5'-IRTX, but not by CB1 antagonist, SR141716. AA-5-HT (1 μM-10 μM) inhibited EJP by ~30% and IJPs by ~50%. The effects of AA-5-HT on junction potentials were reversed by SR141716 and 5`-IRTX. AA-5-HT (20 mg/kg; i.p.) inhibited colonic propulsion by ~30%; SR141716 but not 5`-IRTX reversed this effect. AA-5-HT decreased VMR by ~50%-60%; these effects were not blocked by SR141716 or 5`-IRTX. AA-5-HT increased AEA in the colon. The effects of AA-5-HT on visceral sensation and colonic motility are differentially mediated by CB1, TRPV1 and non-CB1/TRPV1 mechanisms, possibly reflecting the distinct neuromodulatory roles of endocannabinoid and endovanilloid FAAH substrates in the mouse intestine. © 2017 John Wiley & Sons Ltd.

  13. Targeting the urokinase plasminogen activator receptor inhibits ovarian cancer metastasis.

    Science.gov (United States)

    Kenny, Hilary A; Leonhardt, Payton; Ladanyi, Andras; Yamada, S Diane; Montag, Anthony; Im, Hae Kyung; Jagadeeswaran, Sujatha; Shaw, David E; Mazar, Andrew P; Lengyel, Ernst

    2011-02-01

    To understand the functional and preclinical efficacy of targeting the urokinase plasminogen activator receptor (u-PAR) in ovarian cancer. Expression of u-PAR was studied in 162 epithelial ovarian cancers, including 77 pairs of corresponding primary and metastatic tumors. The effect of an antibody against u-PAR (ATN-658) on proliferation, adhesion, invasion, apoptosis, and migration was assessed in 3 (SKOV3ip1, HeyA8, and CaOV3) ovarian cancer cell lines. The impact of the u-PAR antibody on tumor weight, number, and survival was examined in corresponding ovarian cancer xenograft models and the mechanism by which ATN-658 blocks metastasis was explored. Only 8% of all ovarian tumors were negative for u-PAR expression. Treatment of SKOV3ip1, HeyA8, and CaOV3 ovarian cancer cell lines with the u-PAR antibody inhibited cell invasion, migration, and adhesion. In vivo, anti-u-PAR treatment reduced the number of tumors and tumor weight in CaOV3 and SKOV3ip1 xenografts and reduced tumor weight and increased survival in HeyA8 xenografts. Immunostaining of CaOV3 xenograft tumors and ovarian cancer cell lines showed an increase in active-caspase 3 and TUNEL staining. Treatment with u-PAR antibody inhibited α(5)-integrin and u-PAR colocalization on primary human omental extracellular matrix. Anti-u-PAR treatment also decreased the expression of urokinase, u-PAR, β(3)-integrin, and fibroblast growth factor receptor-1 both in vitro and in vivo. This study shows that an antibody against u-PAR reduces metastasis, induces apoptosis, and reduces the interaction between u-PAR and α(5)-integrin. This provides a rationale for targeting the u-PAR pathway in patients with ovarian cancer and for further testing of ATN-658 in this indication. ©2010 AACR.

  14. Human Diversity in a Cell Surface Receptor that Inhibits Autophagy.

    Science.gov (United States)

    Chaudhary, Anu; Leite, Mara; Kulasekara, Bridget R; Altura, Melissa A; Ogahara, Cassandra; Weiss, Eli; Fu, Wenqing; Blanc, Marie-Pierre; O'Keeffe, Michael; Terhorst, Cox; Akey, Joshua M; Miller, Samuel I

    2016-07-25

    Mutations in genes encoding autophagy proteins have been associated with human autoimmune diseases, suggesting that diversity in autophagy responses could be associated with disease susceptibility or severity. A cellular genome-wide association study (GWAS) screen was performed to explore normal human diversity in responses to rapamycin, a microbial product that induces autophagy. Cells from several human populations demonstrated variability in expression of a cell surface receptor, CD244 (SlamF4, 2B4), that correlated with changes in rapamycin-induced autophagy. High expression of CD244 and receptor activation with its endogenous ligand CD48 inhibited starvation- and rapamycin-induced autophagy by promoting association of CD244 with the autophagy complex proteins Vps34 and Beclin-1. The association of CD244 with this complex reduced Vps34 lipid kinase activity. Lack of CD244 is associated with auto-antibody production in mice, and lower expression of human CD244 has previously been implicated in severity of human rheumatoid arthritis and systemic lupus erythematosus, indicating that increased autophagy as a result of low levels of CD244 may alter disease outcomes. Copyright © 2016 Elsevier Ltd. All rights reserved.

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

    DEFF Research Database (Denmark)

    Siebenmann, Christoph; Rasmussen, Peter; Sørensen, Henrik

    2015-01-01

    Hypoxia increases the heart rate (HR) response to exercise but the mechanism(s) remain 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...... combined β-adrenergic and muscarinic receptor inhibition....

  16. Palmitoylethanolamide Inhibits Glutamate Release in Rat Cerebrocortical Nerve Terminals

    Directory of Open Access Journals (Sweden)

    Tzu-Yu Lin

    2015-03-01

    Full Text Available The effect of palmitoylethanolamide (PEA, an endogenous fatty acid amide displaying neuroprotective actions, on glutamate release from rat cerebrocortical nerve terminals (synaptosomes was investigated. PEA inhibited the Ca2+-dependent release of glutamate, which was triggered by exposing synaptosomes to the potassium channel blocker 4-aminopyridine. This release inhibition was concentration dependent, associated with a reduction in cytosolic Ca2+ concentration, and not due to a change in synaptosomal membrane potential. The glutamate release-inhibiting effect of PEA was prevented by the Cav2.1 (P/Q-type channel blocker ω-agatoxin IVA or the protein kinase A inhibitor H89, not affected by the intracellular Ca2+ release inhibitors dantrolene and CGP37157, and partially antagonized by the cannabinoid CB1 receptor antagonist AM281. Based on these results, we suggest that PEA exerts its presynaptic inhibition, likely through a reduction in the Ca2+ influx mediated by Cav2.1 (P/Q-type channels, thereby inhibiting the release of glutamate from rat cortical nerve terminals. This release inhibition might be linked to the activation of presynaptic cannabinoid CB1 receptors and the suppression of the protein kinase A pathway.

  17. GABA-A Receptors Mediate Tonic Inhibition and Neurosteroid Sensitivity in the Brain.

    Science.gov (United States)

    Reddy, Doodipala Samba

    2018-01-01

    Neurosteroids like allopregnanolone (AP) are positive allosteric modulators of synaptic and extrasynaptic GABA-A receptors. AP and related neurosteroids exhibit a greater potency for δ-containing extrasynaptic receptors. The δGABA-A receptors, which are expressed extrasynaptically in the dentate gyrus and other regions, contribute to tonic inhibition, promoting network shunting as well as reducing seizure susceptibility. Levels of endogenous neurosteroids fluctuate with ovarian cycle. Natural and synthetic neurosteroids maximally potentiate tonic inhibition in the hippocampus and provide robust protection against a variety of limbic seizures and status epilepticus. Recently, a consensus neurosteroid pharmacophore model has been proposed at extrasynaptic δGABA-A receptors based on structure-activity relationship for functional activation of tonic currents and seizure protection. Aside from anticonvulsant actions, neurosteroids have been found to be powerful anxiolytic and anesthetic agents. Neurosteroids and Zn 2+ have preferential affinity for δ-containing receptors. Thus, Zn 2+ can prevent neurosteroid activation of extrasynaptic δGABA-A receptor-mediated tonic inhibition. Recently, we demonstrated that Zn 2+ selectively inhibits extrasynaptic δGABA-A receptors and thereby fully prevents AP activation of tonic inhibition and seizure protection. We confirmed that neurosteroids exhibit greater sensitivity at extrasynaptic δGABA-A receptors. Overall, extrasynaptic GABA-A receptors are primary mediators of tonic inhibition in the brain and play a key role in the pathophysiology of epilepsy and other neurological disorders. © 2018 Elsevier Inc. All rights reserved.

  18. Sigma-1 receptor agonists directly inhibit Nav1.2/1.4 channels.

    Directory of Open Access Journals (Sweden)

    Xiao-Fei Gao

    Full Text Available (+-SKF 10047 (N-allyl-normetazocine is a prototypic and specific sigma-1 receptor agonist that has been used extensively to study the function of sigma-1 receptors. (+-SKF 10047 inhibits K(+, Na(+ and Ca2+ channels via sigma-1 receptor activation. We found that (+-SKF 10047 inhibited Na(V1.2 and Na(V1.4 channels independently of sigma-1 receptor activation. (+-SKF 10047 equally inhibited Na(V1.2/1.4 channel currents in HEK293T cells with abundant sigma-1 receptor expression and in COS-7 cells, which barely express sigma-1 receptors. The sigma-1 receptor antagonists BD 1063,BD 1047 and NE-100 did not block the inhibitory effects of (+-SKF-10047. Blocking of the PKA, PKC and G-protein pathways did not affect (+-SKF 10047 inhibition of Na(V1.2 channel currents. The sigma-1 receptor agonists Dextromethorphan (DM and 1,3-di-o-tolyl-guanidine (DTG also inhibited Na(V1.2 currents through a sigma-1 receptor-independent pathway. The (+-SKF 10047 inhibition of Na(V1.2 currents was use- and frequency-dependent. Point mutations demonstrated the importance of Phe(1764 and Tyr(1771 in the IV-segment 6 domain of the Na(V1.2 channel and Phe(1579 in the Na(V1.4 channel for (+-SKF 10047 inhibition. In conclusion, our results suggest that sigma-1 receptor agonists directly inhibit Na(V1.2/1.4 channels and that these interactions should be given special attention for future sigma-1 receptor function studies.

  19. Sulindac metabolites inhibit epidermal growth factor receptor activation and expression

    Directory of Open Access Journals (Sweden)

    Ahnen Dennis

    2005-01-01

    Full Text Available Abstract Background Regular use of nonsteroidal anti-inflammatory drugs (NSAIDs is associated with a decreased mortality from colorectal cancer (CRC. NSAIDs induce apoptotic cell death in colon cancer cells in vitro and inhibit growth of neoplastic colonic mucosa in vivo however, the biochemical mechanisms required for these growth inhibitory effects are not well defined. We previously reported that metabolites of the NSAID sulindac downregulate extracellular-signal regulated kinase 1/2 (ERK1/2 signaling and that this effect is both necessary and sufficient for the apoptotic effects of these drugs. The goal of this project was to specifically test the hypothesis that sulindac metabolites block activation and/or expression of the epidermal growth factor (EGF receptor (EGFR. Methods HT29 human colon cancer cells were treated with EGF, alone, or in the presence of sulindac sulfide or sulindac sulfone. Cells lysates were assayed by immunoblotting for phosphorylated EGFR (pEGFR, pY1068, total EGFR, phosphorylated ERK1/2 (pERK1/2, total ERK1/2, activated caspase-3, and α-tubulin. Results EGF treatment rapidly induced phosphorylation of both EGFR and ERK1/2 in HT29 colon cancer cells. Pretreatment with sulindac metabolites for 24 h blocked EGF-induced phosphorylation of both EGFR and ERK1/2 and decreased total EGFR protein expression. Under basal conditions, downregulation of pEGFR and total EGFR was detected as early as 12 h following sulindac sulfide treatment and persisted through at least 48 h. Sulindac sulfone induced downregulation of pEGFR and total EGFR was detected as early as 1 h and 24 h, respectively, following drug treatment, and persisted through at least 72 h. EGFR downregulation by sulindac metabolites was observed in three different CRC cell lines, occurred prior to the observed downregulation of pERK1/2 and induction of apoptosis by these drugs, and was not dependent of caspase activation. Conclusion These results suggest that

  20. Metabotropic glutamate receptor I (mGluR1) antagonism impairs cocaine-induced conditioned place preference via inhibition of protein synthesis.

    Science.gov (United States)

    Yu, Fei; Zhong, Peng; Liu, Xiaojie; Sun, Dalong; Gao, Hai-Qing; Liu, Qing-Song

    2013-06-01

    Antagonism of group I metabotropic glutamate receptors (mGluR1 and mGluR5) reduces behavioral effects of drugs of abuse, including cocaine. However, the underlying mechanisms remain poorly understood. Activation of mGluR5 increases protein synthesis at synapses. Although mGluR5-induced excessive protein synthesis has been implicated in the pathology of fragile X syndrome, it remains unknown whether group I mGluR-mediated protein synthesis is involved in any behavioral effects of drugs of abuse. We report that group I mGluR agonist DHPG induced more pronounced initial depression of inhibitory postsynaptic currents (IPSCs) followed by modest long-term depression (I-LTD) in dopamine neurons of rat ventral tegmental area (VTA) through the activation of mGluR1. The early component of DHPG-induced depression of IPSCs was mediated by the cannabinoid CB1 receptors, while DHPG-induced I-LTD was dependent on protein synthesis. Western blotting analysis indicates that mGluR1 was coupled to extracellular signal-regulated kinase (ERK) and mammalian target of rapamycin (mTOR) signaling pathways to increase translation. We also show that cocaine conditioning activated translation machinery in the VTA via an mGluR1-dependent mechanism. Furthermore, intra-VTA microinjections of mGluR1 antagonist JNJ16259685 and protein synthesis inhibitor cycloheximide significantly attenuated or blocked the acquisition of cocaine-induced conditioned place preference (CPP) and activation of translation elongation factors. Taken together, these results suggest that mGluR1 antagonism inhibits de novo protein synthesis; this effect may block the formation of cocaine-cue associations and thus provide a mechanism for the reduction in CPP to cocaine.

  1. Phenobarbital indirectly activates the constitutive active androstane receptor (CAR) by inhibition of epidermal growth factor receptor signaling.

    Science.gov (United States)

    Mutoh, Shingo; Sobhany, Mack; Moore, Rick; Perera, Lalith; Pedersen, Lee; Sueyoshi, Tatsuya; Negishi, Masahiko

    2013-05-07

    Phenobarbital is a central nervous system depressant that also indirectly activates nuclear receptor constitutive active androstane receptor (CAR), which promotes drug and energy metabolism, as well as cell growth (and death), in the liver. We found that phenobarbital activated CAR by inhibiting epidermal growth factor receptor (EGFR) signaling. Phenobarbital bound to EGFR and potently inhibited the binding of EGF, which prevented the activation of EGFR. This abrogation of EGFR signaling induced the dephosphorylation of receptor for activated C kinase 1 (RACK1) at Tyr(52), which then promoted the dephosphorylation of CAR at Thr(38) by the catalytic core subunit of protein phosphatase 2A. The findings demonstrated that the phenobarbital-induced mechanism of CAR dephosphorylation and activation is mediated through its direct interaction with and inhibition of EGFR.

  2. G-protein coupling of cannabinoid receptors

    International Nuclear Information System (INIS)

    Glass, M.

    2001-01-01

    Full text: Since the cloning of the cannabinoid CB1 and CB2 receptors in the early 1990's extensive research has focused on understanding their signal transduction pathways. While it has been known for sometime that both receptors can couple to intracellular signalling via pertussis toxin sensitive G-proteins (Gi/Go), the specificity and kinetics of these interactions have only recently been elucidated. We have developed an in situ reconstitution approach to investigating receptor-G-protein interactions. This approach involves chaotropic extraction of receptor containing membranes in order to inactivate or remove endogenous G-proteins. Recombinant or isolated brain G-proteins can then be added back to the receptors, and their activation monitored through the binding of [ 35 S]-GTPγS. This technique has been utilised for an extensive study of cannabinoid receptor mediated activation of G-proteins. In these studies we have established that CB1 couples with high affinity to both Gi and Go type G-proteins. In contrast, CB2 couples strongly to Gi, but has a very low affinity for Go. This finding correlated well with the previous findings that while CB1 and CB2 both couple to the inhibition of adenylate cyclase, CB1 but not CB2 could also inhibit calcium channels. We then examined the ability of a range of cannabinoid agonists to activate the Gi and Go via CB1. Conventional receptor theory suggests that a receptor is either active or inactive with regard to a G-protein and that the active receptor activates all relevant G-proteins equally. However, in this study we found that agonists could produce different degrees of activation, depending on which G-protein was present. Further studies have compared the ability of the two endocannabinoids to drive the activation of Gi or Go. These studies show that agonists can induce multiple forms of activated receptor that differ in their ability to catalyse the activation of Gi or Go. The ability of an agonist to drive a receptor

  3. Mitochondrial ADP/ATP exchange inhibition: a novel off-target mechanism underlying ibipinabant-induced myotoxicity.

    Science.gov (United States)

    Schirris, Tom J J; Ritschel, Tina; Herma Renkema, G; Willems, Peter H G M; Smeitink, Jan A M; Russel, Frans G M

    2015-09-29

    Cannabinoid receptor 1 (CB1R) antagonists appear to be promising drugs for the treatment of obesity, however, serious side effects have hampered their clinical application. Rimonabant, the first in class CB1R antagonist, was withdrawn from the market because of psychiatric side effects. This has led to the search for more peripherally restricted CB1R antagonists, one of which is ibipinabant. However, this 3,4-diarylpyrazoline derivative showed muscle toxicity in a pre-clinical dog study with mitochondrial dysfunction. Here, we studied the molecular mechanism by which ibipinabant induces mitochondrial toxicity. We observed a strong cytotoxic potency of ibipinabant in C2C12 myoblasts. Functional characterization of mitochondria revealed increased cellular reactive oxygen species generation and a decreased ATP production capacity, without effects on the catalytic activities of mitochondrial enzyme complexes I-V or the complex specific-driven oxygen consumption. Using in silico off-target prediction modelling, combined with in vitro validation in isolated mitochondria and mitoplasts, we identified adenine nucleotide translocase (ANT)-dependent mitochondrial ADP/ATP exchange as a novel molecular mechanism underlying ibipinabant-induced toxicity. Minor structural modification of ibipinabant could abolish ANT inhibition leading to a decreased cytotoxic potency, as observed with the ibipinabant derivative CB23. Our results will be instrumental in the development of new types of safer CB1R antagonists.

  4. Differences in receptor binding affinity of several phytocannabinoids do not explain their effects on neural cell cultures.

    Science.gov (United States)

    Rosenthaler, Sarah; Pöhn, Birgit; Kolmanz, Caroline; Huu, Chi Nguyen; Krewenka, Christopher; Huber, Alexandra; Kranner, Barbara; Rausch, Wolf-Dieter; Moldzio, Rudolf

    2014-01-01

    Phytocannabinoids are potential candidates for neurodegenerative disease treatment. Nonetheless, the exact mode of action of major phytocannabinoids has to be elucidated, but both, receptor and non-receptor mediated effects are discussed. Focusing on the often presumed structure-affinity-relationship, Ki values of phytocannabinoids cannabidiol (CBD), cannabidivarin (CBDV), cannabichromene (CBC), cannabigerol (CBG), cannabinol (CBN), THC acid (THCA) and THC to human CB1 and CB2 receptors were detected by using competitive inhibition between radioligand [(3)H]CP-55,940 and the phytocannabinoids. The resulting Ki values to CB1 range from 23.5 nM (THCA) to 14711 nM (CBDV), whereas Ki values to CB2 range from 8.5 nM (THC) to 574.2 nM (CBDV). To study the relationship between binding affinity and effects on neurons, we investigated possible CB1 related cytotoxic properties in murine mesencephalic primary cell cultures and N18TG2 neuroblastoma cell line. Most of the phytocannabinoids did not affect the number of dopaminergic neurons in primary cultures, whereas propidium iodide and resazurin formation assays revealed cytotoxic properties of CBN, CBDV and CBG. However, THC showed positive effects on N18TG2 cell viability at a concentration of 10 μM, whereas CBC and THCA also displayed slightly positive activities. These findings are not linked to the receptor binding affinity therewith pointing to another mechanism than a receptor mediated one. [Corrected] Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Structural basis for vinyl sulfone inhibition of the SmCB1 drug target from the human blood fluke

    Czech Academy of Sciences Publication Activity Database

    Jílková, Adéla; Marešová, Lucie; Rubešová, Petra; Fajtová, Pavla; Horn, Martin; Řezáčová, Pavlína; Brynda, Jiří; McKerrow, J. H.; Caffrey, C. R.; Mareš, Michael

    2017-01-01

    Roč. 284, Suppl 1 (2017), s. 191 ISSN 1742-464X. [FEBS Congress /42./ From Molecules to Cells and Back. 10.09.2017-14.09.2017, Jerusalem] Institutional support: RVO:61388963 Keywords : schistosomiasis * cathepsin B1 * inhibitor Subject RIV: CE - Biochemistry

  6. Competitive inhibition of [3H]dexamethasone binding to mammary glucocorticoid receptor by leupeptin

    International Nuclear Information System (INIS)

    Hsieh, L.C.C.; Su, C.; Markland, F.S. Jr.

    1987-01-01

    The inhibitory effect of leupeptin on [ 3 H]dexamethasone binding to the glucocorticoid receptor from lactating goat mammary cytosol has been studied. Leupeptin (10 mM) caused a significant (about 35%) inhibition of [ 3 H]dexamethasone binding to glucocorticoid receptor. Binding inhibition is further increased following filtration of unlabeled cytosolic receptor through a Bio-Gel A 0.5-m column. Binding inhibition was partially reversed by monothioglycerol at 10 mM concentration. A double reciprocal plot revealed that leupeptin appears to be a competitive inhibitor of [ 3 H]dexamethasone binding to the glucocorticoid receptor. Low salt sucrose density gradient centrifugation revealed that the leupeptin-treated sample formed a slightly larger (approximately 9 S) receptor complex (leupeptin-free complex sediments at 8 S)

  7. Biased Type 1 Cannabinoid Receptor Signaling Influences Neuronal Viability in a Cell Culture Model of Huntington Disease.

    Science.gov (United States)

    Laprairie, Robert B; Bagher, Amina M; Kelly, Melanie E M; Denovan-Wright, Eileen M

    2016-03-01

    Huntington disease (HD) is an inherited, autosomal dominant, neurodegenerative disorder with limited treatment options. Prior to motor symptom onset or neuronal cell loss in HD, levels of the type 1 cannabinoid receptor (CB1) decrease in the basal ganglia. Decreasing CB1 levels are strongly correlated with chorea and cognitive deficit. CB1 agonists are functionally selective (biased) for divergent signaling pathways. In this study, six cannabinoids were tested for signaling bias in in vitro models of medium spiny projection neurons expressing wild-type (STHdh(Q7/Q7)) or mutant huntingtin protein (STHdh(Q111/Q111)). Signaling bias was assessed using the Black and Leff operational model. Relative activity [ΔlogR (τ/KA)] and system bias (ΔΔlogR) were calculated relative to the reference compound WIN55,212-2 for Gαi/o, Gαs, Gαq, Gβγ, and β-arrestin1 signaling following treatment with 2-arachidonoylglycerol (2-AG), anandamide (AEA), CP55,940, Δ(9)-tetrahydrocannabinol (THC), cannabidiol (CBD), and THC+CBD (1:1), and compared between wild-type and HD cells. The Emax of Gαi/o-dependent extracellular signal-regulated kinase (ERK) signaling was 50% lower in HD cells compared with wild-type cells. 2-AG and AEA displayed Gαi/o/Gβγ bias and normalized CB1 protein levels and improved cell viability, whereas CP55,940 and THC displayed β-arrestin1 bias and reduced CB1 protein levels and cell viability in HD cells. CBD was not a CB1 agonist but inhibited THC-dependent signaling (THC+CBD). Therefore, enhancing Gαi/o-biased endocannabinoid signaling may be therapeutically beneficial in HD. In contrast, cannabinoids that are β-arrestin-biased--such as THC found at high levels in modern varieties of marijuana--may be detrimental to CB1 signaling, particularly in HD where CB1 levels are already reduced. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

  8. Effects of Chronic Alcohol Exposure on the Modulation of Ischemia-Induced Glutamate Release via Cannabinoid Receptors in the Dorsal Hippocampus.

    Science.gov (United States)

    Zheng, Lei; Wu, Xiaoda; Dong, Xiao; Ding, Xinli; Song, Cunfeng

    2015-10-01

    Chronic alcohol consumption is a critical contributing factor to ischemic stroke, as it enhances ischemia-induced glutamate release, leading to more severe excitotoxicity and brain damage. But the neural mechanisms underlying this phenomenon are poorly understood. We evaluated the effects of chronic alcohol exposure on the modulation of ischemia-induced glutamate release via CB1 and CB2 cannabinoid receptors during middle cerebral artery occlusion, using in vivo microdialysis coupled with high-performance liquid chromatography, in alcohol-naïve rats or rats after 1 or 30 days of withdrawal from chronic ethanol intake (6% v/v for 14 days). Intra-dorsal hippocampus (DH) infusions of ACEA or JWH133, selective CB1 or CB2 receptor agonists, respectively, decreased glutamate release in the DH in alcohol-naïve rats in a dose-dependent manner. Such an effect was reversed by co-infusions of SR141716A or AM630, selective CB1 or CB2 receptor antagonists, respectively. After 30 days, but not 1 day of withdrawal, ischemia induced an enhancement in glutamate release in the DH, as compared with non-alcohol-treated control group. Intra-DH infusions of JWH133, but not ACEA, inhibited ischemia-induced glutamate release in the DH after 30 days of withdrawal. Finally, 1 day of withdrawal did not alter the protein level of CB1 or CB2 receptors in the DH, as compared to non-alcohol-treated control rats. Whereas 30 days of withdrawal robustly decreased the protein level of CB1 receptors, but failed to alter the protein level of CB2 receptors, in the DH, as compared to non-alcohol-treated control rats. Together, these findings suggest that loss of expression/function of CB1 receptors, but not CB2 receptors in the DH, is correlated with the enhancement of ischemia-induced glutamate release after prolonged alcohol withdrawal. Copyright © 2015 by the Research Society on Alcoholism.

  9. The anabolic steroid nandrolone alters cannabinoid self-administration and brain CB1 receptor density and function

    NARCIS (Netherlands)

    Struik, Dicky; Fadda, Paola; Zara, Tamara; Zamberletti, Erica; Rubino, Tiziana; Parolaro, Daniela; Fratta, Walter; Fattore, Liana

    Clinical and pre-clinical observations indicate that anabolic-androgenic steroids can induce neurobiological changes that alter the rewarding effects of drugs of abuse. In this study, we investigated the effect of the anabolic steroid nandrolone on the rewarding properties of the cannabinoid CBI

  10. Endocannabinoids shape accumbal encoding of cue-motivated behavior via CB1 receptor activation in the ventral tegmentum.

    Science.gov (United States)

    Oleson, Erik B; Beckert, Michael V; Morra, Joshua T; Lansink, Carien S; Cachope, Roger; Abdullah, Rehab A; Loriaux, Amy L; Schetters, Dustin; Pattij, Tommy; Roitman, Mitchell F; Lichtman, Aron H; Cheer, Joseph F

    2012-01-26

    Transient increases in nucleus accumbens (NAc) dopamine concentration are observed when animals are presented with motivationally salient stimuli and are theorized to energize reward seeking. They arise from high-frequency firing of dopamine neurons in the ventral tegmental area (VTA), which also results in the release of endocannabinoids from dopamine cell bodies. In this context, endocannabinoids are thought to regulate reward seeking by modulating dopamine signaling, although a direct link has never been demonstrated. To test this, we pharmacologically manipulated endocannabinoid neurotransmission in the VTA while measuring transient changes in dopamine concentration in the NAc during reward seeking. Disrupting endocannabinoid signaling dramatically reduced, whereas augmenting levels of the endocannabinoid 2-arachidonoylglycerol (2AG) increased, cue-evoked dopamine concentrations and reward seeking. These data suggest that 2AG in the VTA regulates reward seeking by sculpting ethologically relevant patterns of dopamine release during reward-directed behavior. Copyright © 2012 Elsevier Inc. All rights reserved.

  11. Cannabinoid CB1 receptor agonists do not decrease, but may increase, acoustic trauma-induced tinnitus in rats

    Directory of Open Access Journals (Sweden)

    Yiwen eZheng

    2015-03-01

    Full Text Available Tinnitus has been suggested to arise from neuronal hyperactivity in auditory areas of the brain and anti-epileptic drugs are sometimes used to provide relief from tinnitus. Recently, the anti-epileptic properties of the cannabinoid drugs have gained increasing interest; however, the use of cannabinoids as a form of treatment for tinnitus is controversial. In the present study, we tested whether a combination of delta-9-tetrahydrocannabinol (delta-9-THC and cannabidiol (CBD, delivered in a 1:1 ratio, could affect tinnitus perception in a rat model of acoustic trauma-induced tinnitus. Following sham treatment or acoustic trauma, the animals were divided into the following groups: 1 sham (i.e. no acoustic trauma with vehicle treatment; 2 sham with drug treatment (i.e. delta-9-THC + CBD; 3 acoustic trauma-exposed exhibiting tinnitus, with drug treatment; and 4 acoustic trauma-exposed exhibiting no tinnitus, with drug treatment. The animals received either the vehicle or the cannabinoid drugs every day, 30 min before the tinnitus behavioural testing. Acoustic trauma caused a significant increase in the auditory brainstem response (ABR thresholds in the exposed animals, indicating hearing loss; however, there was a partial recovery over 6 months. Acoustic trauma did not always result in tinnitus; however among those that did exhibit tinnitus, some of them had tinnitus at multiple frequencies while others had it only at a single frequency. The cannabinoids significantly increased the number of tinnitus animals in the exposed-tinnitus group, but not in the sham group. The results suggest that cannabinoids may promote the development of tinnitus, especially when there is pre-existing hearing damage.

  12. Effects of amphetamine on dopamine release in the rat nucleus accumbens shell region depend on cannabinoid CB1 receptor activation

    NARCIS (Netherlands)

    Kleijn, J.; Wiskerke, J.; Cremers, T.I.F.H.; Schoffelmeer, A.N.M.; Westerink, B.H.C.; Pattij, T.

    2012-01-01

    The psychostimulant drug amphetamine is often prescribed to treat Attention-Deficit/Hyperactivity Disorder. The behavioral effects of the psychostimulant drug amphetamine depend on its ability to increase monoamine neurotransmission in brain regions such as the nucleus accumbens (NAC) and medial

  13. A bacterial tyrosine phosphatase inhibits plant pattern recognition receptor activation

    Science.gov (United States)

    Perception of pathogen-associated molecular patterns (PAMPs) by surface-localised pattern-recognition receptors (PRRs) is a key component of plant innate immunity. Most known plant PRRs are receptor kinases and initiation of PAMP-triggered immunity (PTI) signalling requires phosphorylation of the PR...

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

    Science.gov (United States)

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

    2005-11-21

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

  15. Increased NMDA receptor inhibition at an increased Sevoflurane MAC

    Directory of Open Access Journals (Sweden)

    Brosnan Robert J

    2012-06-01

    Full Text Available Abstract Background Sevoflurane potently enhances glycine receptor currents and more modestly decreases NMDA receptor currents, each of which may contribute to immobility. This modest NMDA receptor antagonism by sevoflurane at a minimum alveolar concentration (MAC could be reciprocally related to large potentiation of other inhibitory ion channels. If so, then reduced glycine receptor potency should increase NMDA receptor antagonism by sevoflurane at MAC. Methods Indwelling lumbar subarachnoid catheters were surgically placed in 14 anesthetized rats. Rats were anesthetized with sevoflurane the next day, and a pre-infusion sevoflurane MAC was measured in duplicate using a tail clamp method. Artificial CSF (aCSF containing either 0 or 4 mg/mL strychnine was then infused intrathecally at 4 μL/min, and the post-infusion baseline sevoflurane MAC was measured. Finally, aCSF containing strychnine (either 0 or 4 mg/mL plus 0.4 mg/mL dizocilpine (MK-801 was administered intrathecally at 4 μL/min, and the post-dizocilpine sevoflurane MAC was measured. Results Pre-infusion sevoflurane MAC was 2.26%. Intrathecal aCSF alone did not affect MAC, but intrathecal strychnine significantly increased sevoflurane requirement. Addition of dizocilpine significantly decreased MAC in all rats, but this decrease was two times larger in rats without intrathecal strychnine compared to rats with intrathecal strychnine, a statistically significant (P  Conclusions Glycine receptor antagonism increases NMDA receptor antagonism by sevoflurane at MAC. The magnitude of anesthetic effects on a given ion channel may therefore depend on the magnitude of its effects on other receptors that modulate neuronal excitability.

  16. Intracellular postsynaptic cannabinoid receptors link thyrotropin-releasing hormone receptors to TRPC-like channels in thalamic paraventricular nucleus neurons.

    Science.gov (United States)

    Zhang, L; Kolaj, M; Renaud, L P

    2015-12-17

    In rat thalamic paraventricular nucleus of thalamus (PVT) neurons, activation of thyrotropin-releasing hormone (TRH) receptors enhances excitability via concurrent decrease in G protein-coupled inwardly-rectifying potassium (GIRK)-like and activation of transient receptor potential cation (TRPC)4/5-like cationic conductances. An exploration of intracellular signaling pathways revealed the TRH-induced current to be insensitive to phosphatidylinositol-specific phospholipase C (PI-PLC) inhibitors, but reduced by D609, an inhibitor of phosphatidylcholine-specific PLC (PC-PLC). A corresponding change in the I-V relationship implied suppression of the cationic component of the TRH-induced current. Diacylglycerol (DAG) is a product of the hydrolysis of PC. Studies focused on the isolated cationic component of the TRH-induced response revealed a reduction by RHC80267, an inhibitor of DAG lipase, the enzyme involved in the hydrolysis of DAG to the endocannabinoid 2-arachidonoylglycerol (2-AG). Further investigation revealed enhancement of the cationic component in the presence of either JZL184 or WWL70, inhibitors of enzymes involved in the hydrolysis of 2-AG. A decrease in the TRH-induced response was noted in the presence of rimonabant or SR144528, membrane permeable CB1 and CB2 receptor antagonists, respectively. A decrease in the TRH-induced current by intracellular, but not by bath application of the membrane impermeable peptide hemopressin, selective for CB1 receptors, suggests a postsynaptic intracellular localization of these receptors. The TRH-induced current was increased in the presence of arachidonyl-2'-chloroethylamide (ACEA) or JWH133, CB1 and CB2 receptor agonists, respectively. The PI3-kinase inhibitor LY294002, known to inhibit TRPC translocation, decreased the response to TRH. In addition, a TRH-induced enhancement of the low-threshold spike was prevented by both rimonabant, and SR144528. TRH had no influence on excitatory or inhibitory miniature

  17. Cannabinoid receptor activation in the rostral ventrolateral medulla oblongata evokes cardiorespiratory effects in anaesthetised rats

    Science.gov (United States)

    Padley, James R; Li, Qun; Pilowsky, Paul M; Goodchild, Ann K

    2003-01-01

    The nature of the cardiorespiratory effects mediated by cannabinoids in the hindbrain is poorly understood. In the present study we investigated whether cannabinoid receptor activation in the rostral ventrolateral medulla oblongata (RVLM) affects cardiovascular and/or respiratory function. Initially, we looked for evidence of CB1 receptor gene expression in rostral and caudal sections of the rat ventrolateral medulla (VLM) using reverse transcription–polymerase chain reaction. Second, the potent cannabinoid receptor agonists WIN55,212-2 (0.05, 0.5 or 5 pmol per 50 nl) and HU-210 (0.5 pmol per 50 nl) or the CB1 receptor antagonist/inverse agonist AM281 (1 pmol per 100 nl) were microinjected into the RVLM of urethane-anaesthetised, immobilised and mechanically ventilated male Sprague–Dawley rats (n=22). Changes in splanchnic nerve activity (sSNA), phrenic nerve activity (PNA), mean arterial pressure (MAP) and heart rate (HR) in response to cannabinoid administration were recorded. The CB1 receptor gene was expressed throughout the VLM. Unilateral microinjection of WIN55,212-2 into the RVLM evoked short-latency, dose-dependent increases in sSNA (0.5 pmol; 175±8%, n=5) and MAP (0.5 pmol; 26±3%, n=8) and abolished PNA (0.5 pmol; duration of apnoea: 5.4±0.4 s, n=8), with little change in HR (P<0.005). HU-210, structurally related to Δ9-tetrahydrocannabinol (THC), evoked similar effects when microinjected into the RVLM (n=4). Surprisingly, prior microinjection of AM281 produced agonist-like effects, as well as significantly attenuated the response to subsequent injection of WIN55,212-2 (0.5 pmol, n=4). The present study reveals CB1 receptor gene expression in the rat VLM and demonstrates sympathoexcitation, hypertension and respiratory inhibition in response to RVLM-administered cannabinoids. These findings suggest a novel link between CB1 receptors in this region of the hindbrain and the central cardiorespiratory effects of cannabinoids. The extent to which these

  18. Pumpkin seed extract: Cell growth inhibition of hyperplastic and cancer cells, independent of steroid hormone receptors.

    Science.gov (United States)

    Medjakovic, Svjetlana; Hobiger, Stefanie; Ardjomand-Woelkart, Karin; Bucar, Franz; Jungbauer, Alois

    2016-04-01

    Pumpkin seeds have been known in folk medicine as remedy for kidney, bladder and prostate disorders since centuries. Nevertheless, pumpkin research provides insufficient data to back up traditional beliefs of ethnomedical practice. The bioactivity of a hydro-ethanolic extract of pumpkin seeds from the Styrian pumpkin, Cucurbita pepo L. subsp. pepo var. styriaca, was investigated. As pumpkin seed extracts are standardized to cucurbitin, this compound was also tested. Transactivational activity was evaluated for human androgen receptor, estrogen receptor and progesterone receptor with in vitro yeast assays. Cell viability tests with prostate cancer cells, breast cancer cells, colorectal adenocarcinoma cells and a hyperplastic cell line from benign prostate hyperplasia tissue were performed. As model for non-hyperplastic cells, effects on cell viability were tested with a human dermal fibroblast cell line (HDF-5). No transactivational activity was found for human androgen receptor, estrogen receptor and progesterone receptor, for both, extract and cucurbitin. A cell growth inhibition of ~40-50% was observed for all cell lines, with the exception of HDF-5, which showed with ~20% much lower cell growth inhibition. Given the receptor status of some cell lines, a steroid-hormone receptor independent growth inhibiting effect can be assumed. The cell growth inhibition for fast growing cells together with the cell growth inhibition of prostate-, breast- and colon cancer cells corroborates the ethnomedical use of pumpkin seeds for a treatment of benign prostate hyperplasia. Moreover, due to the lack of androgenic activity, pumpkin seed applications can be regarded as safe for the prostate. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  19. Stimulation of accumbal GABAA receptors inhibits delta2-, but not delta1-, opioid receptor-mediated dopamine efflux in the nucleus accumbens of freely moving rats.

    Science.gov (United States)

    Aono, Yuri; Kiguchi, Yuri; Watanabe, Yuriko; Waddington, John L; Saigusa, Tadashi

    2017-11-15

    The nucleus accumbens contains delta-opioid receptors that may reduce inhibitory neurotransmission. Reduction in GABA A receptor-mediated inhibition of accumbal dopamine release due to delta-opioid receptor activation should be suppressed by stimulating accumbal GABA A receptors. As delta-opioid receptors are divided into delta2- and delta1-opioid receptors, we analysed the effects of the GABA A receptor agonist muscimol on delta2- and delta1-opioid receptor-mediated accumbal dopamine efflux in freely moving rats using in vivo microdialysis. Drugs were administered intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 25-50min infusions. The delta2-opioid receptor agonist deltorphin II (25.0nmol)- and delta1-opioid receptor agonist DPDPE (5.0nmol)-induced increases in dopamine efflux were inhibited by the delta2-opioid receptor antagonist naltriben (1.5nmol) and the delta1-opioid receptor antagonist BNTX (150.0pmol), respectively. Muscimol (250.0pmol) inhibited deltorphin II (25.0nmol)-induced dopamine efflux. The GABA A receptor antagonist bicuculline (50.0pmol), which failed to affect deltorphin II (25.0nmol)-induced dopamine efflux, counteracted the inhibitory effect of muscimol on deltorphin II-induced dopamine efflux. Neither muscimol (250.0pmol) nor bicuculline (50.0 and 500.0pmol) altered DPDPE (5.0nmol)-induced dopamine efflux. The present results show that reduction in accumbal GABA A receptor-mediated inhibition of dopaminergic activity is necessary to produce delta2-opioid receptor-induced increase in accumbal dopamine efflux. This study indicates that activation of delta2- but not delta1-opioid receptors on the cell bodies and/or terminals of accumbal GABAergic interneurons inhibits GABA release and, accordingly, decreases GABA A receptor-mediated inhibition of dopaminergic terminals, resulting in enhanced accumbal dopamine efflux. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Azadirachtin interacts with retinoic acid receptors and inhibits retinoic acid-mediated biological responses.

    Science.gov (United States)

    Thoh, Maikho; Babajan, Banaganapalli; Raghavendra, Pongali B; Sureshkumar, Chitta; Manna, Sunil K

    2011-02-11

    Considering the role of retinoids in regulation of more than 500 genes involved in cell cycle and growth arrest, a detailed understanding of the mechanism and its regulation is useful for therapy. The extract of the medicinal plant Neem (Azadirachta indica) is used against several ailments especially for anti-inflammatory, anti-itching, spermicidal, anticancer, and insecticidal activities. In this report we prove the detailed mechanism on the regulation of retinoic acid-mediated cell signaling by azadirachtin, active components of neem extract. Azadirachtin repressed all trans-retinoic acid (ATRA)-mediated nuclear transcription factor κB (NF-κB) activation, not the DNA binding but the NF-κB-dependent gene expression. It did not inhibit IκBα degradation, IκBα kinase activity, or p65 phosphorylation and its nuclear translocation but inhibited NF-κB-dependent reporter gene expression. Azadirachtin inhibited TRAF6-mediated, but not TRAF2-mediated NF-κB activation. It inhibited ATRA-induced Sp1 and CREB (cAMP-response element-binding protein) DNA binding. Azadirachtin inhibited ATRA binding with retinoid receptors, which is supported by biochemical and in silico evidences. Azadirachtin showed strong interaction with retinoid receptors. It suppressed ATRA-mediated removal of retinoid receptors, bound with DNA by inhibiting ATRA binding to its receptors. Overall, our data suggest that azadirachtin interacts with retinoic acid receptors and suppresses ATRA binding, inhibits falling off the receptors, and activates transcription factors like CREB, Sp1, NF-κB, etc. Thus, azadirachtin exerts anti-inflammatory and anti-metastatic responses by a novel pathway that would be beneficial for further anti-inflammatory and anti-cancer therapies.

  1. Azadirachtin Interacts with Retinoic Acid Receptors and Inhibits Retinoic Acid-mediated Biological Responses*

    Science.gov (United States)

    Thoh, Maikho; Babajan, Banaganapalli; Raghavendra, Pongali B.; Sureshkumar, Chitta; Manna, Sunil K.

    2011-01-01

    Considering the role of retinoids in regulation of more than 500 genes involved in cell cycle and growth arrest, a detailed understanding of the mechanism and its regulation is useful for therapy. The extract of the medicinal plant Neem (Azadirachta indica) is used against several ailments especially for anti-inflammatory, anti-itching, spermicidal, anticancer, and insecticidal activities. In this report we prove the detailed mechanism on the regulation of retinoic acid-mediated cell signaling by azadirachtin, active components of neem extract. Azadirachtin repressed all trans-retinoic acid (ATRA)-mediated nuclear transcription factor κB (NF-κB) activation, not the DNA binding but the NF-κB-dependent gene expression. It did not inhibit IκBα degradation, IκBα kinase activity, or p65 phosphorylation and its nuclear translocation but inhibited NF-κB-dependent reporter gene expression. Azadirachtin inhibited TRAF6-mediated, but not TRAF2-mediated NF-κB activation. It inhibited ATRA-induced Sp1 and CREB (cAMP-response element-binding protein) DNA binding. Azadirachtin inhibited ATRA binding with retinoid receptors, which is supported by biochemical and in silico evidences. Azadirachtin showed strong interaction with retinoid receptors. It suppressed ATRA-mediated removal of retinoid receptors, bound with DNA by inhibiting ATRA binding to its receptors. Overall, our data suggest that azadirachtin interacts with retinoic acid receptors and suppresses ATRA binding, inhibits falling off the receptors, and activates transcription factors like CREB, Sp1, NF-κB, etc. Thus, azadirachtin exerts anti-inflammatory and anti-metastatic responses by a novel pathway that would be beneficial for further anti-inflammatory and anti-cancer therapies. PMID:21127062

  2. Acute Stress Suppresses Synaptic Inhibition and Increases Anxiety via Endocannabinoid Release in the Basolateral Amygdala.

    Science.gov (United States)

    Di, Shi; Itoga, Christy A; Fisher, Marc O; Solomonow, Jonathan; Roltsch, Emily A; Gilpin, Nicholas W; Tasker, Jeffrey G

    2016-08-10

    Stress and glucocorticoids stimulate the rapid mobilization of endocannabinoids in the basolateral amygdala (BLA). Cannabinoid receptors in the BLA contribute to anxiogenesis and fear-memory formation. We tested for rapid glucocorticoid-induced endocannabinoid regulation of synaptic inhibition in the rat BLA. Glucocorticoid application to amygdala slices elicited a rapid, nonreversible suppression of spontaneous, but not evoked, GABAergic synaptic currents in BLA principal neurons; the effect was also seen with a membrane-impermeant glucocorticoid, but not with intracellular glucocorticoid application, implicating a membrane-associated glucocorticoid receptor. The glucocorticoid suppression of GABA currents was not blocked by antagonists of nuclear corticosteroid receptors, or by inhibitors of gene transcription or protein synthesis, but was blocked by inhibiting postsynaptic G-protein activity, suggesting a postsynaptic nongenomic steroid signaling mechanism that stimulates the release of a retrograde messenger. The rapid glucocorticoid-induced suppression of inhibition was prevented by blocking CB1 receptors and 2-arachidonoylglycerol (2-AG) synthesis, and it was mimicked and occluded by CB1 receptor agonists, indicating it was mediated by the retrograde release of the endocannabinoid 2-AG. The rapid glucocorticoid effect in BLA neurons in vitro was occluded by prior in vivo acute stress-induced, or prior in vitro glucocorticoid-induced, release of endocannabinoid. Acute stress also caused an increase in anxiety-like behavior that was attenuated by blocking CB1 receptor activation and inhibiting 2-AG synthesis in the BLA. Together, these findings suggest that acute stress causes a long-lasting suppression of synaptic inhibition in BLA neurons via a membrane glucocorticoid receptor-induced release of 2-AG at GABA synapses, which contributes to stress-induced anxiogenesis. We provide a cellular mechanism in the basolateral amygdala (BLA) for the rapid stress

  3. Nuclear receptor TLX inhibits TGF-β signaling in glioblastoma

    Energy Technology Data Exchange (ETDEWEB)

    Johansson, Erik; Zhai, Qiwei [Sahlgrenska Cancer Center at the Sahlgrenska Academy, University of Gothenburg, Box 425, SE 405 30 Gothenburg (Sweden); Zeng, Zhao-jun [Sahlgrenska Cancer Center at the Sahlgrenska Academy, University of Gothenburg, Box 425, SE 405 30 Gothenburg (Sweden); Molecular Biology Research Center, School of Life Sciences, Central South University, 110, Xiangya Road, Changsha, Hunan 410078 (China); Yoshida, Takeshi [Sahlgrenska Cancer Center at the Sahlgrenska Academy, University of Gothenburg, Box 425, SE 405 30 Gothenburg (Sweden); Funa, Keiko, E-mail: keiko.funa@gu.se [Sahlgrenska Cancer Center at the Sahlgrenska Academy, University of Gothenburg, Box 425, SE 405 30 Gothenburg (Sweden)

    2016-05-01

    TLX (also called NR2E1) is an orphan nuclear receptor that maintains stemness of neuronal stem cells. TLX is highly expressed in the most malignant form of glioma, glioblastoma multiforme (GBM), and is important for the proliferation and maintenance of the stem/progenitor cells of the tumor. Transforming Growth Factor-β (TGF-β) is a cytokine regulating many different cellular processes such as differentiation, migration, adhesion, cell death and proliferation. TGF-β has an important function in cancer where it can work as either a tumor suppressor or oncogene, depending on the cancer type and stage of tumor development. Since glioblastoma often have dysfunctional TGF-β signaling we wanted to find out if there is any interaction between TLX and TGF-β in glioblastoma cells. We demonstrate that knockdown of TLX enhances the canonical TGF-β signaling response in glioblastoma cell lines. TLX physically interacts with and stabilizes Smurf1, which can ubiquitinate and target TGF-β receptor II for degradation, whereas knockdown of TLX leads to stabilization of TGF-β receptor II, increased nuclear translocation of Smad2/3 and enhanced expression of TGF-β target genes. The interaction between TLX and TGF-β may play an important role in the regulation of proliferation and tumor-initiating properties of glioblastoma cells. - Highlights: • TLX knockdown enhances TGF-β dependent Smad signaling in glioblastoma cells • TLX knockdown increases the protein level of TGF-β receptor II. • TLX stabilizes and retains Smurf1 in the cytoplasm. • TLX enhances Smurf1-dependent ubiquitination and degradation of TGF-β receptor II.

  4. Nuclear receptor TLX inhibits TGF-β signaling in glioblastoma

    International Nuclear Information System (INIS)

    Johansson, Erik; Zhai, Qiwei; Zeng, Zhao-jun; Yoshida, Takeshi; Funa, Keiko

    2016-01-01

    TLX (also called NR2E1) is an orphan nuclear receptor that maintains stemness of neuronal stem cells. TLX is highly expressed in the most malignant form of glioma, glioblastoma multiforme (GBM), and is important for the proliferation and maintenance of the stem/progenitor cells of the tumor. Transforming Growth Factor-β (TGF-β) is a cytokine regulating many different cellular processes such as differentiation, migration, adhesion, cell death and proliferation. TGF-β has an important function in cancer where it can work as either a tumor suppressor or oncogene, depending on the cancer type and stage of tumor development. Since glioblastoma often have dysfunctional TGF-β signaling we wanted to find out if there is any interaction between TLX and TGF-β in glioblastoma cells. We demonstrate that knockdown of TLX enhances the canonical TGF-β signaling response in glioblastoma cell lines. TLX physically interacts with and stabilizes Smurf1, which can ubiquitinate and target TGF-β receptor II for degradation, whereas knockdown of TLX leads to stabilization of TGF-β receptor II, increased nuclear translocation of Smad2/3 and enhanced expression of TGF-β target genes. The interaction between TLX and TGF-β may play an important role in the regulation of proliferation and tumor-initiating properties of glioblastoma cells. - Highlights: • TLX knockdown enhances TGF-β dependent Smad signaling in glioblastoma cells • TLX knockdown increases the protein level of TGF-β receptor II. • TLX stabilizes and retains Smurf1 in the cytoplasm. • TLX enhances Smurf1-dependent ubiquitination and degradation of TGF-β receptor II.

  5. Resistance to diet-induced adiposity in cannabinoid receptor-1 deficient mice is not due to impaired adipocyte function

    Directory of Open Access Journals (Sweden)

    Oosterveer Maaike H

    2011-12-01

    Full Text Available Abstract Background Overactivity and/or dysregulation of the endocannabinoid system (ECS contribute to development of obesity. In vitro studies indicate a regulatory role for the cannabinoid receptor 1 (CB1 in adipocyte function and CB1-receptor deficient (CB1-/- mice are resistant to high fat diet-induced obesity. Whether this phenotype of CB1-/- mice is related to altered fat metabolism in adipose tissue is unknown. Methods We evaluated adipose tissue differentiation/proliferation markers and quantified lipogenic and lipolytic activities in fat tissues of CB1-/- and CB1+/+ mice fed a high-fat (HF or a high-fat/fish oil (HF/FO diet as compared to animals receiving a low-fat chow diet. Comparison between HF diet and HF/FO diet allowed to investigate the influence of dietary fat quality on adipose tissue biology in relation to CB1 functioning. Results The adiposity-resistant phenotype of the CB1-/- mice was characterized by reduced fat mass and adipocyte size in HF and HF/FO-fed CB1-/- mice in parallel to a significant increase in energy expenditure as compared to CB1+/+ mice. The expression levels of adipocyte differentiation and proliferation markers were however maintained in these animals. Consistent with unaltered lipogenic gene expression, the fatty acid synthesis rates in adipose tissues from CB1-/- and CB1+/+ mice were unchanged. Whole-body and adipose-specific lipoprotein lipase (LPL activities were also not altered in CB1-/- mice. Conclusions These findings indicate that protection against diet-induced adiposity in CB1-deficient mice is not related to changes in adipocyte function per se, but rather results from increased energy dissipation by oxidative and non-oxidative pathways.

  6. Nuclear receptor TLX inhibits TGF-β signaling in glioblastoma.

    Science.gov (United States)

    Johansson, Erik; Zhai, Qiwei; Zeng, Zhao-Jun; Yoshida, Takeshi; Funa, Keiko

    2016-05-01

    TLX (also called NR2E1) is an orphan nuclear receptor that maintains stemness of neuronal stem cells. TLX is highly expressed in the most malignant form of glioma, glioblastoma multiforme (GBM), and is important for the proliferation and maintenance of the stem/progenitor cells of the tumor. Transforming Growth Factor-β (TGF-β) is a cytokine regulating many different cellular processes such as differentiation, migration, adhesion, cell death and proliferation. TGF-β has an important function in cancer where it can work as either a tumor suppressor or oncogene, depending on the cancer type and stage of tumor development. Since glioblastoma often have dysfunctional TGF-β signaling we wanted to find out if there is any interaction between TLX and TGF-β in glioblastoma cells. We demonstrate that knockdown of TLX enhances the canonical TGF-β signaling response in glioblastoma cell lines. TLX physically interacts with and stabilizes Smurf1, which can ubiquitinate and target TGF-β receptor II for degradation, whereas knockdown of TLX leads to stabilization of TGF-β receptor II, increased nuclear translocation of Smad2/3 and enhanced expression of TGF-β target genes. The interaction between TLX and TGF-β may play an important role in the regulation of proliferation and tumor-initiating properties of glioblastoma cells. Copyright © 2016. Published by Elsevier Inc.

  7. Probenecid inhibits α-adrenergic receptor-mediated vasoconstriction in the human leg vasculature

    DEFF Research Database (Denmark)

    Nyberg, Michael Permin; Piil, Peter Bergmann; Kiehn, Oliver Thistrup

    2018-01-01

    to α1- and α2-adrenergic receptor stimulation in the human forearm and leg vasculature of young healthy male subjects (23±3 years). By use of immunolabeling and confocal microscopy, Panx1 channels were found to be expressed in vascular smooth muscle cells of arterioles in human leg skeletal muscle....... Probenecid treatment increased (Padrenergic receptor stimulation) by ≈15%, whereas the response to the α1-agonist phenylephrine was unchanged. Inhibition...

  8. Dopamine inhibits somatolactin gene expression in tilapia pituitary cells through the dopamine D2 receptors.

    Science.gov (United States)

    Jiang, Quan; Lian, Anji; He, Qi

    2016-07-01

    Dopamine (DA) is an important neurotransmitter in the central nervous system of vertebrates and possesses key hypophysiotropic functions. Early studies have shown that DA has a potent inhibitory effect on somatolactin (SL) release in fish. However, the mechanisms responsible for DA inhibition of SL gene expression are largely unknown. To this end, tilapia DA type-1 (D1) and type-2 (D2) receptor transcripts were examined in the neurointermediate lobe (NIL) of the tilapia pituitary by real-time PCR. In tilapia, DA not only was effective in inhibiting SL mRNA levels in vivo and in vitro, but also could abolish pituitary adenylate cyclase-activating polypeptide (PACAP)- and salmon gonadotropin-releasing hormone (sGnRH)-stimulated SL gene expression at the pituitary level. In parallel studies, the specific D2 receptor agonists quinpirole and bromocriptine could mimic the DA-inhibited SL gene expression. Furthermore, the D2 receptor antagonists domperidone and (-)-sulpiride could abolish the SL response to DA or the D2 agonist quinpirole, whereas D1 receptor antagonists SCH23390 and SKF83566 were not effective in this respect. In primary cultures of tilapia NIL cells, D2 agonist quinpirole-inhibited cAMP production could be blocked by co-treatment with the D2 antagonist domperidone and the ability of forskolin to increase cAMP production was also inhibited by quinpirole. Using a pharmacological approach, the AC/cAMP pathway was shown to be involved in quinpirole-inhibited SL mRNA expression. These results provide evidence that DA can directly inhibit SL gene expression at the tilapia pituitary level via D2 receptor through the AC/cAMP-dependent mechanism. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Modulating Estrogen Receptor-related Receptor-α Activity Inhibits Cell Proliferation*

    OpenAIRE

    Bianco, Stéphanie; Lanvin, Olivia; Tribollet, Violaine; Macari, Claire; North, Sophie; Vanacker, Jean-Marc

    2009-01-01

    High expression of the estrogen receptor-related receptor (ERR)-α in human tumors is correlated to a poor prognosis, suggesting an involvement of the receptor in cell proliferation. In this study, we show that a synthetic compound (XCT790) that modulates the activity of ERRα reduces the proliferation of various cell lines and blocks the G1/S transition of the cell cycle in an ERR...

  10. Inhibition of TRAIL-induced apoptosis and forced internalization of TRAIL receptor 1 by adenovirus proteins.

    Science.gov (United States)

    Tollefson, A E; Toth, K; Doronin, K; Kuppuswamy, M; Doronina, O A; Lichtenstein, D L; Hermiston, T W; Smith, C A; Wold, W S

    2001-10-01

    Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) induces apoptosis through two receptors, TRAIL-R1 (also known as death receptor 4) and TRAIL-R2 (also known as death receptor 5), that are members of the TNF receptor superfamily of death domain-containing receptors. We show that human adenovirus type 5 encodes three proteins, named RID (previously named E3-10.4K/14.5K), E3-14.7K, and E1B-19K, that independently inhibit TRAIL-induced apoptosis of infected human cells. This conclusion was derived from studies using wild-type adenovirus, adenovirus replication-competent mutants that lack one or more of the RID, E3-14.7K, and E1B-19K genes, and adenovirus E1-minus replication-defective vectors that express all E3 genes, RID plus E3-14.7K only, RID only, or E3-14.7K only. RID inhibits TRAIL-induced apoptosis when cells are sensitized to TRAIL either by adenovirus infection or treatment with cycloheximide. RID induces the internalization of TRAIL-R1 from the cell surface, as shown by flow cytometry and indirect immunofluorescence for TRAIL-R1. TRAIL-R1 was internalized in distinct vesicles which are very likely to be endosomes and lysosomes. TRAIL-R1 is degraded, as indicated by the disappearance of the TRAIL-R1 immunofluorescence signal. Degradation was inhibited by bafilomycin A1, a drug that prevents acidification of vesicles and the sorting of receptors from late endosomes to lysosomes, implying that degradation occurs in lysosomes. RID was also shown previously to internalize and degrade another death domain receptor, Fas, and to prevent apoptosis through Fas and the TNF receptor. RID was shown previously to force the internalization and degradation of the epidermal growth factor receptor. E1B-19K was shown previously to block apoptosis through Fas, and both E1B-19K and E3-14.7K were found to prevent apoptosis through the TNF receptor. These findings suggest that the receptors for TRAIL, Fas ligand, and TNF play a role in limiting virus

  11. Activation of dopamine D3 receptors inhibits reward-related learning induced by cocaine.

    Science.gov (United States)

    Kong, H; Kuang, W; Li, S; Xu, M

    2011-03-10

    Memories of learned associations between the rewarding properties of drugs and environmental cues contribute to craving and relapse in humans. The mesocorticolimbic dopamine (DA) system is involved in reward-related learning induced by drugs of abuse. DA D3 receptors are preferentially expressed in mesocorticolimbic DA projection areas. Genetic and pharmacological studies have shown that DA D3 receptors suppress locomotor-stimulant effects of cocaine and reinstatement of cocaine-seeking behaviors. Activation of the extracellular signal-regulated kinase (ERK) induced by acute cocaine administration is also inhibited by D3 receptors. How D3 receptors modulate cocaine-induced reward-related learning and associated changes in cell signaling in reward circuits in the brain, however, have not been fully investigated. In the present study, we show that D3 receptor mutant mice exhibit potentiated acquisition of conditioned place preference (CPP) at low doses of cocaine compared to wild-type mice. Activation of ERK and CaMKIIα, but not the c-Jun N-terminal kinase and p38, in the nucleus accumbens, amygdala and prefrontal cortex is also potentiated in D3 receptor mutant mice compared to that in wild-type mice following CPP expression. These results support a model in which D3 receptors modulate reward-related learning induced by low doses of cocaine by inhibiting activation of ERK and CaMKIIα in reward circuits in the brain. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

  12. Inhibition of radiation-induced polyuria by histamine receptor antagonists

    Energy Technology Data Exchange (ETDEWEB)

    Donlon, M.A.; Melia, J.A.; Helgeson, E.A.; Wolfe, W.W.

    1986-03-01

    In previous studies the authors have demonstrated that gamma radiation results in polyuria, which is preceded by polydypsia. This suggests that the increased thirst elicited by radiation causes increased urinary volume (UV). Histamine, which is released following radiation exposure, also elicits drinking by nonirradiated rats when administered exogenously. In this study the authors have investigated both the role of water deprivation and the effect of histamine receptor antagonists (HRA) on radiation-induced polyuria. Sprague-Dawley rats were housed individually in metabolic cages. Water was allowed ad libitum except in deprivation experiments where water was removed for 24 hr immediately following radiation. Cimetidine (CIM), an H2 HRA, and dexbromopheniramine (DXB), an H1 HRA, were administered i.p. (16 and 1 mg/kg, respectively) 30 min prior to irradiation (950 rads from a cobalt source). UV was determined at 24-hr intervals for 3 days preceding irradiation and 24 hr postirradiation. UV in DXB treated rats was significantly reduced 24 hr postirradiation (CON = 427 +/- 54%; DXB = 247 +/- 39% of preirradiated CON) compared to postirradiation control values. CIM did not affect postirradiation UV. These data suggest that radiation-induced polyuria is caused by polydypsia which is, in part, mediated by histamine induced by an H1 receptor.

  13. Inhibition of radiation-induced polyuria by histamine receptor antagonists

    International Nuclear Information System (INIS)

    Donlon, M.A.; Melia, J.A.; Helgeson, E.A.; Wolfe, W.W.

    1986-01-01

    In previous studies the authors have demonstrated that gamma radiation results in polyuria, which is preceded by polydypsia. This suggests that the increased thirst elicited by radiation causes increased urinary volume (UV). Histamine, which is released following radiation exposure, also elicits drinking by nonirradiated rats when administered exogenously. In this study the authors have investigated both the role of water deprivation and the effect of histamine receptor antagonists (HRA) on radiation-induced polyuria. Sprague-Dawley rats were housed individually in metabolic cages. Water was allowed ad libitum except in deprivation experiments where water was removed for 24 hr immediately following radiation. Cimetidine (CIM), an H2 HRA, and dexbromopheniramine (DXB), an H1 HRA, were administered i.p. (16 and 1 mg/kg, respectively) 30 min prior to irradiation (950 rads from a cobalt source). UV was determined at 24-hr intervals for 3 days preceding irradiation and 24 hr postirradiation. UV in DXB treated rats was significantly reduced 24 hr postirradiation (CON = 427 +/- 54%; DXB = 247 +/- 39% of preirradiated CON) compared to postirradiation control values. CIM did not affect postirradiation UV. These data suggest that radiation-induced polyuria is caused by polydypsia which is, in part, mediated by histamine induced by an H1 receptor

  14. Pharmacological or genetic orexin 1 receptor inhibition attenuates MK-801 induced glutamate release in mouse cortex

    Directory of Open Access Journals (Sweden)

    Leah eAluisio

    2014-05-01

    Full Text Available The orexin/hypocretin neuropeptides are produced by a cluster of neurons within the lateral posterior hypothalamus and participate in neuronal regulation by activating their receptors (OX1 and OX2 receptors. The orexin system projects widely through the brain and functions as an interface between multiple regulatory systems including wakefulness, energy balance, stress, reward and emotion. Recent studies have demonstrated that orexins and glutamate interact at the synaptic level and that orexins facilitate glutamate actions. We tested the hypothesis that orexins modulate glutamate signaling via OX1 receptors by monitoring levels of glutamate in frontal cortex of freely moving mice using enzyme coated biosensors under inhibited OX1 receptor conditions. MK-801, an NMDA receptor antagonist, was administered subcutaneously (0.178 mg/kg to indirectly disinhibit pyramidal neurons and therefore increase cortical glutamate release. In wild-type mice, pretreatment with the OX1 receptor antagonist GSK-1059865 (10 mg/kg S.C. which had no effect by itself, significantly attenuated the cortical glutamate release elicited by MK-801. OX1 receptor knockout mice had a blunted glutamate release response to MK-801 and exhibited about half of the glutamate release observed in wild-type mice in agreement with the data obtained with transient blockade of OX1 receptors. These results indicate that pharmacological (transient or genetic (permanent inhibition of the OX1 receptor similarly interfere with glutamatergic function in the cortex. Selectively targeting the OX1 receptor with an antagonist may normalize hyperglutamatergic states and thus may represent a novel therapeutic strategy for the treatment of various psychiatric disorders associated with hyperactive states.

  15. Antidepressants inhibit P2X4 receptor function: a possible involvement in neuropathic pain relief

    Directory of Open Access Journals (Sweden)

    Tozaki-Saitoh Hidetoshi

    2009-04-01

    Full Text Available Abstract Background Neuropathic pain is characterized by pain hypersensitivity to innocuous stimuli (tactile allodynia that is nearly always resistant to known treatments such as non-steroidal anti-inflammatory drugs or even opioids. It has been reported that some antidepressants are effective for treating neuropathic pain. However, the underlying molecular mechanisms are not well understood. We have recently demonstrated that blocking P2X4 receptors in the spinal cord reverses tactile allodynia after peripheral nerve injury in rats, implying that P2X4 receptors are a key molecule in neuropathic pain. We investigated a possible role of antidepressants as inhibitors of P2X4 receptors and analysed their analgesic mechanism using an animal model of neuropathic pain. Results Antidepressants strongly inhibited ATP-mediated Ca2+ responses in P2X4 receptor-expressing 1321N1 cells, which are known to have no endogenous ATP receptors. Paroxetine exhibited the most powerful inhibition of calcium influx via rat and human P2X4 receptors, with IC50 values of 2.45 μM and 1.87 μM, respectively. Intrathecal administration of paroxetine produced a striking antiallodynic effect in an animal model of neuropathic pain. Co-administration of WAY100635, ketanserin or ondansetron with paroxetine induced no significant change in the antiallodynic effect of paroxetine. Furthermore, the antiallodynic effect of paroxetine was observed even in rats that had received intrathecal pretreatment with 5,7-dihydroxytryptamine, which dramatically depletes spinal 5-hydroxytryptamine. Conclusion These results suggest that paroxetine acts as a potent analgesic in the spinal cord via a mechanism independent of its inhibitory effect on serotonin transporters. Powerful inhibition on P2X4 receptors may underlie the analgesic effect of paroxetine, and it is possible that some antidepressants clinically used in patients with neuropathic pain show antiallodynic effects, at least in part

  16. B cell receptor inhibition as a target for CLL therapy.

    Science.gov (United States)

    Jeyakumar, Deepa; O'Brien, Susan

    2016-03-01

    Inhibitors of the B cell receptor (BCR) represent an attractive therapeutic option for patients with chronic lymphocytic leukemia. Recently approved inhibitors of Bruton's tyrosine kinase (ibrutinib) and phosphatidylinositol 3-kinase (idelalisib), are promising agents because they are generally well tolerated and highly effective. These agents may be particularly important in the treatment of older patients who are less able to tolerate the myelosuppression (and infections) associated with chemoimmunotherapy. As a class of medications, BCR inhibitors have some unique side effects including redistribution lymphocytosis. Ibrutinib has specific toxicities including increased risk for bleeding and atrial fibrillation. Idelalisib also has some unique toxicities consisting of transaminitis, diarrhea and pneumonitis. Ongoing clinical trials are evaluating these agents in combination with antibodies, chemotherapy and other small molecules. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Immunohistochemical analysis of cannabinoid receptor 1 expression in steatotic rat livers.

    Science.gov (United States)

    Zduniak, Krzysztof; Ziółkowski, Piotr; Regnell, Pontus; Tollet-Egnell, Petra; Åkesson, Lina; Cooper, Martin E

    2016-04-01

    The primary aim of the present study was to determine the expression levels of cannabinoid receptor type 1 (CB1) in steatotic rat livers. The secondary aim was to clarify whether steatosis and inflammation are more marked in areas with increased CB1 overexpression. For ethical and economic reasons, the present study investigated tissue from archived liver blocks, which were obtained from 38 rats that had been euthanized during the course of previous research at the Karolinska Institute of the Karolinska University Hospital (Stockholm, Sweden) and Lund University (Malmö, Sweden). Liver tissue fixed in formalin and embedded in paraffin was used that had been sourced from 36 male Sprague Dawley rats (age, 7 weeks) and 2 rats (age, 180 days) lacking normal leptin receptors. The rat liver tissue was stained with antibodies against CB1 and counterstained with hematoxylin. The expression of CB1 and the number of cells overexpressing CB1 were determined. Steatosis was scored according to the Dixon scoring system. CB1 overexpression and steatosis were detected in hepatocytes from all 38 livers sampled. The expression of CB1 was more marked in hepatocytes localized next to portal triads. Near the central veins, the expression was significantly weaker. Steatosis was more marked in areas of increased CB1 overexpression. Lymphocyte infiltration was more commonly observed in areas of increased CB1 overexpression. Therefore, the present results indicate that CB1 receptors are overexpressed in areas with steatosis, and indicate that CB1 in hepatocytes contributes to the formation of steatosis in rats, even prior to its progression to steatohepatitis. These results are consistent with publications reporting that CB1 in hepatocytes increases lipogenesis and contributes to inflammation.

  18. Epidermal Growth Factor Receptor Transactivation by the Cannabinoid Receptor (CB1) and Transient Receptor Potential Vanilloid 1 (TRPV1) Induces Differential Responses in Corneal Epithelial Cells

    Science.gov (United States)

    2010-01-01

    inflammatory cytokine release in corneal epithelium through MAPK signaling. J. Cell Physiol. 213, 730e739. Zhang, J., Li , H., Wang, J., Dong, Z., Mian ...Kang, S.S., Li , T., Xu, D., Reinach, P.S., Lu, L., 2000. Inhibitory effect of PGE2 on EGF- induced MAP kinase activity and rabbit corneal epithelial

  19. Inhibition of neointima formation by local delivery of estrogen receptor alpha and beta specific agonists

    NARCIS (Netherlands)

    Krom, Y.D.; Pires, N.M.M.; Jukema, J.W.; Vries, M.R. de; Frants, R.R.; Havekes, L.M.; Dijk, K.W. van; Quax, P.H.A.

    2007-01-01

    Objective: Neointima formation is the underlying mechanism of (in-stent) restenosis. 17β-Estradiol (E2) is known to inhibit injury-induced neointima formation and post-angioplasty restenosis. Estrogen receptor alpha (ERα) has been demonstrated to mediate E2 anti-restenotic properties. However, the

  20. Mechanism of inhibition of growth hormone receptor signaling by suppressor of cytokine signaling proteins

    DEFF Research Database (Denmark)

    Hansen, J A; Lindberg, K; Hilton, D J

    1999-01-01

    In this study we have investigated the role of suppressor of cytokine signaling (SOCS) proteins in GH receptor-mediated signaling. GH-induced transcription was inhibited by SOCS-1 and SOCS-3, while SOCS-2 and cytokine inducible SH2-containing protein (CIS) had no effect By using chimeric SOCS pro...

  1. Molecular requirements for inhibition of the chemokine receptor CCR8--probe-dependent allosteric interactions

    DEFF Research Database (Denmark)

    Rummel, Pia Cwarzko; Arfelt, K N; Baumann, L

    2012-01-01

    Here we present a novel series of CCR8 antagonists based on a naphthalene-sulfonamide structure. This structure differs from the predominant pharmacophore for most small-molecule CC-chemokine receptor antagonists, which in fact activate CCR8, suggesting that CCR8 inhibition requires alternative...

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

    Science.gov (United States)

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

    1991-11-26

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

  3. Activation of the sympathetic nervous system mediates hypophagic and anxiety-like effects of CB₁ receptor blockade.

    Science.gov (United States)

    Bellocchio, Luigi; Soria-Gómez, Edgar; Quarta, Carmelo; Metna-Laurent, Mathilde; Cardinal, Pierre; Binder, Elke; Cannich, Astrid; Delamarre, Anna; Häring, Martin; Martín-Fontecha, Mar; Vega, David; Leste-Lasserre, Thierry; Bartsch, Dusan; Monory, Krisztina; Lutz, Beat; Chaouloff, Francis; Pagotto, Uberto; Guzman, Manuel; Cota, Daniela; Marsicano, Giovanni

    2013-03-19

    Complex interactions between periphery and the brain regulate food intake in mammals. Cannabinoid type-1 (CB1) receptor antagonists are potent hypophagic agents, but the sites where this acute action is exerted and the underlying mechanisms are not fully elucidated. To dissect the mechanisms underlying the hypophagic effect of CB1 receptor blockade, we combined the acute injection of the CB1 receptor antagonist rimonabant with the use of conditional CB1-knockout mice, as well as with pharmacological modulation of different central and peripheral circuits. Fasting/refeeding experiments revealed that CB1 receptor signaling in many specific brain neurons is dispensable for the acute hypophagic effects of rimonabant. CB1 receptor antagonist-induced hypophagia was fully abolished by peripheral blockade of β-adrenergic transmission, suggesting that this effect is mediated by increased activity of the sympathetic nervous system. Consistently, we found that rimonabant increases gastrointestinal metabolism via increased peripheral β-adrenergic receptor signaling in peripheral organs, including the gastrointestinal tract. Blockade of both visceral afferents and glutamatergic transmission in the nucleus tractus solitarii abolished rimonabant-induced hypophagia. Importantly, these mechanisms were specifically triggered by lipid-deprivation, revealing a nutrient-specific component acutely regulated by CB1 receptor blockade. Finally, peripheral blockade of sympathetic neurotransmission also blunted central effects of CB1 receptor blockade, such as fear responses and anxiety-like behaviors. These data demonstrate that, independently of their site of origin, important effects of CB1 receptor blockade are expressed via activation of peripheral sympathetic activity. Thus, CB1 receptors modulate bidirectional circuits between the periphery and the brain to regulate feeding and other behaviors.

  4. P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions.

    Science.gov (United States)

    Giroud, Charline; Marin, Mariana; Hammonds, Jason; Spearman, Paul; Melikyan, Gregory B

    2015-09-01

    HIV-1 Env glycoprotein-mediated fusion is initiated upon sequential binding of Env to CD4 and the coreceptor CXCR4 or CCR5. Whereas these interactions are thought to be necessary and sufficient to promote HIV-1 fusion, other host factors can modulate this process. Previous studies reported potent inhibition of HIV-1 fusion by selective P2X1 receptor antagonists, including NF279, and suggested that these receptors play a role in HIV-1 entry. Here we investigated the mechanism of antiviral activity of NF279 and found that this compound does not inhibit HIV-1 fusion by preventing the activation of P2X1 channels but effectively blocks the binding of the virus to CXCR4 or CCR5. The notion of an off-target effect of NF279 on HIV-1 fusion is supported by the lack of detectable expression of P2X1 receptors in cells used in fusion experiments and by the fact that the addition of ATP or the enzymatic depletion of ATP in culture medium does not modulate viral fusion. Importantly, NF279 fails to inhibit HIV-1 fusion with cell lines and primary macrophages when added at an intermediate stage downstream of Env-CD4-coreceptor engagement. Conversely, in the presence of NF279, HIV-1 fusion is arrested downstream of CD4 binding but prior to coreceptor engagement. NF279 also antagonizes the signaling function of CCR5, CXCR4, and another chemokine receptor, as evidenced by the suppression of calcium responses elicited by specific ligands and by recombinant gp120. Collectively, our results demonstrate that NF279 is a dual HIV-1 coreceptor inhibitor that interferes with the functional engagement of CCR5 and CXCR4 by Env. Inhibition of P2X receptor activity suppresses HIV-1 fusion and replication, suggesting that P2X signaling is involved in HIV-1 entry. However, mechanistic experiments conducted in this study imply that P2X1 receptor is not expressed in target cells or involved in viral fusion. Instead, we found that inhibition of HIV-1 fusion by a specific P2X1 receptor antagonist, NF

  5. Resveratrol inhibits PDGF receptor mitogenic signaling in mesangial cells: role of PTP1B

    Science.gov (United States)

    Venkatesan, Balachandar; Ghosh-Choudhury, Nandini; Das, Falguni; Mahimainathan, Lenin; Kamat, Amrita; Kasinath, Balakuntalam S.; Abboud, Hanna E.; Choudhury, Goutam Ghosh

    2008-01-01

    Mesangioproliferative glomerulonephritis is associated with overactive PDGF receptor signal transduction. We show that the phytoalexin resveratrol dose dependently inhibits PDGF-induced DNA synthesis in mesangial cells with an IC50 of 10 μM without inducing apoptosis. Remarkably, the increased SIRT1 deacetylase activity induced by resveratrol was not necessary for this inhibitory effect. Resveratrol significantly blocked PDGF-stimulated c-Src and Akt kinase activation, resulting in reduced cyclin D1 expression and attenuated pRb phosphorylation and cyclin-dependent kinase-2 (CDK2) activity. Furthermore, resveratrol inhibited PDGFR phosphorylation at the PI 3 kinase and Grb-2 binding sites tyrosine-751 and tyrosine-716, respectively. This deficiency in PDGFR phosphorylation resulted in significant inhibition of PI 3 kinase and Erk1/2 MAPK activity. Interestingly, resveratrol increased the activity of protein tyrosine phosphatase PTP1B, which dephosphorylates PDGF-stimulated phosphorylation at tyrosine-751 and tyrosine-716 on PDGFR with concomitant reduction in Akt and Erk1/2 kinase activity. PTP1B significantly inhibited PDGF-induced DNA synthesis without inducing apoptosis. These results for the first time provide evidence that the stilbene resveratrol targets PTP1B to inhibit PDGFR mitogenic signaling.—Venkatesan, B., Ghosh-Choudhury, N., Das, F., Mahimainathan, L., Kamat, A., Kasinath, B. S., Abboud, H. E., Choudhury, G. G. Resveratrol inhibits PDGF receptor mitogenic signaling in mesangial cells: role of PTP1B. PMID:18567737

  6. Modulating Estrogen Receptor-related Receptor-α Activity Inhibits Cell Proliferation*

    Science.gov (United States)

    Bianco, Stéphanie; Lanvin, Olivia; Tribollet, Violaine; Macari, Claire; North, Sophie; Vanacker, Jean-Marc

    2009-01-01

    High expression of the estrogen receptor-related receptor (ERR)-α in human tumors is correlated to a poor prognosis, suggesting an involvement of the receptor in cell proliferation. In this study, we show that a synthetic compound (XCT790) that modulates the activity of ERRα reduces the proliferation of various cell lines and blocks the G1/S transition of the cell cycle in an ERRα-dependent manner. XCT790 induces, in a p53-independent manner, the expression of the cell cycle inhibitor p21waf/cip1 at the protein, mRNA, and promoter level, leading to an accumulation of hypophosphorylated Rb. Finally, XCT790 reduces cell tumorigenicity in Nude mice. PMID:19546226

  7. Acutely increasing δGABAA receptor activity impairs memory and inhibits synaptic plasticity in the hippocampus

    Science.gov (United States)

    Whissell, Paul D.; Eng, Dave; Lecker, Irene; Martin, Loren J.; Wang, Dian-Shi; Orser, Beverley A.

    2013-01-01

    Extrasynaptic γ-aminobutyric acid type A (GABAA) receptors that contain the δ subunit (δGABAA receptors) are expressed in several brain regions including the dentate gyrus (DG) and CA1 subfields of the hippocampus. Drugs that increase δGABAA receptor activity have been proposed as treatments for a variety of disorders including insomnia, epilepsy and chronic pain. Also, long-term pretreatment with the δGABAA receptor–preferring agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) enhances discrimination memory and increases neurogenesis in the DG. Despite the potential therapeutic benefits of such treatments, the effects of acutely increasing δGABAA receptor activity on memory behaviors remain unknown. Here, we studied the effects of THIP (4 mg/kg, i.p.) on memory performance in wild-type (WT) and δGABAA receptor null mutant (Gabrd−/−) mice. Additionally, the effects of THIP on long-term potentiation (LTP), a molecular correlate of memory, were studied within the DG and CA1 subfields of the hippocampus using electrophysiological recordings of field potentials in hippocampal slices. The results showed that THIP impaired performance in the Morris water maze, contextual fear conditioning and object recognition tasks in WT mice but not Gabrd−/− mice. Furthermore, THIP inhibited LTP in hippocampal slices from WT but not Gabrd−/− mice, an effect that was blocked by GABAA receptor antagonist bicuculline. Thus, acutely increasing δGABAA receptor activity impairs memory behaviors and inhibits synaptic plasticity. These results have important implications for the development of therapies aimed at increasing δGABAA receptor activity. PMID:24062648

  8. Acutely increasing δGABAA receptor activity impairs memory and inhibits synaptic plasticity in the hippocampus

    Directory of Open Access Journals (Sweden)

    Paul David Whissell

    2013-09-01

    Full Text Available Extrasynaptic γ-aminobutyric acid type A (GABAA receptors that contain the δ subunit (δGABAA receptors are expressed in several brain regions including the dentate gyrus (DG and CA1 subfields of the hippocampus. Drugs that increase δGABAA receptor activity have been proposed as treatments for a variety of disorders including insomnia, epilepsy and chronic pain. Also, long-term pretreatment with the δGABAA receptor–preferring agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP enhances discrimination memory and increases neurogenesis in the DG. Despite the potential therapeutic benefits of such treatments, the effects of acutely increasing δGABAA receptor activity on memory behaviors remain unknown. Here, we studied the effects of THIP (4 mg/kg, i.p. on memory performance in wild-type (WT and δGABAA receptor null mutant (Gabrd–/– mice. Additionally, the effects of THIP on long-term potentiation (LTP, a molecular correlate of memory, were studied within the DG and CA1 subfields of the hippocampus using electrophysiological recordings of field potentials in hippocampal slices. The results showed that THIP impaired performance in the Morris water maze, contextual fear conditioning and object recognition tasks in WT mice but not Gabrd–/– mice. Furthermore, THIP inhibited LTP in hippocampal slices from WT but not Gabrd–/– mice, an effect that was blocked by GABAA receptor antagonist bicuculline. Thus, acutely increasing δGABAA receptor activity impairs memory behaviors and inhibits synaptic plasticity. These results have important implications for the development of therapies aimed at increasing δGABAA receptor activity.

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

    Directory of Open Access Journals (Sweden)

    Maelle Jospin

    2009-12-01

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

  10. Hispolon inhibits the growth of estrogen receptor positive human breast cancer cells through modulation of estrogen receptor alpha

    International Nuclear Information System (INIS)

    Jang, Eun Hyang; Jang, Soon Young; Cho, In-Hye; Hong, Darong; Jung, Bom; Park, Min-Ju; Kim, Jong-Ho

    2015-01-01

    Human estrogen receptor α (ERα) is a nuclear transcription factor that is a major therapeutic target in breast cancer. The transcriptional activity of ERα is regulated by certain estrogen-receptor modulators. Hispolon, isolated from Phellinus linteus, a traditional medicinal mushroom called Sanghwang in Korea, has been used to treat various pathologies, such as inflammation, gastroenteric disorders, lymphatic diseases, and cancers. In this latter context, Hispolon has been reported to exhibit therapeutic efficacy against various cancer cells, including melanoma, leukemia, hepatocarcinoma, bladder cancer, and gastric cancer cells. However, ERα regulation by Hispolon has not been reported. In this study, we investigated the effects of Hispolon on the growth of breast cancer cells. We found that Hispolon decreased expression of ERα at both mRNA and the protein levels in MCF7 and T47D human breast cancer cells. Luciferase reporter assays showed that Hispolon decreased the transcriptional activity of ERα. Hispolon treatment also inhibited expression of the ERα target gene pS2. We propose that Hispolon, an anticancer drug extracted from natural sources, inhibits cell growth through modulation of ERα in estrogen-positive breast cancer cells and is a candidate for use in human breast cancer chemotherapy. - Highlights: • Hispolon decreased ERα expression at both mRNA and protein levels. • Hispolon decreased ERα transcriptional activity. • Hispolon treatment inhibited expression of ERα target gene pS2. • Shikonin is a candidate chemotherapeutic target in the treatment of human breast cancer

  11. Hispolon inhibits the growth of estrogen receptor positive human breast cancer cells through modulation of estrogen receptor alpha

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Eun Hyang; Jang, Soon Young; Cho, In-Hye [Department of Pharmacy, Graduate School, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemun-gu, Seoul 130-701 (Korea, Republic of); Hong, Darong [Department of Life and Nanopharmaceutical Science, Graduate School, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemun-gu, Seoul 130-701 (Korea, Republic of); Jung, Bom; Park, Min-Ju [Department of Pharmacy, Graduate School, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemun-gu, Seoul 130-701 (Korea, Republic of); Kim, Jong-Ho, E-mail: jonghokim@khu.ac.kr [Department of Pharmacy, Graduate School, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemun-gu, Seoul 130-701 (Korea, Republic of)

    2015-08-07

    Human estrogen receptor α (ERα) is a nuclear transcription factor that is a major therapeutic target in breast cancer. The transcriptional activity of ERα is regulated by certain estrogen-receptor modulators. Hispolon, isolated from Phellinus linteus, a traditional medicinal mushroom called Sanghwang in Korea, has been used to treat various pathologies, such as inflammation, gastroenteric disorders, lymphatic diseases, and cancers. In this latter context, Hispolon has been reported to exhibit therapeutic efficacy against various cancer cells, including melanoma, leukemia, hepatocarcinoma, bladder cancer, and gastric cancer cells. However, ERα regulation by Hispolon has not been reported. In this study, we investigated the effects of Hispolon on the growth of breast cancer cells. We found that Hispolon decreased expression of ERα at both mRNA and the protein levels in MCF7 and T47D human breast cancer cells. Luciferase reporter assays showed that Hispolon decreased the transcriptional activity of ERα. Hispolon treatment also inhibited expression of the ERα target gene pS2. We propose that Hispolon, an anticancer drug extracted from natural sources, inhibits cell growth through modulation of ERα in estrogen-positive breast cancer cells and is a candidate for use in human breast cancer chemotherapy. - Highlights: • Hispolon decreased ERα expression at both mRNA and protein levels. • Hispolon decreased ERα transcriptional activity. • Hispolon treatment inhibited expression of ERα target gene pS2. • Shikonin is a candidate chemotherapeutic target in the treatment of human breast cancer.

  12. Enzalutamide inhibits androgen receptor-positive bladder cancer cell growth.

    Science.gov (United States)

    Kawahara, Takashi; Ide, Hiroki; Kashiwagi, Eiji; El-Shishtawy, Kareem A; Li, Yi; Reis, Leonardo O; Zheng, Yichun; Miyamoto, Hiroshi

    2016-10-01

    Emerging preclinical evidence suggests that androgen-mediated androgen receptor (AR) signals promote bladder cancer progression. However, little is known about the efficacy of an AR signaling inhibitor, enzalutamide, in the growth of bladder cancer cells. In this study, we compared the effects of enzalutamide and 2 other classic antiandrogens, flutamide and bicalutamide, on androgen-induced bladder cancer cell proliferation, migration, and invasion as well as tumor growth in vivo. Thiazolyl blue cell viability assay, flow cytometry, scratch wound-healing assay, transwell invasion assay, real-time polymerase chain reaction, and reporter gene assay were performed in AR-positive (e.g., UMUC3, TCCSUP, and 647V-AR) and AR-negative (e.g., UMUC3-AR-short hairpin RNA [shRNA], TCCSUP-AR-shRNA, 647V) bladder cancer lines treated with dihydrotestosterone and each AR antagonist. We also used a mouse xenograft model for bladder cancer. Dihydrotestosterone increased bladder cancer cell proliferation, migration, and invasion indicating that endogenous or exogenous AR was functional. Enzalutamide, hydroxyflutamide, and bicalutamide showed similar inhibitory effects, without significant agonist activity, on androgen-mediated cell viability/apoptosis, cell migration, and cell invasion in AR-positive lines. No significant effects of dihydrotestosterone as well as AR antagonists on the growth of AR-negative cells were seen. Correspondingly, in UMUC3 cells, these AR antagonists down-regulated androgen-induced expression of AR, matrix metalloproteinase-2, and interleukin-6. Androgen-enhanced AR-mediated transcriptional activity was also blocked by each AR antagonist exhibiting insignificant agonist activity. In UMUC3 xenograft-bearing mice, oral gavage treatment with each antiandrogen retarded tumor growth, and only enzalutamide demonstrated a statistically significant suppression compared with mock treatment. Our current data support recent observations indicating the involvement of

  13. Poxvirus-encoded TNF decoy receptors inhibit the biological activity of transmembrane TNF.

    Science.gov (United States)

    Pontejo, Sergio M; Alejo, Ali; Alcami, Antonio

    2015-10-01

    Poxviruses encode up to four different soluble TNF receptors, named cytokine response modifier B (CrmB), CrmC, CrmD and CrmE. These proteins mimic the extracellular domain of the cellular TNF receptors to bind and inhibit the activity of TNF and, in some cases, other TNF superfamily ligands. Most of these ligands are released after the enzymic cleavage of a membrane precursor. However, transmembrane TNF (tmTNF) is not only a precursor of soluble TNF but also exerts specific pro-inflammatory and immunological activities. Here, we report that viral TNF receptors bound and inhibited tmTNF and describe some interesting differences in their activity against the soluble cytokine. Thus, CrmE, which does not inhibit mouse soluble TNF, could block murine tmTNF-induced cytotoxicity. We propose that this anti-tmTNF effect should be taken into consideration when assessing the role of viral TNF decoy receptors in the pathogenesis of poxvirus.

  14. Insulin signaling inhibits the 5-HT2C receptor in choroid plexus via MAP kinase

    Directory of Open Access Journals (Sweden)

    Guan Kunliang

    2003-06-01

    Full Text Available Abstract Background G protein-coupled receptors (GPCRs interact with heterotrimeric GTP-binding proteins (G proteins to modulate acute changes in intracellular messenger levels and ion channel activity. In contrast, long-term changes in cellular growth, proliferation and differentiation are often mediated by tyrosine kinase receptors and certain GPCRs by activation of mitogen-activated protein (MAP kinases. Complex interactions occur between these signaling pathways, but the specific mechanisms of such regulatory events are not well-understood. In particular it is not clear whether GPCRs are modulated by tyrosine kinase receptor-MAP kinase pathways. Results Here we describe tyrosine kinase receptor regulation of a GPCR via MAP kinase. Insulin reduced the activity of the 5-HT2C receptor in choroid plexus cells which was blocked by the MAP kinase kinase (MEK inhibitor, PD 098059. We demonstrate that the inhibitory effect of insulin and insulin-like growth factor type 1 (IGF-1 on the 5-HT2C receptor is dependent on tyrosine kinase, RAS and MAP kinase. The effect may be receptor-specific: insulin had no effect on another GPCR that shares the same G protein signaling pathway as the 5-HT2C receptor. This effect is also direct: activated MAP kinase mimicked the effect of insulin, and removing a putative MAP kinase site from the 5-HT2C receptor abolished the effect of insulin. Conclusion These results show that insulin signaling can inhibit 5-HT2C receptor activity and suggest that MAP kinase may play a direct role in regulating the function of a specific GPCR.

  15. The selective estrogen receptor modulator raloxifene inhibits neutrophil extracellular trap formation.

    Directory of Open Access Journals (Sweden)

    Roxana Flores

    2016-12-01

    Full Text Available Raloxifene is a selective estrogen receptor modulator typically prescribed for the prevention/treatment of osteoporosis in postmenopausal women. Although raloxifene is known to have anti-inflammatory properties, its effect on human neutrophils, the primary phagocytic leukocytes of the immune system, remain poorly understood. Here, through a screen of pharmacologically active small molecules, we find that raloxifene prevents neutrophil cell death in response to the classical activator phorbol 12-myristate 13-acetate (PMA, a compound known to induce formation of DNA-based neutrophil extracellular traps (NETs. Inhibition of PMA-induced NET production by raloxifene was confirmed using quantitative and imaging-based assays. Human neutrophils from both male and female donors express the nuclear estrogen receptors ERα and ERβ, known targets of raloxifene. Like raloxifene, selective antagonists of these receptors inhibit PMA-induced NET production. Furthermore, raloxifene inhibited PMA-induced ERK phosphorylation but not reactive oxygen species (ROS production, pathways known to be key modulators of NET production. Finally, we found that raloxifene inhibited PMA-induced, NET-based killing of the leading human bacterial pathogen, methicillin-resistant Staphylococcus aureus (MRSA. Our results reveal that raloxifene is a potent modulator of neutrophil function and NET production.

  16. Inhibition of Estrogen Receptor Action by the Orphan Receptors, SHP and DAX-1

    National Research Council Canada - National Science Library

    DiRenzo, James

    2002-01-01

    In support of DoD grant # DAMD17-99-1-9163, we present our progress towards understanding the function of mechanisms of action of two orphan nuclear receptors, SHP and DAX-I as inhibitors of ER alpha and ER beta action...

  17. Effects on food intake and blood lipids of cannabinoid receptor 1 antagonist treatment in lean rats.

    Science.gov (United States)

    Bennetzen, Marianne F; Nielsen, Maria P; Richelsen, Bjørn; Pedersen, Steen B

    2008-11-01

    Endocannabinoids act through the cannabinoid receptor 1 (CB1) and has both orexigenic and peripheral metabolic effects. It is not yet fully understood whether all the beneficial effects on the metabolic profile by CB1 antagonism are induced by the weight loss or also by direct peripheral effects. The present study was intended to further elucidate this question and to investigate whether tolerance development to the hypophagic effect could be attenuated by cyclic treatment. We performed an intervention study in 40 lean rats over 4 weeks. The rats were divided in four groups: a control group, two groups treated with the CB1 antagonist Rimonabant either continuously or cyclically, and one group pair fed with the continuous Rimonabant group to obtain the same body weight. During the first 6 days, food intake was less in the continuous Rimonabant group compared to the control group (P acids (nonesterified fatty acid, NEFA) were significantly reduced in both treated groups compared to the untreated groups, and levels of triglycerides showed the same tendency. Cyclic treatment with Rimonabant is able to inhibit tolerance development on food intake, which resulted in reduction in body weight. Rimonabant treatment is associated with reduced serum levels of glycerol, NEFA, and triglyceride which seem independent of body weight changes.

  18. Striatal D1- and D2-type dopamine receptors are linked to motor response inhibition in human subjects.

    Science.gov (United States)

    Robertson, Chelsea L; Ishibashi, Kenji; Mandelkern, Mark A; Brown, Amira K; Ghahremani, Dara G; Sabb, Fred; Bilder, Robert; Cannon, Tyrone; Borg, Jacqueline; London, Edythe D

    2015-04-15

    Motor response inhibition is mediated by neural circuits involving dopaminergic transmission; however, the relative contributions of dopaminergic signaling via D1- and D2-type receptors are unclear. Although evidence supports dissociable contributions of D1- and D2-type receptors to response inhibition in rats and associations of D2-type receptors to response inhibition in humans, the relationship between D1-type receptors and response inhibition has not been evaluated in humans. Here, we tested whether individual differences in striatal D1- and D2-type receptors are related to response inhibition in human subjects, possibly in opposing ways. Thirty-one volunteers participated. Response inhibition was indexed by stop-signal reaction time on the stop-signal task and commission errors on the continuous performance task, and tested for association with striatal D1- and D2-type receptor availability [binding potential referred to nondisplaceable uptake (BPND)], measured using positron emission tomography with [(11)C]NNC-112 and [(18)F]fallypride, respectively. Stop-signal reaction time was negatively correlated with D1- and D2-type BPND in whole striatum, with significant relationships involving the dorsal striatum, but not the ventral striatum, and no significant correlations involving the continuous performance task. The results indicate that dopamine D1- and D2-type receptors are associated with response inhibition, and identify the dorsal striatum as an important locus of dopaminergic control in stopping. Moreover, the similar contribution of both receptor subtypes suggests the importance of a relative balance between phasic and tonic dopaminergic activity subserved by D1- and D2-type receptors, respectively, in support of response inhibition. The results also suggest that the stop-signal task and the continuous performance task use different neurochemical mechanisms subserving motor response inhibition. Copyright © 2015 the authors 0270-6474/15/355990-08$15.00/0.

  19. Basolateral P2X receptors mediate inhibition of NaCl transport in mouse medullary thick ascending limb (mTAL)

    DEFF Research Database (Denmark)

    Marques, Rita D; de Bruijn, Pauline I.A.; Sørensen, Mads Vaarby

    2012-01-01

    Extracellular nucleotides regulate epithelial transport via luminal and basolateral P2 receptors. Renal epithelia express multiple P2 receptors, which mediate significant inhibition of solute absorption. Recently, we identified several P2 receptors in the medullary thick ascending limb (m...

  20. Blocking beta 2-adrenergic receptor inhibits dendrite ramification in a mouse model of Alzheimer's disease.

    Science.gov (United States)

    Wu, Qin; Sun, Jin-Xia; Song, Xiang-He; Wang, Jing; Xiong, Cun-Quan; Teng, Fei-Xiang; Gao, Cui-Xiang

    2017-09-01

    Dendrite ramification affects synaptic strength and plays a crucial role in memory. Previous studies revealed a correlation between beta 2-adrenergic receptor dysfunction and Alzheimer's disease (AD), although the mechanism involved is still poorly understood. The current study investigated the potential effect of the selective β 2 -adrenergic receptor antagonist, ICI 118551 (ICI), on Aβ deposits and AD-related cognitive impairment. Morris water maze test results demonstrated that the performance of AD-transgenic (TG) mice treated with ICI (AD-TG/ICI) was significantly poorer compared with NaCl-treated AD-TG mice (AD-TG/NaCl), suggesting that β 2 -adrenergic receptor blockage by ICI might reduce the learning and memory abilities of mice. Golgi staining and immunohistochemical staining revealed that blockage of the β 2 -adrenergic receptor by ICI treatment decreased the number of dendritic branches, and ICI treatment in AD-TG mice decreased the expression of hippocampal synaptophysin and synapsin 1. Western blot assay results showed that the blockage of β 2 -adrenergic receptor increased amyloid-β accumulation by downregulating hippocampal α-secretase activity and increasing the phosphorylation of amyloid precursor protein. These findings suggest that blocking the β 2 -adrenergic receptor inhibits dendrite ramification of hippocampal neurons in a mouse model of AD.

  1. Acute stimulation of brain mu opioid receptors inhibits glucose-stimulated insulin secretion via sympathetic innervation.

    Science.gov (United States)

    Tudurí, Eva; Beiroa, Daniel; Stegbauer, Johannes; Fernø, Johan; López, Miguel; Diéguez, Carlos; Nogueiras, Rubén

    2016-11-01

    Pancreatic insulin-secreting β-cells express opioid receptors, whose activation by opioid peptides modulates hormone secretion. Opioid receptors are also expressed in multiple brain regions including the hypothalamus, where they play a role in feeding behavior and energy homeostasis, but their potential role in central regulation of glucose metabolism is unknown. Here, we investigate whether central opioid receptors participate in the regulation of insulin secretion and glucose homeostasis in vivo. C57BL/6J mice were acutely treated by intracerebroventricular (i.c.v.) injection with specific agonists for the three main opioid receptors, kappa (KOR), delta (DOR) and mu (MOR) opioid receptors: activation of KOR and DOR did not alter glucose tolerance, whereas activation of brain MOR with the specific agonist DAMGO blunted glucose-stimulated insulin secretion (GSIS), reduced insulin sensitivity, increased the expression of gluconeogenic genes in the liver and, consequently, impaired glucose tolerance. Pharmacological blockade of α2A-adrenergic receptors prevented DAMGO-induced glucose intolerance and gluconeogenesis. Accordingly, DAMGO failed to inhibit GSIS and to impair glucose tolerance in α2A-adrenoceptor knockout mice, indicating that the effects of central MOR activation on β-cells are mediated via sympathetic innervation. Our results show for the first time a new role of the central opioid system, specifically the MOR, in the regulation of insulin secretion and glucose metabolism. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Activation of GABAB receptors inhibits protein kinase B /Glycogen Synthase Kinase 3 signaling

    Directory of Open Access Journals (Sweden)

    Lu Frances Fangjia

    2012-11-01

    Full Text Available Abstract Accumulated evidence has suggested that potentiation of cortical GABAergic inhibitory neurotransmission may be a key mechanism in the treatment of schizophrenia. However, the downstream molecular mechanisms related to GABA potentiation remain unexplored. Recent studies have suggested that dopamine D2 receptor antagonists, which are used in the clinical treatment of schizophrenia, modulate protein kinase B (Akt/glycogen synthase kinase (GSK-3 signaling. Here we report that activation of GABAB receptors significantly inhibits Akt/GSK-3 signaling in a β-arrestin-dependent pathway. Agonist stimulation of GABAB receptors enhances the phosphorylation of Akt (Thr-308 and enhances the phosphorylation of GSK-3α (Ser-21/β (Ser-9 in both HEK-293T cells expressing GABAB receptors and rat hippocampal slices. Furthermore, knocking down the expression of β-arrestin2 using siRNA abolishes the GABAB receptor-mediated modulation of GSK-3 signaling. Our data may help to identify potentially novel targets through which GABAB receptor agents may exert therapeutic effects in the treatment of schizophrenia.

  3. Inhibition of amygdaloid dopamine D2 receptors impairs emotional learning measured with fear-potentiated startle.

    Science.gov (United States)

    Greba, Q; Gifkins, A; Kokkinidis, L

    2001-04-27

    Considerable advances have been made in understanding the neurocircuitry underlying the acquisition and expression of Pavlovian conditioned fear responses. Within the complex cellular and molecular processes mediating fearfulness, amygdaloid dopamine (DA), originating from cells in the ventral tegmental area (VTA) of the midbrain, is thought to contribute to fear-motivated responding. Considering that blockade of DA D(2) receptors is a common mechanism of action for antipsychotic agents, we hypothesized that inhibition of D(2) receptors in the amygdala may be involved in the antiparanoid effects of these drugs. To assess the role of amygdaloid DA D(2) receptors in aversive emotionality, the D(2) receptor antagonist raclopride was infused into the amygdala prior to Pavlovian fear conditioning. Potentiated startle was used as a behavioral indicator of fear and anxiety. Classical fear conditioning and acoustic startle testing were conducted in a single session allowing for the concomitant assessment of shock reactivity with startle enhancement. Depending on dose, the results found conditioned fear acquisition and retention to be impaired following administration of raclopride into the amygdala. Additionally, the learning deficit was dissociated from shock detection and from fear expression assessed with the shock sensitization of acoustic startle. These findings further refine the known neural mechanisms of amygdala-based emotional learning and memory and were interpreted to suggest that, along with D(1) receptors, D(2) receptors in the amygdala may mediate the formation and the retention of newly-acquired fear associations.

  4. Inhibition of allergen-induced basophil activation by ASM-024, a nicotinic receptor ligand.

    Science.gov (United States)

    Watson, Brittany M; Oliveria, John Paul; Nusca, Graeme M; Smith, Steven G; Beaudin, Sue; Dua, Benny; Watson, Rick M; Assayag, Evelynne Israël; Cormier, Yvon F; Sehmi, Roma; Gauvreau, Gail M

    2014-01-01

    Nicotinic acetylcholine receptors (nAChRs) were identified on eosinophils and shown to regulate inflammatory responses, but nAChR expression on basophils has not been explored yet. We investigated surface receptor expression of nAChR α4, α7 and α1/α3/α5 subunits on basophils. Furthermore, we examined the effects of ASM-024, a synthetic nicotinic ligand, on in vitro anti-IgE and in vivo allergen-induced basophil activation. Basophils were enriched from the peripheral blood of allergic donors and the expression of nAChR subunits and muscarinic receptors was determined. Purified basophils were stimulated with anti-IgE in the presence of ASM-024 with or without muscarinic or nicotinic antagonists for the measurement of CD203c expression and histamine release. The effect of 9 days of treatment with 50 and 200 mg ASM-024 on basophil CD203c expression was examined in the blood of mild allergic asthmatics before and after allergen inhalation challenge. nAChR α4, α7 and α1/α3/α5 receptor subunit expression was detected on basophils. Stimulation of basophils with anti-IgE increased CD203c expression and histamine release, which was inhibited by ASM-024 (10(-5) to 10(-)(3) M, p ASM-024 was reversed in the presence of muscarinic and nicotinic antagonists. In subjects with mild asthma, ASM-024 inhalation significantly inhibited basophil CD203c expression measured 24 h after allergen challenge (p = 0.03). This study shows that ASM-024 inhibits IgE- and allergen-induced basophil activation through both nicotinic and muscarinic receptors, and suggests that ASM-024 may be an efficacious agent for modulating allergic asthma responses. © 2015 S. Karger AG, Basel.

  5. Full Fatty Acid Amide Hydrolase Inhibition Combined with Partial Monoacylglycerol Lipase Inhibition: Augmented and Sustained Antinociceptive Effects with Reduced Cannabimimetic Side Effects in Mice.

    Science.gov (United States)

    Ghosh, Sudeshna; Kinsey, Steven G; Liu, Qing-Song; Hruba, Lenka; McMahon, Lance R; Grim, Travis W; Merritt, Christina R; Wise, Laura E; Abdullah, Rehab A; Selley, Dana E; Sim-Selley, Laura J; Cravatt, Benjamin F; Lichtman, Aron H

    2015-08-01

    Inhibition of fatty acid amide hydrolase (FAAH) or monoacylglycerol lipase (MAGL), the primary hydrolytic enzymes for the respective endocannabinoids N-arachidonoylethanolamine (AEA) and 2-arachidonylglycerol (2-AG), produces antinociception but with minimal cannabimimetic side effects. Although selective inhibitors of either enzyme often show partial efficacy in various nociceptive models, their combined blockade elicits augmented antinociceptive effects, but side effects emerge. Moreover, complete and prolonged MAGL blockade leads to cannabinoid receptor type 1 (CB1) receptor functional tolerance, which represents another challenge in this potential therapeutic strategy. Therefore, the present study tested whether full FAAH inhibition combined with partial MAGL inhibition would produce sustained antinociceptive effects with minimal cannabimimetic side effects. Accordingly, we tested a high dose of the FAAH inhibitor PF-3845 (N-​3-​pyridinyl-​4-​[[3-​[[5-​(trifluoromethyl)-​2-​pyridinyl]oxy]phenyl]methyl]-​1-​piperidinecarboxamide; 10 mg/kg) given in combination with a low dose of the MAGL inhibitor JZL184 [4-nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate] (4 mg/kg) in mouse models of inflammatory and neuropathic pain. This combination of inhibitors elicited profound increases in brain AEA levels (>10-fold) but only 2- to 3-fold increases in brain 2-AG levels. This combination produced significantly greater antinociceptive effects than single enzyme inhibition and did not elicit common cannabimimetic effects (e.g., catalepsy, hypomotility, hypothermia, and substitution for Δ(9)-tetrahydrocannabinol in the drug-discrimination assay), although these side effects emerged with high-dose JZL184 (i.e., 100 mg/kg). Finally, repeated administration of this combination did not lead to tolerance to its antiallodynic actions in the carrageenan assay or CB1 receptor functional tolerance. Thus, full FAAH inhibition

  6. Microbial Culturomics Application for Global Health: Noncontiguous Finished Genome Sequence and Description of Pseudomonas massiliensis Strain CB-1T sp. nov. in Brazil.

    Science.gov (United States)

    Bardet, Lucie; Cimmino, Teresa; Buffet, Clémence; Michelle, Caroline; Rathored, Jaishriram; Tandina, Fatalmoudou; Lagier, Jean-Christophe; Khelaifia, Saber; Abrahão, Jônatas; Raoult, Didier; Rolain, Jean-Marc

    2018-02-01

    Culturomics is a new postgenomics field that explores the microbial diversity of the human gut coupled with taxono-genomic strategy. Culturomics, and the microbiome science more generally, are anticipated to transform global health diagnostics and inform the ways in which gut microbial diversity contributes to human health and disease, and by extension, to personalized medicine. Using culturomics, we report in this study the description of strain CB1 T ( = CSUR P1334 = DSM 29075), a new species isolated from a stool specimen from a 37-year-old Brazilian woman. This description includes phenotypic characteristics and complete genome sequence and annotation. Strain CB1 T is a gram-negative aerobic and motile bacillus, exhibits neither catalase nor oxidase activities, and presents a 98.3% 16S rRNA sequence similarity with Pseudomonas putida. The 4,723,534 bp long genome contains 4239 protein-coding genes and 74 RNA genes, including 15 rRNA genes (5 16S rRNA, 4 23S rRNA, and 6 5S rRNA) and 59 tRNA genes. Strain CB1 T was named Pseudomonas massiliensis sp. nov. and classified into the family Pseudomonadaceae. This study demonstrates the usefulness of microbial culturomics in exploration of human microbiota in diverse geographies and offers new promise for incorporating new omics technologies for innovation in diagnostic medicine and global health.

  7. The Cellular Prion Protein Prevents Copper-Induced Inhibition of P2X4 Receptors

    Directory of Open Access Journals (Sweden)

    Ramón A. Lorca

    2011-01-01

    Full Text Available Although the physiological function of the cellular prion protein (PrPC remains unknown, several evidences support the notion of its role in copper homeostasis. PrPC binds Cu2+ through a domain composed by four to five repeats of eight amino acids. Previously, we have shown that the perfusion of this domain prevents and reverses the inhibition by Cu2+ of the adenosine triphosphate (ATP-evoked currents in the P2X4 receptor subtype, highlighting a modulatory role for PrPC in synaptic transmission through regulation of Cu2+ levels. Here, we study the effect of full-length PrPC in Cu2+ inhibition of P2X4 receptor when both are coexpressed. PrPC expression does not significantly change the ATP concentration-response curve in oocytes expressing P2X4 receptors. However, the presence of PrPC reduces the inhibition by Cu2+ of the ATP-elicited currents in these oocytes, confirming our previous observations with the Cu2+ binding domain. Thus, our observations suggest a role for PrPC in modulating synaptic activity through binding of extracellular Cu2+.

  8. Distinct roles of synaptic and extrasynaptic GABAA receptors in striatal inhibition dynamics

    Directory of Open Access Journals (Sweden)

    Ruixi eLuo

    2013-11-01

    Full Text Available Striatonigral and striatopallidal projecting medium spiny neurons (MSNs express dopamine D1 (D1+ and D2 receptors (D2+, respectively. Both classes receive extensive GABAergic input via expression of synaptic, perisynaptic and extrasynaptic GABAA receptors. The activation patterns of different presynaptic GABAergic neurons produce transient and sustained GABAA receptor-mediated conductance that fulfill distinct physiological roles. We performed single and dual whole cell recordings from striatal neurons in mice expressing fluorescent proteins in interneurons and MSNs. We report specific inhibitory dynamics produced by distinct activation patterns of presynaptic GABAergic neurons as source of synaptic, perisynaptic and extrasynaptic inhibition. Synaptic GABAA receptors in MSNs contain the α2, γ2 and a β subunit. In addition, there is evidence for the developmental increase of the α1 subunit that contributes to faster inhibitory postsynaptic current (IPSC. Tonic GABAergic currents in MSNs from adult mice are carried by extrasynaptic receptors containing the α4 and δ subunit, while in younger mice this current is mediated by receptors that contain the α5 subunit. Both forms of tonic currents are differentially expressed in D1+ and D2+ MSNs. This study extends these findings by relating presynaptic activation with pharmacological analysis of inhibitory conductance in mice where the β3 subunit is conditionally removed in fluorescently labeled D2+ MSNs and in mice with global deletion of the δ subunit. Our results show that responses to low doses of gaboxadol (2μM, a GABAA receptor agonist with preference to δ subunit, are abolished in the δ but not the β3 subunit knock out mice. This suggests that the β3 subunit is not a component of the adult extrasynaptic receptor pool, in contrast to what has been shown for tonic current in young mice. Deletion of the β3 subunit from D2+ MSNs however, removed slow spontaneous IPSCs, implicating its

  9. The future of type 1 cannabinoid receptor allosteric ligands.

    Science.gov (United States)

    Alaverdashvili, Mariam; Laprairie, Robert B

    2018-02-01

    Allosteric modulation of the type 1 cannabinoid receptor (CB1R) holds great therapeutic potential. This is because allosteric modulators do not possess intrinsic efficacy, but instead augment (positive allosteric modulation) or diminish (negative allosteric modulation) the receptor's response to endogenous ligand. Consequently, CB1R allosteric modulators have an effect ceiling which allows for the tempering of CB1R signaling without the desensitization, tolerance, dependence, and psychoactivity associated with orthosteric compounds. Pain, movement disorders, epilepsy, obesity are all potential therapeutic targets for CB1R allosteric modulation. Several challenges exist for the development of CB1R allosteric modulators, such as receptor subtype specificity, translation to in vivo systems, and mixed allosteric/agonist/inverse agonist activity. Despite these challenges, elucidation of crystal structures of CB1R and compound design based on structure-activity relationships will advance the field. In this review, we will cover recent progress for CB1R allosteric modulators and discuss the future promise of this research.

  10. GABA(B) receptor modulation of feedforward inhibition through hippocampal neurogliaform cells.

    Science.gov (United States)

    Price, Christopher J; Scott, Ricardo; Rusakov, Dmitri A; Capogna, Marco

    2008-07-02

    Feedforward inhibition of neurons is a fundamental component of information flow control in the brain. We studied the roles played by neurogliaform cells (NGFCs) of stratum lacunosum moleculare of the hippocampus in providing feedforward inhibition to CA1 pyramidal cells. We recorded from synaptically coupled pairs of anatomically identified NGFCs and CA1 pyramidal cells and found that, strikingly, a single presynaptic action potential evoked a biphasic unitary IPSC (uIPSC), consisting of two distinct components mediated by GABA(A) and GABA(B) receptors. A GABA(B) receptor-mediated unitary response has not previously been observed in hippocampal excitatory neurons. The decay of the GABA(A) receptor-mediated response was slow (time constant = 50 ms), and was tightly regulated by presynaptic GABA(B) receptors. Surprisingly, the GABA(B) receptor ligands baclofen and (2S)-3-{[(1S)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl}(phenylmethyl)phosphinic acid (CGP55845), while affecting the NGFC-mediated uIPSCs, had no effect on action potential-evoked presynaptic Ca2+ signals monitored in individual axonal boutons of NGFCs with two-photon microscopy. In contrast, baclofen clearly depressed presynaptic Ca2+ transients in non-NGF interneurons. Changes in extracellular Ca2+ concentration that mimicked the effects of baclofen or CGP55845 on uIPSCs significantly altered presynaptic Ca2+ transients. Electrophysiological data suggest that GABA(B) receptors expressed by NGFCs contribute to the dynamic control of the excitatory input to CA1 pyramidal neurons from the temporoammonic path. The NGFC-CA1 pyramidal cell connection therefore provides a unique and subtle mechanism to shape the integration time domain for signals arriving via a major excitatory input to CA1 pyramidal cells.

  11. Effects of protease-activated receptor 1 inhibition on anxiety and fear following status epilepticus.

    Science.gov (United States)

    Bogovyk, Ruslan; Lunko, Oleksii; Fedoriuk, Mihail; Isaev, Dmytro; Krishtal, Oleg; Holmes, Gregory L; Isaeva, Elena

    2017-02-01

    Protease-activated receptor 1 (PAR1) is an important contributor to the pathogenesis of a variety of brain disorders associated with a risk of epilepsy development. Using the lithium-pilocarpine model of temporal lobe epilepsy (TLE), we recently showed that inhibition of this receptor during the first ten days after pilocarpine-induced status epilepticus (SE) results in substantial anti-epileptogenic and neuroprotective effects. As PAR1 is expressed in the central nervous system regions of importance for processing emotional reactions, including amygdala and hippocampus, and TLE is frequently associated with a chronic alteration of the functions of these regions, we tested the hypothesis that PAR1 inhibition could modulate emotionally driven behavioral responses of rats experiencing SE. We showed that SE induces a chronic decrease in the animals' anxiety-related behavior and an increase of locomotor activity. PAR1 inhibition after SE abolished the alteration of the anxiety level but does not affect the increase of locomotor activity in the open field and elevated plus maze tests. Moreover, while PAR1 inhibition produces an impairment of memory recall in the context fear conditioning paradigm in the control group, it substantially improves contextual and cued fear learning in rats experiencing SE. These data suggest that PAR1-dependent signaling is involved in the mechanisms underlying emotional disorders in epilepsy. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Different mechanisms are involved in the antibody mediated inhibition of ligand binding to the urokinase receptor

    DEFF Research Database (Denmark)

    List, K; Høyer-Hansen, G; Rønne, E

    1999-01-01

    Certain monoclonal antibodies are capable of inhibiting the biological binding reactions of their target proteins. At the molecular level, this type of effect may be brought about by completely different mechanisms, such as competition for common binding determinants, steric hindrance or interfer......Certain monoclonal antibodies are capable of inhibiting the biological binding reactions of their target proteins. At the molecular level, this type of effect may be brought about by completely different mechanisms, such as competition for common binding determinants, steric hindrance......) can be employed as a highly useful tool to characterize the inhibitory mechanism of specific antagonist antibodies. Two inhibitory antibodies against uPAR, mAb R3 and mAb R5, were shown to exhibit competitive and non-competitive inhibition, respectively, of ligand binding to the receptor. The former...

  13. Anterior cingulate serotonin 1B receptor binding is associated with emotional response inhibition

    DEFF Research Database (Denmark)

    da Cunha-Bang, Sofi; Hjordt, Liv Vadskjær; Dam, Vibeke Høyrup

    2017-01-01

    -offender controls, completed an emotional Go/NoGo task requiring inhibition of prepotent motor responses to emotional facial expressions. We also measured cerebral serotonin 1B receptor (5-HT1BR) binding with [11C]AZ10419369 positron emission tomography within regions of the frontal cortex. We hypothesized that 5......-HT1BR would be positively associated with false alarms (failures to inhibit nogo responses) in the context of aversive (angry and fearful) facial expressions. Across groups, we found that frontal cortex 5-HT1BR binding was positively correlated with false alarms when angry faces were go stimuli......Serotonin has a well-established role in emotional processing and is a key neurotransmitter in impulsive aggression, presumably by facilitating response inhibition and regulating subcortical reactivity to aversive stimuli. In this study 44 men, of whom 19 were violent offenders and 25 were non...

  14. Computer modeling of Cannabinoid receptor type 1

    Directory of Open Access Journals (Sweden)

    Sapundzhi Fatima

    2018-01-01

    Full Text Available Cannabinoid receptors are important class of receptors as they are involved in various physiological processes such as appetite, pain-sensation, mood, and memory. It is important to design receptor-selective ligands in order to treat a particular disorder. The aim of the present study is to model the structure of cannabinoid receptor CB1 and to perform docking between obtained models and known ligands. Two models of CBR1 were prepared with two different methods (Modeller of Chimera and MOE. They were used for docking with GOLD 5.2. It was established a high correlation between inhibitory constant Ki of CB1 cannabinoid ligands and the ChemScore scoring function of GOLD, which concerns both models. This suggests that the models of the CB1 receptors obtained could be used for docking studies and in further investigation and design of new potential, selective and active cannabinoids with the desired effects.

  15. Gambogic acid inhibits multiple myeloma mediated osteoclastogenesis through suppression of chemokine receptor CXCR4 signaling pathways.

    Science.gov (United States)

    Pandey, Manoj K; Kale, Vijay P; Song, Chunhua; Sung, Shen-shu; Sharma, Arun K; Talamo, Giampaolo; Dovat, Sinisa; Amin, Shantu G

    2014-10-01

    Bone disease, characterized by the presence of lytic lesions and osteoporosis is the hallmark of multiple myeloma (MM). Stromal cell-derived factor 1α (SDF-1α) and its receptor, CXC chemokine receptor 4 (CXCR4), has been implicated as a regulator of bone resorption, suggesting that agents that can suppress SDF1α/CXCR4 signaling might inhibit osteoclastogenesis, a process closely linked to bone resorption. We, therefore, investigated whether gambogic acid (GA), a xanthone, could inhibit CXCR4 signaling and suppress osteoclastogenesis induced by MM cells. Through docking studies we predicted that GA directly interacts with CXCR4. This xanthone down-regulates the expression of CXCR4 on MM cells in a dose- and time-dependent manner. The down-regulation of CXCR4 was not due to proteolytic degradation, but rather GA suppresses CXCR4 mRNA expression by inhibiting nuclear factor-kappa B (NF-κB) DNA binding. This was further confirmed by quantitative chromatin immunoprecipitation assay, as GA inhibits p65 binding at the CXCR4 promoter. GA suppressed SDF-1α-induced chemotaxis of MM cells and downstream signaling of CXCR4 by inhibiting phosphorylation of Akt, p38, and Erk1/2 in MM cells. GA abrogated the RANKL-induced differentiation of macrophages to osteoclasts in a dose- and time-dependent manner. In addition, we found that MM cells induced differentiation of macrophages to osteoclasts, and that GA suppressed this process. Importantly, suppression of osteoclastogenesis by GA was mediated through IL-6 inhibition. Overall, our results show that GA is a novel inhibitor of CXCR4 expression and has a strong potential to suppress osteoclastogenesis mediated by MM cells. Published by Elsevier Inc.

  16. Exercise reduces adipose tissue via cannabinoid receptor type 1 which is regulated by peroxisome proliferator-activated receptor

    International Nuclear Information System (INIS)

    Yan Zhencheng; Liu Daoyan; Zhang Lili; Shen Chenyi; Ma Qunli; Cao Tingbing; Wang Lijuan; Nie Hai; Zidek, Walter; Tepel, Martin; Zhu Zhiming

    2007-01-01

    Obesity is one major cardiovascular risk factor. We tested effects of endurance exercise on cannabinoid receptor type 1 (CB1) and peroxisome proliferator-activated receptor-δ (PPAR-δ)-dependent pathways in adipose tissue. Male Wistar rats were randomly assigned to standard laboratory chow or a high-fat diet without and with regular endurance exercise. Exercise in rats on high-fat diet significantly reduced visceral fat mass, blood pressure, and adipocyte size (each p < 0.05). Adipocyte hypertrophy induced by high-fat diet was accompanied by increased CB1 expression in adipose tissue, whereas exercise significantly reduced CB1 expression (each p < 0.05). CB1 receptor expression and adipocyte differentiation were directly regulated by PPAR-δ. Adipocyte hypertrophy induced by high-fat diet was accompanied by reduced PPAR-δ. Furthermore, selective silencing of PPAR-δ by RNA interference in 3T3-L1-preadipocyte cells significantly increased CB1 expression from 1.00 ± 0.06 (n = 3) to 1.91 ± 0.06 (n = 3; p < 0.01) and increased adipocyte differentiation, whereas adenovirus-mediated overexpression of PPAR-δ significantly reduced CB1 expression to 0.39 ± 0.03 (n = 3; p < 0.01) and reduced adipocyte differentiation. In the presence of the CB1 antagonist rimonabant adipocyte differentiation in stimulated 3T3 L1 preadipocyte cells was significantly reduced. The study indicates that high-fat diet-induced hypertrophy of adipocytes is associated with increased CB1 receptor expression which is directly regulated by PPAR-δ. Both CB1 and PPAR-δ are intimately involved in therapeutic interventions against a most important cardiovascular risk factor

  17. Hemin inhibits internalization of transferrin by reticulocytes and promotes phosphorylation of the membrane transferrin receptor

    International Nuclear Information System (INIS)

    Cox, T.M.; O'Donnell, M.W.; Aisen, P.; London, I.M.

    1985-01-01

    Addition of hemin to reticulocytes inhibits incorporation of iron from transferrin. Heme also regulates protein synthesis in immature erythroid cells through its effects on phosphorylation of the initiation factor eIF-2. The authors have examined its effects on endocytosis of iron-transferrin and phosphorylation of the transferrin receptor. Hemin reduced iron transport but increased cell-associated transferrin. During uptake of 125 I-labeled transferrin in the steady state, the use of a washing technique to dissociate bound transferrin on the cell membrane showed that radioligand accumulated on the surface of hemin-treated cells. Receptor phosphorylation was investigated by immunoprecipitation of reticulocyte extracts after metabolic labeling with [ 32 P]P/sub i/. In the absence of ligand, phosphorylated receptor was chiefly localized on cell stroma. Exposure to transferrin increased cytosolic phosphorylated receptor from 15-30% to approximately 50% of the total, an effect overcome by hemin treatment. The findings suggest a possible relationship of phosphorylation to endocytosis of the transferrin receptor in reticulocytes

  18. Constitutive cannabinoid 1 and mu opioid receptor activity in the ventral tegmental area: occurrence, function and therapeutic relevance

    NARCIS (Netherlands)

    Meye, F.J.

    2012-01-01

    Cannabinoid 1 receptors (CB1Rs) play a crucial role in regulating systems dedicated to processing rewards and emotions. It was known that in artificial systems, CB1Rs can exhibit activity that is independent of the typical agonist-driven form. However, it remained largely unclear whether this

  19. MiR-34a inhibits colon cancer proliferation and metastasis by inhibiting platelet-derived growth factor receptor α.

    Science.gov (United States)

    Li, Chunyan; Wang, Yulin; Lu, Shuming; Zhang, Zhuqing; Meng, Hua; Liang, Lina; Zhang, Yan; Song, Bo

    2015-11-01

    The microRNA (miRNA), miR‑34a is significant in colon cancer progression. In the present study, the role of miR‑34a in colon cancer cell proliferation and metastasis was investigated. It was found that the expression of miR‑34a in colon cancer tissues and cell lines was lower when compared with that of normal tissues and cells. Further research demonstrated that miR‑34a inhibited cell proliferation, induced G1 phase arrest, and suppressed metastasis and epithelial mesenchymal transition in colon cancer cells. Bioinformatic prediction indicated that platelet‑derived growth factor receptor α (PDGFRA) was a potential target gene of miR‑34a and a luciferase assay identified that PDGFRA was a novel direct target gene of miR‑34a. In addition, assays of western blot analyses and quantitative reverse‑transcription polymerase chain reaction confirmed that miR‑34a decreased PDGFRA mRNA expression and protein levels in colon cancer cells. Assessment of cellular function indicated that miR‑34a inhibited colon cancer progression via PDGFRA. These findings demonstrate that miR‑34a may act as a negative regulator in colon cancer by targeting PDGFRA.

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

    International Nuclear Information System (INIS)

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

    1989-01-01

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

  1. The aryl hydrocarbon receptor ligand ITE inhibits TGFβ1-induced human myofibroblast differentiation.

    Science.gov (United States)

    Lehmann, Geniece M; Xi, Xia; Kulkarni, Ajit A; Olsen, Keith C; Pollock, Stephen J; Baglole, Carolyn J; Gupta, Shikha; Casey, Ann E; Huxlin, Krystel R; Sime, Patricia J; Feldon, Steven E; Phipps, Richard P

    2011-04-01

    Fibrosis can occur in any human tissue when the normal wound healing response is amplified. Such amplification results in fibroblast proliferation, myofibroblast differentiation, and excessive extracellular matrix deposition. Occurrence of these sequelae in organs such as the eye or lung can result in severe consequences to health. Unfortunately, medical treatment of fibrosis is limited by a lack of safe and effective therapies. These therapies may be developed by identifying agents that inhibit critical steps in fibrotic progression; one such step is myofibroblast differentiation triggered by transforming growth factor-β1 (TGFβ1). In this study, we demonstrate that TGFβ1-induced myofibroblast differentiation is blocked in human fibroblasts by a candidate endogenous aryl hydrocarbon receptor (AhR) ligand 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE). Our data show that ITE disrupts TGFβ1 signaling by inhibiting the nuclear translocation of Smad2/3/4. Although ITE functions as an AhR agonist, and biologically persistent AhR agonists, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin, cause severe toxic effects, ITE exhibits no toxicity. Interestingly, ITE effectively inhibits TGFβ1-driven myofibroblast differentiation in AhR(-/-) fibroblasts: Its ability to inhibit TGFβ1 signaling is AhR independent. As supported by the results of this study, the small molecule ITE inhibits myofibroblast differentiation and may be useful clinically as an antiscarring agent. Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  2. Midazolam inhibits hippocampal long-term potentiation and learning through dual central and peripheral benzodiazepine receptor activation and neurosteroidogenesis

    OpenAIRE

    Tokuda, Kazuhiro; O’Dell, Kazuko A.; Izumi, Yukitoshi; Zorumski, Charles F.

    2010-01-01

    Benzodiazepines (BDZs) enhance γ-aminobutyric acid-A (GABAA) receptor inhibition by direct actions on central BDZ receptors (CBRs). Although some BDZs also bind mitochondrial receptors (translocator protein 18kDa, TSPO) and promote the synthesis of GABA-enhancing neurosteroids, the role of neurosteroids in the clinical effects of BDZs is unknown. In rat hippocampal slices, we compared midazolam, an anesthetic BDZ with clonazepam, an anticonvulsant/anxiolytic BDZ that activates CBRs selectivel...

  3. A novel thromboxane receptor antagonist, nstpbp5185, inhibits platelet aggregation and thrombus formation in animal models.

    Science.gov (United States)

    Huang, Shiu-Wen; Kuo, Heng-Lan; Hsu, Ming-Tsung; Tseng, Yufeng Jane; Lin, Shu-Wha; Kuo, Sheng-Chu; Peng, Hui-Chin; Lien, Jin-Cherng; Huang, Tur-Fu

    2016-08-01

    A novel benzimidazole derivative, nstpbp5185, was discovered through in vitro and in vivo evaluations for antiplatelet activity. Thromaboxane receptor (TP) is important in vascular physiology, haemostasis and pathophysiological thrombosis. Nstpbp5185 concentration-dependently inhibited human platelet aggregation caused by collagen, arachidonic acid and U46619. Nstpbp5185 caused a right-shift of the concentration-response curve of U46619 and competitively inhibited the binding of 3H-SQ-29548 to TP receptor expressed on HEK-293 cells, with an IC50 of 0.1 µM, indicating that nstpbp5185 is a TP antagonist. In murine thrombosis models, nstpbp5185 significantly prolonged the latent period in triggering platelet plug formation in mesenteric and FeCl3-induced thrombi formation, and increased the survival rate in pulmonary embolism model with less bleeding than aspirin. This study suggests nstpbp5185, an orally selective anti-thrombotic agent, acting through blockade of TXA2 receptor, may be efficacious for prevention or treatment of pathologic thrombosis.

  4. Oestrogen inhibits human colonic motility by a non-genomic cell membrane receptor-dependent mechanism.

    LENUS (Irish Health Repository)

    Hogan, A M

    2012-02-01

    BACKGROUND: Classical effects of oestrogen involve activation of target genes after binding nuclear receptors. Oestrogenic effects too rapid for DNA transcription (non-genomic) are known to occur. The effect of oestrogen on colonic motility is unknown despite the prevalence of gastrointestinal symptoms in pregnant and premenopausal women. METHODS: Histologically normal colon was obtained from proximal resection margins of colorectal carcinoma specimens. Circular smooth muscle strips were microdissected and suspended in organ baths under 1 g of tension. After equilibration, they were exposed to 17beta-oestradiol (n = 8) or bovine serum albumin (BSA)-conjugated 17beta-oestradiol (n = 8). Fulvestrant, an oestrogen receptor antagonist, was added to some baths (n = 8). Other strips were exposed to calphostin C or cycloheximide. Carbachol was added in increasing concentrations and contractile activity was recorded isometrically. RESULTS: Oestrogen inhibited colonic contractility (mean difference 19.7 per cent; n = 8, P < 0.001). In keeping with non-genomic, rapid-onset steroid action, the effect was apparent within minutes and reversible. It was observed with both 17beta-oestradiol and BSA-conjugated oestrogen, and was not altered by cycloheximide. Effects were inhibited by fulvestrant, suggesting receptor mediation. CONCLUSION: Oestrogen decreases contractility in human colonic smooth muscle by a non-genomic mechanism involving cell membrane coupling.

  5. MiR-125a TNF receptor-associated factor 6 to inhibit osteoclastogenesis

    International Nuclear Information System (INIS)

    Guo, Li-Juan; Liao, Lan; Yang, Li; Li, Yu; Jiang, Tie-Jian

    2014-01-01

    MicroRNAs (miRNAs) play important roles in osteoclastogenesis and bone resorption. In the present study, we found that miR-125a was dramatically down-regulated during macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL) induced osteoclastogenesis of circulating CD14+ peripheral blood mononuclear cells (PBMCs). Overexpression of miR-125a in CD14+ PBMCs inhibited osteoclastogenesis, while inhibition of miR-125a promoted osteoclastogenesis. TNF receptor-associated factor 6 (TRAF6), a transduction factor for RANKL/RANK/NFATc1 signal, was confirmed to be a target of miR-125a. EMSA and ChIP assays confirmed that NFATc1 bound to the promoter of the miR-125a. Overexpression of NFATc1 inhibited miR-125a transcription, and block of NFATc1 expression attenuated RANKL-regulated miR-125a transcription. Here, we reported that miR-125a played a biological function in osteoclastogenesis through a novel TRAF6/ NFATc1/miR-125a regulatory feedback loop. It suggests that regulation of miR-125a expression may be a potential strategy for ameliorating metabolic disease. - Highlights: • MiR-125a was significantly down-regulated in osteoclastogenesis of CD14+ PBMCs. • MiR-125a inhibited osteoclast differentiation by targeting TRAF6. • NFATc1 inhibited miR-125a transciption by binding to the promoter of miR-125a. • TRAF6/NFATc1 and miR-125a form a regulatory feedback loop in osteoclastogenesis

  6. Tributyltin and triphenyltin inhibit osteoclast differentiation through a retinoic acid receptor-dependent signaling pathway

    International Nuclear Information System (INIS)

    Yonezawa, Takayuki; Hasegawa, Shin-ichi; Ahn, Jae-Yong; Cha, Byung-Yoon; Teruya, Toshiaki; Hagiwara, Hiromi; Nagai, Kazuo; Woo, Je-Tae

    2007-01-01

    Organotin compounds, such as tributyltin (TBT) and triphenyltin (TPT), have been widely used in agriculture and industry. Although these compounds are known to have many toxic effects, including endocrine-disrupting effects, their effects on bone resorption are unknown. In this study, we investigated the effects of organotin compounds, such as monobutyltin (MBT), dibutyltin (DBT), TBT, and TPT, on osteoclast differentiation using mouse monocytic RAW264.7 cells. MBT and DBT had no effects, whereas TBT and TPT dose-dependently inhibited osteoclast differentiation at concentrations of 3-30 nM. Treatment with a retinoic acid receptor (RAR)-specific antagonist, Ro41-5253, restored the inhibition of osteoclastogenesis by TBT and TPT. TBT and TPT reduced receptor activator of nuclear factor-κB ligand (RANKL) induced nuclear factor of activated T cells (NFAT) c1 expression, and the reduction in NFATc1 expression was recovered by Ro41-5253. Our results suggest that TBT and TPT suppress osteoclastogenesis by inhibiting RANKL-induced NFATc1 expression via an RAR-dependent signaling pathway

  7. CDB-4124, a progesterone receptor modulator, inhibits mammary carcinogenesis by suppressing cell proliferation and inducing apoptosis.

    Science.gov (United States)

    Wiehle, Ronald; Lantvit, Daniel; Yamada, Tohru; Christov, Konstantin

    2011-03-01

    CDB-4124 (Proellex or telapristone acetate) is a modulator of progesterone receptor (PR) signaling, which is currently employed in preclinical studies for prevention and treatment of breast cancer and has been used in clinical studies for treatment of uterine fibroids and endometriosis. Here we provide evidence for its action on steroid hormone-signaling, cell cycle-regulated genes and in vivo on mammary carcinogenesis. When CDB-4124 is given to rats at 200 mg/kg for 24 months, it prevents the development of spontaneous mammary hyperplastic and premalignant lesions. Also, CDB-4124 given as subcutaneous pellets at two different doses suppressed, dose dependently, N-methyl-N-nitrosourea (MNU)-induced mammary carcinogenesis. The high dose (30 mg, over 84 days) increased tumor latency from 66 ± 24 days to 87 ± 20 days (P CDB-4124 inhibited cell proliferation and induced apoptosis in MNU-induced mammary tumors, which correlated with a decreased proportion of PR(+) tumor cells and with decreased serum progesterone. CDB-4124 did not affect serum estradiol. In a mechanistic study employing T47D cells we found that CDB-4124 suppressed G(1)/G(0)-S transition by inhibiting CDK2 and CDK4 expressions, which correlated with inhibition of estrogen receptor (ER) expression. Taken together, these data indicate that CDB-4124 can suppress the development of precancerous lesions and carcinogen-induced ER(+) mammary tumors in rats, and may have implications for prevention and treatment of human breast cancer.

  8. GLP-1 receptor stimulation depresses heart rate variability and inhibits neurotransmission to cardiac vagal neurons.

    Science.gov (United States)

    Griffioen, Kathleen J; Wan, Ruiqian; Okun, Eitan; Wang, Xin; Lovett-Barr, Mary Rachael; Li, Yazhou; Mughal, Mohamed R; Mendelowitz, David; Mattson, Mark P

    2011-01-01

    glucagon-like peptide 1 (GLP-1) is an incretin hormone released from the gut in response to food intake. Whereas GLP-1 acts in the periphery to inhibit glucagon secretion and stimulate insulin release, it also acts in the central nervous system to mediate autonomic control of feeding, body temperature, and cardiovascular function. Because of its role as an incretin hormone, GLP-1 receptor analogs are used as a treatment for type 2 diabetes. Central or peripheral administration of GLP-1 increases blood pressure and heart rate, possibly by activating brainstem autonomic nuclei and increasing vagus nerve activity. However, the mechanism(s) by which GLP-1 receptor stimulation affects cardiovascular function are unknown. We used the long-lasting GLP-1 receptor agonist Exendin-4 (Ex-4) to test the hypothesis that GLP-1 signalling modulates central parasympathetic control of heart rate. using a telemetry system, we assessed heart rate in mice during central Ex-4 administration. Heart rate was increased by both acute and chronic central Ex-4 administration. Spectral analysis indicated that the high frequency and low frequency powers of heart rate variability were diminished by Ex-4 treatment. Finally, Ex-4 decreased both excitatory glutamatergic and inhibitory glycinergic neurotransmission to preganglionic parasympathetic cardiac vagal neurons. these data suggest that central GLP-1 receptor stimulation diminishes parasympathetic modulation of the heart thereby increasing heart rate.

  9. GABA type a receptor trafficking and the architecture of synaptic inhibition.

    Science.gov (United States)

    Lorenz-Guertin, Joshua M; Jacob, Tija C

    2018-03-01

    Ubiquitous expression of GABA type A receptors (GABA A R) in the central nervous system establishes their central role in coordinating most aspects of neural function and development. Dysregulation of GABAergic neurotransmission manifests in a number of human health disorders and conditions that in certain cases can be alleviated by drugs targeting these receptors. Precise changes in the quantity or activity of GABA A Rs localized at the cell surface and at GABAergic postsynaptic sites directly impact the strength of inhibition. The molecular mechanisms constituting receptor trafficking to and from these compartments therefore dictate the efficacy of GABA A R function. Here we review the current understanding of how GABA A Rs traffic through biogenesis, plasma membrane transport, and degradation. Emphasis is placed on discussing novel GABAergic synaptic proteins, receptor and scaffolding post-translational modifications, activity-dependent changes in GABA A R confinement, and neuropeptide and neurosteroid mediated changes. We further highlight modern techniques currently advancing the knowledge of GABA A R trafficking and clinically relevant neurodevelopmental diseases connected to GABAergic dysfunction. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 78: 238-270, 2018. © 2017 Wiley Periodicals, Inc.

  10. Chronic ethanol consumption in rats produces opioid antinociceptive tolerance through inhibition of mu opioid receptor endocytosis.

    Directory of Open Access Journals (Sweden)

    Li He

    Full Text Available It is well known that the mu-opioid receptor (MOR plays an important role in the rewarding properties of ethanol. However, it is less clear how chronic ethanol consumption affects MOR signaling. Here, we demonstrate that rats with prolonged voluntary ethanol consumption develop antinociceptive tolerance to opioids. Signaling through the MOR is controlled at many levels, including via the process of endocytosis. Importantly, agonists at the MOR that promote receptor endocytosis, such as the endogenous peptides enkephalin and β-endorphin, show a reduced propensity to promote antinociceptive tolerance than do agonists, like morphine, which do not promote receptor endocytosis. These observations led us to examine whether chronic ethanol consumption produced opioid tolerance by interfering with MOR endocytosis. Indeed, here we show that chronic ethanol consumption inhibits the endocytosis of MOR in response to opioid peptide. This loss of endocytosis was accompanied by a dramatic decrease in G protein coupled receptor kinase 2 (GRK2 protein levels after chronic drinking, suggesting that loss of this component of the trafficking machinery could be a mechanism by which endocytosis is lost. We also found that MOR coupling to G-protein was decreased in ethanol-drinking rats, providing a functional explanation for loss of opioid antinociception. Together, these results suggest that chronic ethanol drinking alters the ability of MOR to endocytose in response to opioid peptides, and consequently, promotes tolerance to the effects of opioids.

  11. Mactosylceramide Prevents Glial Cell Overgrowth by Inhibiting Insulin and Fibroblast Growth Factor Receptor Signaling

    DEFF Research Database (Denmark)

    Gerdøe-Kristensen, Stine; Lund, Viktor K; Wandall, Hans H

    2017-01-01

    , in which the mannosyltransferase Egghead controls conversion of glucosylceramide (GlcCer) to mactosylceramide (MacCer). Lack of elongated GSL in egghead (egh) mutants causes overgrowth of subperineurial glia (SPG), largely due to aberrant activation of phosphatidylinositol 3-kinase (PI3K). However, to what...... of the Drosophila Insulin Receptor (InR) and the FGFR homolog Heartless (Htl) in wild type SPG, and is suppressed by inhibiting Htl and InR activity in egh. Knockdown of GlcCer synthase in the SPG fails to suppress glial overgrowth in egh nerves, and slightly promotes overgrowth in wild type, suggesting that RTK...... hyperactivation is caused by absence of MacCer and not by GlcCer accumulation. We conclude that an early product in GSL biosynthesis, MacCer, prevents inappropriate activation of Insulin and Fibroblast Growth Factor Receptors in Drosophila glia. This article is protected by copyright. All rights reserved....

  12. A peptide antagonist of the ErbB1 receptor inhibits receptor activation, tumor cell growth and migration in vitro and xenograft tumor growth in vivo

    DEFF Research Database (Denmark)

    Xu, Ruodan; Povlsen, Gro Klitgaard; Soroka, Vladislav

    2010-01-01

    The epidermal growth factor family of receptor tyrosine kinases (ErbBs) plays essential roles in tumorigenesis and cancer disease progression, and therefore has become an attractive target for structure-based drug design. ErbB receptors are activated by ligand-induced homo- and heterodimerization...... constitutes part of the dimerization arm of ErbB3. Inherbin3 binds to the extracellular domains of all four ErbB receptors, with the lowest peptide binding affinity for ErbB4. Inherbin3 functions as an antagonist of epidermal growth factor (EGF)-ErbB1 signaling. We show that Inherbin3 inhibits EGF-induced Erb....... Structural studies have revealed that ErbB receptor dimers are stabilized by receptor-receptor interactions, primarily mediated by a region in the second extracellular domain, termed the "dimerization arm". The present study is the first biological characterization of a peptide, termed Inherbin3, which...

  13. Targeting receptor for advanced glycation end products (RAGE) expression induces apoptosis and inhibits prostate tumor growth

    International Nuclear Information System (INIS)

    Elangovan, Indira; Thirugnanam, Sivasakthivel; Chen, Aoshuang; Zheng, Guoxing; Bosland, Maarten C.; Kajdacsy-Balla, André; Gnanasekar, Munirathinam

    2012-01-01

    Highlights: ► Targeting RAGE by RNAi induces apoptosis in prostate cancer cells. ► Silencing RAGE expression abrogates rHMGB1 mediated cell proliferation. ► Down regulation of RAGE by RNAi inhibits PSA secretion of prostate cancer cells. ► Knock down of RAGE abrogates prostate tumor growth in vivo. ► Disruption of RAGE expression in prostate tumor activates death receptors. -- Abstract: Expression of receptor for advanced glycation end products (RAGE) plays a key role in the progression of prostate cancer. However, the therapeutic potential of targeting RAGE expression in prostate cancer is not yet evaluated. Therefore in this study, we have investigated the effects of silencing the expression of RAGE by RNAi approach both in vitro and in vivo. The results of this study showed that down regulation of RAGE expression by RNAi inhibited the cell proliferation of androgen-dependent (LNCaP) and androgen-independent (DU-145) prostate cancer cells. Furthermore, targeting RAGE expression resulted in apoptotic elimination of these prostate cancer cells by activation of caspase-8 and caspase-3 death signaling. Of note, the levels of prostate specific antigen (PSA) were also reduced in LNCaP cells transfected with RAGE RNAi constructs. Importantly, the RAGE RNAi constructs when administered in nude mice bearing prostate tumors, inhibited the tumor growth by targeting the expression of RAGE, and its physiological ligand, HMGB1 and by up regulating death receptors DR4 and DR5 expression. Collectively, the results of this study for the first time show that targeting RAGE by RNAi may be a promising alternative therapeutic strategy for treating prostate cancer.

  14. Characterization of niphatenones that inhibit androgen receptor N-terminal domain.

    Directory of Open Access Journals (Sweden)

    Carmen A Banuelos

    Full Text Available Androgen ablation therapy causes a temporary reduction in tumor burden in patients with advanced prostate cancer. Unfortunately the malignancy will return to form lethal castration-recurrent prostate cancer (CRPC. The androgen receptor (AR remains transcriptionally active in CRPC in spite of castrate levels of androgens in the blood. AR transcriptional activity resides in its N-terminal domain (NTD. Possible mechanisms of continued AR transcriptional activity may include, at least in part, expression of constitutively active splice variants of AR that lack the C-terminal ligand-binding domain (LBD. Current therapies that target the AR LBD, would not be effective against these AR variants. Currently no drugs are clinically available that target the AR NTD which should be effective against these AR variants as well as full-length AR. Niphatenones were originally isolated and identified in active extracts from Niphates digitalis marine sponge. Here we begin to characterize the mechanism of niphatenones in blocking AR transcriptional activity. Both enantiomers had similar IC50 values of 6 µM for inhibiting the full-length AR in a functional transcriptional assay. However, (S-niphatenone had significantly better activity against the AR NTD compared to (R-niphatenone. Consistent with niphatenones binding to and inhibiting transactivation of AR NTD, niphatenones inhibited AR splice variant. Niphatenone did not affect the transcriptional activity of the related progesterone receptor, but slightly decreased glucocorticoid receptor (GR activity and covalently bound to GR activation function-1 (AF-1 region. Niphatenone blocked N/C interactions of AR without altering either AR protein levels or its intracellular localization in response to androgen. Alkylation with glutathione suggests that niphatenones are not a feasible scaffold for further drug development.

  15. Characterization of Notch1 antibodies that inhibit signaling of both normal and mutated Notch1 receptors.

    Directory of Open Access Journals (Sweden)

    Miguel Aste-Amézaga

    2010-02-01

    Full Text Available Notch receptors normally play a key role in guiding a variety of cell fate decisions during development and differentiation of metazoan organisms. On the other hand, dysregulation of Notch1 signaling is associated with many different types of cancer as well as tumor angiogenesis, making Notch1 a potential therapeutic target.Here we report the in vitro activities of inhibitory Notch1 monoclonal antibodies derived from cell-based and solid-phase screening of a phage display library. Two classes of antibodies were found, one directed against the EGF-repeat region that encompasses the ligand-binding domain (LBD, and the second directed against the activation switch of the receptor, the Notch negative regulatory region (NRR. The antibodies are selective for Notch1, inhibiting Jag2-dependent signaling by Notch1 but not by Notch 2 and 3 in reporter gene assays, with EC(50 values as low as 5+/-3 nM and 0.13+/-0.09 nM for the LBD and NRR antibodies, respectively, and fail to recognize Notch4. While more potent, NRR antibodies are incomplete antagonists of Notch1 signaling. The antagonistic activity of LBD, but not NRR, antibodies is strongly dependent on the activating ligand. Both LBD and NRR antibodies bind to Notch1 on human tumor cell lines and inhibit the expression of sentinel Notch target genes, including HES1, HES5, and DTX1. NRR antibodies also strongly inhibit ligand-independent signaling in heterologous cells transiently expressing Notch1 receptors with diverse NRR "class I" point mutations, the most common type of mutation found in human T-cell acute lymphoblastic leukemia (T-ALL. In contrast, NRR antibodies failed to antagonize Notch1 receptors bearing rare "class II" or "class III" mutations, in which amino acid insertions generate a duplicated or constitutively sensitive metalloprotease cleavage site. Signaling in T-ALL cell lines bearing class I mutations is partially refractory to inhibitory antibodies as compared to cell

  16. Dopamine inhibition of anterior pituitary adenylate cyclase is mediated through the high-affinity state of the D2 receptor

    International Nuclear Information System (INIS)

    Borgundvaag, B.; George, S.R.

    1985-01-01

    The diterpinoid forskolin stimulated adenylate cyclase activity (measured by conversion of [ 3 H]-ATP to [ 3 H]-cAMP) in anterior pituitary from male and female rats. Inhibition of stimulated adenylate cyclase activity by potent dopaminergic agonists was demonstrable only in female anterior pituitary. The inhibition of adenylate cyclase activity displayed a typically dopaminergic rank order of agonist potencies and could be completely reversed by a specific dopamine receptor antagonist. The IC 50 values of dopamine agonist inhibition of adenylate cyclase activity correlated with equal molarity with the dissociation constant of the high-affinity dopamine agonist-detected receptor binding site and with the IC 50 values for inhibition of prolactin secretion. These findings support the hypothesis that it is the high-affinity form of the D 2 dopamine receptor in anterior pituitary which is responsible for mediating the dopaminergic function of attenuating adenylate cyclase activity. 12 references, 4 figures, 1 table

  17. Cobaltous chloride and hypoxia inhibit aryl hydrocarbon receptor-mediated responses in breast cancer cells

    International Nuclear Information System (INIS)

    Khan, Shaheen; Liu Shengxi; Stoner, Matthew; Safe, Stephen

    2007-01-01

    The aryl hydrocarbon receptor (AhR) is expressed in estrogen receptor (ER)-positive ZR-75 breast cancer cells. Treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces CYP1A1 protein and mRNA levels and also activates inhibitory AhR-ERα crosstalk associated with hormone-induced reporter gene expression. In ZR-75 cells grown under hypoxia, induction of these AhR-mediated responses by TCDD was significantly inhibited. This was not accompanied by decreased nuclear AhR levels or decreased interaction of the AhR complex with the CYP1A1 gene promoter as determined in a chromatin immunoprecipitation assay. Hypoxia-induced loss of Ah-responsiveness was not associated with induction of hypoxia-inducible factor-1α or other factors that sequester the AhR nuclear translocation (Arnt) protein, and overexpression of Arnt under hypoxia did not restore Ah-responsiveness. The p65 subunit of NFκB which inhibits AhR-mediated transactivation was not induced by hypoxia and was primarily cytosolic in ZR-75 cells grown under hypoxic and normoxic conditions. In ZR-75 cells maintained under hypoxic conditions for 24 h, BRCA1 (an enhancer of AhR-mediated transactivation in breast cancer cells) was significantly decreased and this contributed to loss of Ah-responsiveness. In cells grown under hypoxia for 6 h, BRCA1 was not decreased, but induction of CYP1A1 by TCDD was significantly decreased. Cotreatment of ZR-75 cells with TCDD plus the protein synthesis inhibitor cycloheximide for 6 h enhanced CYP1A1 expression in cells grown under hypoxia and normoxia. These results suggest that hypoxia rapidly induces protein(s) that inhibit Ah-responsiveness and these may be similar to constitutively expressed inhibitors of Ah-responsiveness (under normoxia) that are also inhibited by cycloheximide

  18. Inhibition of thyroid hormone receptor locally in the retina is a therapeutic strategy for retinal degeneration.

    Science.gov (United States)

    Ma, Hongwei; Yang, Fan; Butler, Michael R; Belcher, Joshua; Redmond, T Michael; Placzek, Andrew T; Scanlan, Thomas S; Ding, Xi-Qin

    2017-08-01

    Thyroid hormone (TH) signaling regulates cell proliferation, differentiation, and metabolism. Recent studies have implicated TH signaling in cone photoreceptor viability. Using mouse models of retinal degeneration, we demonstrated that antithyroid drug treatment and targeting iodothyronine deiodinases (DIOs) to suppress cellular tri-iodothyronine (T3) production or increase T3 degradation preserves cones. In this work, we investigated the effectiveness of inhibition of the TH receptor (TR). Two genes, THRA and THRB , encode TRs; THRB 2 has been associated with cone viability. Using TR antagonists and Thrb2 deletion, we examined the effects of TR inhibition. Systemic and ocular treatment with the TR antagonists NH-3 and 1-850 increased cone density by 30-40% in the Rpe65 -/- mouse model of Leber congenital amaurosis and reduced the number of TUNEL + cells. Cone survival was significantly improved in Rpe65 -/- and Cpfl1 (a model of achromatopsia with Pde6c defect) mice with Thrb2 deletion. Ventral cone density in Cpfl1/Thrb2 -/- and Rpe65 -/- / Thrb2 -/- mice was increased by 1- to 4-fold, compared with age-matched controls. Moreover, the expression levels of TR were significantly higher in the cone-degeneration retinas, suggesting locally elevated TR signaling. This work shows that the effects of antithyroid treatment or targeting DIOs were likely mediated by TRs and that suppressing TR protects cones. Our findings support the view that inhibition of TR locally in the retina is a therapeutic strategy for retinal degeneration management.-Ma, H., Yang, F., Butler, M. R., Belcher, J., Redmond, T. M., Placzek, A. T., Scanlan, T. S., Ding, X.-Q. Inhibition of thyroid hormone receptor locally in the retina is a therapeutic strategy for retinal degeneration. © FASEB.

  19. δ-Tocopherol inhibits receptor tyrosine kinase-induced AKT activation in prostate cancer cells.

    Science.gov (United States)

    Wang, Hong; Hong, Jungil; Yang, Chung S

    2016-11-01

    The cancer preventive activity of vitamin E is suggested by epidemiological studies and supported by animal studies with vitamin E forms, γ-tocopherol and δ-tocopherol (δ-T). Several recent large-scale cancer prevention trials with high dose of α-tocopherol, however, yielded disappointing results. Whether vitamin E prevents or promotes cancer is a serious concern. A better understanding of the molecular mechanisms of action of the different forms of tocopherols would enhance our understanding of this topic. In this study, we demonstrated that δ-T was the most effective tocopherol form in inhibiting prostate cancer cell growth, by inducing cell cycle arrest and apoptosis. By profiling the effects of δ-T on the cell signaling using the phospho-kinase array, we found that the most inhibited target was the phosphorylation of AKT on T308. Further study on the activation of AKT by EGFR and IGFR revealed that δ-T attenuated the EGF/IGF-induced activation of AKT (via the phosphorylation of AKT on T308 induced by the activation of PIK3). Expression of dominant active PIK3 and AKT in prostate cancer cell line DU145 in which PIK3, AKT, and PTEN are wild type caused the cells to be reflectory to the inhibition of δ-T, supporting that δ-T inhibits the PIK3-mediated activation of AKT. Our data also suggest that δ-T interferes with the EGF-induced EGFR internalization, which leads to the inhibition of the receptor tyrosine kinase-dependent activation of AKT. In summary, our results revealed a novel mechanism of δ-T in inhibiting prostate cancer cell growth, supporting the cancer preventive activity δ-T. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

  20. Halothane inhibits the cholinergic-receptor-mediated influx of calcium in primary culture of bovine adrenal medulla cells

    International Nuclear Information System (INIS)

    Yashima, N.; Wada, A.; Izumi, F.

    1986-01-01

    Adrenal medulla cells are cholinoceptive cells. Stimulation of the acetylcholine receptor causes the influx of Ca to the cells, and Ca acts as the coupler of the stimulus-secretion coupling. In this study, the authors investigated the effects of halothane on the receptor-mediated influx of 45 Ca using cultured bovine adrenal medulla cells. Halothane at clinical concentrations (0.5-2%) inhibited the influx of 45 Ca caused by carbachol, with simultaneous inhibition of catecholamine secretion. The influx of 45 Ca and the secretion of catecholamines caused by K depolarization were inhibited by a large concentration of Mg, which competes with Ca at Ca channels, but not inhibited by halothane. Inhibition of the 45 Ca influx by halothane was not overcome by increase in the carbachol concentration. Inhibition of the 45 Ca influx by halothane was examined in comparison with that caused by a large concentration of Mg by the application of Scatchard analysis as the function of the external Ca concentration. Halothane decreased the maximal influx of 45 Ca without altering the apparent kinetic constant of Ca to Ca channels. On the contrary, a large concentration of Mg increased the apparent kinetic constant without altering the maximal influx of 45 Ca. Based on these findings, the authors suggest that inhibition of the 45 Ca influx by halothane was not due to the direct competitive inhibition of Ca channels, nor to the competitive antagonism of agonist-receptor interaction. As a possibility, halothane seems to inhibit the receptor-mediated activation of Ca channels through the interference of coupling between the receptor and Ca channels

  1. The cannabinoid-1 receptor is abundantly expressed in striatal striosomes and striosome-dendron bouquets of the substantia nigra.

    Directory of Open Access Journals (Sweden)

    Margaret I Davis

    Full Text Available Presynaptic cannabinoid-1 receptors (CB1-R bind endogenous and exogenous cannabinoids to modulate neurotransmitter release. CB1-Rs are expressed throughout the basal ganglia, including striatum and substantia nigra, where they play a role in learning and control of motivated actions. However, the pattern of CB1-R expression across different striatal compartments, microcircuits and efferent targets, and the contribution of different CB1-R-expressing neurons to this pattern, are unclear. We use a combination of conventional techniques and novel genetic models to evaluate CB1-R expression in striosome (patch and matrix compartments of the striatum, and in nigral targets of striatal medium spiny projection neurons (MSNs. CB1-R protein and mRNA follow a descending dorsolateral-to-ventromedial intensity gradient in the caudal striatum, with elevated expression in striosomes relative to the surrounding matrix. The lateral predominance of striosome CB1-Rs contrasts with that of the classical striosomal marker, the mu opioid receptor (MOR, which is expressed most prominently in rostromedial striosomes. The dorsolateral-to-ventromedial CB1-R gradient is similar to Drd2 dopamine receptor immunoreactivity and opposite to Substance P. This topology of CB1-R expression is maintained downstream in the globus pallidus and substantia nigra. Dense CB1-R-expressing striatonigral fibers extend dorsally within the substantia nigra pars reticulata, and colocalize with bundles of ventrally extending, striosome-targeted, dendrites of dopamine-containing neurons in the substantia nigra pars compacta (striosome-dendron bouquets. Within striatum, CB1-Rs colocalize with fluorescently labeled MSN collaterals within the striosomes. Cre recombinase-mediated deletion of CB1-Rs from cortical projection neurons or MSNs, and MSN-selective reintroduction of CB1-Rs in knockout mice, demonstrate that the principal source of CB1-Rs in dorsolateral striosomes is local MSN collaterals

  2. Upregulation of Cannabinoid Type 1 Receptors in Dopamine D2 Receptor Knockout Mice Is Reversed by Chronic Forced Ethanol Consumption

    Energy Technology Data Exchange (ETDEWEB)

    Thanos, P.K.; Wang, G.; Thanos, P.K.; Gopez, V.; Delis, F.; Michaelides, M.; Grand, D.K.; Wang, G.-J.; Kunos, G.; Volkow, N.D.

    2011-01-01

    The anatomical proximity of the cannabinoid type 1 (CNR1/CB1R) and the dopamine D2 receptors (DRD2), their ability to form CB1R-DRD2 heteromers, their opposing roles in locomotion, and their involvement in ethanol's reinforcing and addictive properties prompted us to study the levels and distribution of CB1R after chronic ethanol intake, in the presence and absence of DRD2. We monitored the drinking patterns and locomotor activity of Drd2+/+ and Drd2-/- mice consuming either water or a 20% (v/v) ethanol solution (forced ethanol intake) for 6 months and used the selective CB1 receptor antagonist [{sup 3}H]SR141716A to quantify CB1R levels in different brain regions with in vitro receptor autoradiography. We found that the lack of DRD2 leads to a marked upregulation (approximately 2-fold increase) of CB1R in the cerebral cortex, the caudate-putamen, and the nucleus accumbens, which was reversed by chronic ethanol intake. The results suggest that DRD2-mediated dopaminergic neurotransmission and chronic ethanol intake exert an inhibitory effect on cannabinoid receptor expression in cortical and striatal regions implicated in the reinforcing and addictive properties of ethanol.

  3. Dual angiotensin receptor and neprilysin inhibition as an alternative to angiotensin-converting enzyme inhibition in patients with chronic systolic heart failure

    DEFF Research Database (Denmark)

    McMurray, John J V; Packer, Milton; Desai, Akshay S

    2013-01-01

    and natriuresis, inhibit abnormal growth, suppress the RAAS and sympathetic nervous system, and augment parasympathetic activity. The best understood of these mediators are the natriuretic peptides which are metabolized by the enzyme neprilysin. LCZ696 belongs to a new class of drugs, the angiotensin receptor...

  4. Inhibition of the CSF-1 receptor sensitizes ovarian cancer cells to cisplatin.

    Science.gov (United States)

    Yu, Rong; Jin, Hao; Jin, Congcong; Huang, Xuefeng; Lin, Jinju; Teng, Yili

    2018-03-01

    Ovarian cancer is one of the most common female malignancies, and cisplatin-based chemotherapy is routinely used in locally advanced ovarian cancer patients. Acquired or de novo cisplatin resistance remains the barrier to patient survival, and the mechanisms of cisplatin resistance are still not well understood. In the current study, we found that colony-stimulating-factor-1 receptor (CSF-1R) was upregulated in cisplatin-resistant SK-OV-3 and CaoV-3 cells. Colony-stimulating-factor-1 receptor knockdown suppressed proliferation and enhanced apoptosis in cisplatin-resistant SK-OV-3 and CaoV-3 cells. However, CSF-1R overexpression had inverse effects. While parental SK-OV-3 and CaoV-3 cells were more resistant to cisplatin after CSF-1R overexpression, CSF-1R knockdown in SK-OV-3 and CaoV-3 cells promoted cisplatin sensitivity. Overexpression and knockdown studies also showed that CSF-1R significantly promoted active AKT and ERK1/2 signalling pathways in cisplatin-resistant cells. Furthermore, a combination of cisplatin and CSF-1R inhibitor effectively inhibited tumour growth in xenografts. Taken together, our results provide the first evidence that CSF-1R inhibition can sensitize cisplatin-refractory ovarian cancer cells. This study may help to increase understanding of the molecular mechanisms underlying cisplatin resistance in tumours. Copyright © 2018 John Wiley & Sons, Ltd.

  5. Inhibition of androgen receptor by decoy molecules delays progression to castration-recurrent prostate cancer.

    Directory of Open Access Journals (Sweden)

    Jae-Kyung Myung

    Full Text Available Androgen receptor (AR is a member of the steroid receptor family and a therapeutic target for all stages of prostate cancer. AR is activated by ligand binding within its C-terminus ligand-binding domain (LBD. Here we show that overexpression of the AR NTD to generate decoy molecules inhibited both the growth and progression of prostate cancer in castrated hosts. Specifically, it was shown that lentivirus delivery of decoys delayed hormonal progression in castrated hosts as indicated by increased doubling time of tumor volume, prolonged time to achieve pre-castrate levels of serum prostate-specific antigen (PSA and PSA nadir. These clinical parameters are indicative of delayed hormonal progression and improved therapeutic response and prognosis. Decoys reduced the expression of androgen-regulated genes that correlated with reduced in situ interaction of the AR with androgen response elements. Decoys did not reduce levels of AR protein or prevent nuclear localization of the AR. Nor did decoys interact directly with the AR. Thus decoys did not inhibit AR transactivation by a dominant negative mechanism. This work provides evidence that the AR NTD plays an important role in the hormonal progression of prostate cancer and supports the development of AR antagonists that target the AR NTD.

  6. Fisetin inhibits liver cancer growth in a mouse model: Relation to dopamine receptor.

    Science.gov (United States)

    Liu, Xiang-Feng; Long, Hai-Jiao; Miao, Xiong-Ying; Liu, Guo-Li; Yao, Hong-Liang

    2017-07-01

    Fisetin (3,3',4',7-tetrahydroxyflavone), a natural abundant flavonoid, is produced in different vegetables and fruits. Fisetin has been reported to relate to various positive biological effects, including anti-proliferative, anticancer, anti-oxidative and neuroprotective effects. Dopamine receptors (DRs) belonging to G protein‑coupled receptor family, are known as the target of ~50% of all modern medicinal drugs. DRs consist of various proteins, functioning as transduction of intracellular signals for extracellular stimuli. We found that fisetin performed as DR2 agonist to suppress liver cancer cells proliferation, migration and invasion. Caspase-3 signaling was activated to induce apoptosis for fisetin administration. Furthermore, TGF‑β1 was also inhibited in fisetin-treated liver cancer cells, reducing epithelial-mesenchymal transition (EMT). Additionally, fisetin downregulated VEGFR1, p-ERK1/2, p38 and pJNK, ameliorating liver cancer progression. In vivo, the orthotopically implanted tumors from mice were inhibited by fisetin adminisatration accompanied by prolonged survival rate and higher levels of dopamine. Together, the results indicated a novel therapeutic strategy to suppress liver cancer progression associated with DR2 regulation, indicating that dopamine might be of importance in liver cancer progression.

  7. Inhibition of fibroblast growth factor receptor 3-dependent lung adenocarcinoma with a human monoclonal antibody

    Directory of Open Access Journals (Sweden)

    Yongjun Yin

    2016-05-01

    Full Text Available Activating mutations in fibroblast growth factor receptor 3 (FGFR3 have been identified in multiple types of human cancer and in congenital birth defects. In human lung cancer, fibroblast growth factor 9 (FGF9, a high-affinity ligand for FGFR3, is overexpressed in 10% of primary resected non-small cell lung cancer (NSCLC specimens. Furthermore, in a mouse model where FGF9 can be induced in lung epithelial cells, epithelial proliferation and ensuing tumorigenesis is dependent on FGFR3. To develop new customized therapies for cancers that are dependent on FGFR3 activation, we have used this mouse model to evaluate a human monoclonal antibody (D11 with specificity for the extracellular ligand-binding domain of FGFR3, that recognizes both human and mouse forms of the receptor. Here, we show that D11 effectively inhibits signaling through FGFR3 in vitro, inhibits the growth of FGFR3-dependent FGF9-induced lung adenocarcinoma in mice, and reduces tumor-associated morbidity. Given the potency of FGF9 in this mouse model and the absolute requirement for signaling through FGFR3, this study validates the D11 antibody as a potentially useful and effective reagent for treating human cancers or other pathologies that are dependent on activation of FGFR3.

  8. STC1 interference on calcitonin family of receptors signaling during osteoblastogenesis via adenylate cyclase inhibition.

    Science.gov (United States)

    Terra, Silvia R; Cardoso, João Carlos R; Félix, Rute C; Martins, Leo Anderson M; Souza, Diogo Onofre G; Guma, Fatima C R; Canário, Adelino Vicente M; Schein, Vanessa

    2015-03-05

    Stanniocalcin 1 (STC1) and calcitonin gene-related peptide (CGRP) are involved in bone formation/remodeling. Here we investigate the effects of STC1 on functional heterodimer complex CALCRL/RAMP1, expression and activity during osteoblastogenesis. STC1 did not modify CALCRL and ramp1 gene expression during osteoblastogenesis when compared to controls. However, plasma membrane spatial distribution of CALCRL/RAMP1 was modified in 7-day pre-osteoblasts exposed to either CGRP or STC1, and both peptides induced CALCRL and RAMP1 assembly. CGRP, but not STC1 stimulated cAMP accumulation in 7-day osteoblasts and in CALCRL/RAMP1 transfected HEK293 cells. Furthermore, STC1 inhibited forskolin stimulated cAMP accumulation of HEK293 cells, but not in CALCRL/RAMP1 transfected HEK293 cells. However, STC1 inhibited cAMP accumulation in calcitonin receptor (CTR) HEK293 transfected cells stimulated by calcitonin. In conclusion, STC1 signals through inhibitory G-protein modulates CGRP receptor spatial localization during osteoblastogenesis and may function as a regulatory factor interacting with calcitonin peptide members during bone formation. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  9. [Expression of cannabinoid receptor I during mice skin incised wound healing course].

    Science.gov (United States)

    Zhao, Zhen-bin; Guan, Da-wei; Liu, Wei-wei; Wang, Tao; Fan, Yan-yan; Cheng, Zi-hui; Zheng, Ji-long; Hu, Geng-yi

    2010-08-01

    To investigate the expression of cannabinoid receptor I (CB1R) during mice skin incised wound healing course and time-dependent changes of CB1R in wound age determination. The changes of CBIR expression in skin incised wound were detected by immunohistochemistry and Western blotting. The control group showed a low expression of CB1R detected mainly in epidermis, hair follicles, sebaceous gland and dermomuscular layer. CB1R expression was undetectable in neutrophils in the wound specimens from 6h to 12h post-injury. CB1R positive cells were mostly mononuclear cells (MNCs) and fibroblastic cells (FBCs) from 1 d to 5 d post-injury. CB1R positive cells were mostly FBCs from 7 d to 14d post-injury. The ratio of the CB1R positive cells increased gradually in the wound specimens from 6 h to 3 d post-injury, reached peak level at 5 d, and then decreased gradually from 7d to 14 d post-injury. The positive bands of CB1R were observed in all time points of the wound healing course by Western blotting. The expression peak showed at 5 d post-injury. CB1R is activated during the wound healing course. The expression of CB1R is found in mononuclear cells, which could be involved in inflammation reaction. CBIR is observed in fibroblastic cells, which could participate in the wound healing. CB1R may be a potentially useful marker for determination of wound healing age.

  10. Small Molecules from Nature Targeting G-Protein Coupled Cannabinoid Receptors: Potential Leads for Drug Discovery and Development

    Directory of Open Access Journals (Sweden)

    Charu Sharma

    2015-01-01

    Full Text Available The cannabinoid molecules are derived from Cannabis sativa plant which acts on the cannabinoid receptors types 1 and 2 (CB1 and CB2 which have been explored as potential therapeutic targets for drug discovery and development. Currently, there are numerous cannabinoid based synthetic drugs used in clinical practice like the popular ones such as nabilone, dronabinol, and Δ9-tetrahydrocannabinol mediates its action through CB1/CB2 receptors. However, these synthetic based Cannabis derived compounds are known to exert adverse psychiatric effect and have also been exploited for drug abuse. This encourages us to find out an alternative and safe drug with the least psychiatric adverse effects. In recent years, many phytocannabinoids have been isolated from plants other than Cannabis. Several studies have shown that these phytocannabinoids show affinity, potency, selectivity, and efficacy towards cannabinoid receptors and inhibit endocannabinoid metabolizing enzymes, thus reducing hyperactivity of endocannabinoid systems. Also, these naturally derived molecules possess the least adverse effects opposed to the synthetically derived cannabinoids. Therefore, the plant based cannabinoid molecules proved to be promising and emerging therapeutic alternative. The present review provides an overview of therapeutic potential of ligands and plants modulating cannabinoid receptors that may be of interest to pharmaceutical industry in search of new and safer drug discovery and development for future therapeutics.

  11. Lactobacillus bulgaricus OLL1181 activates the aryl hydrocarbon receptor pathway and inhibits colitis

    Science.gov (United States)

    Takamura, Takeyuki; Harama, Daisuke; Fukumoto, Suguru; Nakamura, Yuki; Shimokawa, Naomi; Ishimaru, Kayoko; Ikegami, Shuji; Makino, Seiya; Kitamura, Masanori; Nakao, Atsuhito

    2011-01-01

    Increasing evidence suggests that the aryl hydrocarbon receptor (AhR) pathway has an important role in the regulation of inflammatory responses. Most recently, we have shown that the activation of the AhR pathway by a potent AhR agonist inhibits the development of dextran sodium sulfate (DSS)-induced colitis, a model of human ulcerative colitis, by the induction of prostaglandin E2 (PGE2) in the large intestine. Because several strains of probiotic lactic acid bacteria have been reported to inhibit DSS-induced colitis by unidentified mechanisms, we hypothesized that particular strains of lactic acid bacterium might have the potential to activate the AhR pathway, thereby inhibiting DSS-induced colitis. This study investigated whether there are specific lactic acid bacterial strains that can activate the AhR pathway, and if so, whether this AhR-activating potential is associated with suppression of DSS-induced colitis. By using AhR signaling reporter cells, we found that Lactobacillus bulgaricus OLL1181 had the potential to activate the AhR pathway. OLL1181 also induced the mRNA expression of cytochrome P450 family 1A1 (CYP1A1), a target gene of the AhR pathway, in human colon cells, which was inhibited by the addition of an AhR antagonist, α-naphthoflavon (αNF). In addition, mice treated orally with OLL1181 showed an increase in CYP1A1 mRNA expression in the large intestine and amelioration of DSS-induced colitis. Thus, OLL1181 can induce activation of the intestinal AhR pathway and inhibit DSS-induced colitis in mice. This strain of lactic acid bacterium has therefore the potential to activate the AhR pathway, which may be able to suppress colitis. PMID:21321579

  12. Purinergic receptor antagonists inhibit odorant-mediated CREB phosphorylation in sustentacular cells of mouse olfactory epithelium.

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2012-02-01

    BACKGROUND: Extracellular nucleotides have long been known to play neuromodulatory roles and to be involved in intercellular signalling. In the olfactory system, ATP is released by olfactory neurons, and exogenous ATP can evoke an increase in intracellular calcium concentration in sustentacular cells, the nonneuronal supporting cells of the olfactory epithelium. Here we investigate the hypothesis that olfactory neurons communicate with sustentacular cells via extracellular ATP and purinergic receptor activation. RESULTS: Here we show that exposure of mice to a mixture of odorants induced a significant increase in the levels of the transcription factor CREB phosphorylated at Ser-133 in the nuclei of both olfactory sensory neurons and sustentacular cells. This activation was dependent on adenylyl cyclase III-mediated olfactory signaling and on activation of P2Y purinergic receptors on sustentacular cells. Purinergic receptor antagonists inhibited odorant-dependent CREB phosphorylation specifically in the nuclei of the sustentacular cells. CONCLUSION: Our results point to a possible role for extracellular nucleotides in mediating intercellular communication between the neurons and sustentacular cells of the olfactory epithelium in response to odorant exposure. Maintenance of extracellular ionic gradients and metabolism of noxious chemicals by sustentacular cells may therefore be regulated in an odorant-dependent manner by olfactory sensory neurons.

  13. Purinergic receptor antagonists inhibit odorant-mediated CREB phosphorylation in sustentacular cells of mouse olfactory epithelium

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2011-08-22

    Abstract Background Extracellular nucleotides have long been known to play neuromodulatory roles and to be involved in intercellular signalling. In the olfactory system, ATP is released by olfactory neurons, and exogenous ATP can evoke an increase in intracellular calcium concentration in sustentacular cells, the nonneuronal supporting cells of the olfactory epithelium. Here we investigate the hypothesis that olfactory neurons communicate with sustentacular cells via extracellular ATP and purinergic receptor activation. Results Here we show that exposure of mice to a mixture of odorants induced a significant increase in the levels of the transcription factor CREB phosphorylated at Ser-133 in the nuclei of both olfactory sensory neurons and sustentacular cells. This activation was dependent on adenylyl cyclase III-mediated olfactory signaling and on activation of P2Y purinergic receptors on sustentacular cells. Purinergic receptor antagonists inhibited odorant-dependent CREB phosphorylation specifically in the nuclei of the sustentacular cells. Conclusion Our results point to a possible role for extracellular nucleotides in mediating intercellular communication between the neurons and sustentacular cells of the olfactory epithelium in response to odorant exposure. Maintenance of extracellular ionic gradients and metabolism of noxious chemicals by sustentacular cells may therefore be regulated in an odorant-dependent manner by olfactory sensory neurons.

  14. Phosphorylation inhibits DNA-binding of alternatively spliced aryl hydrocarbon receptor nuclear translocator

    International Nuclear Information System (INIS)

    Kewley, Robyn J.; Whitelaw, Murray L.

    2005-01-01

    The basic helix-loop-helix/PER-ARNT-SIM homology (bHLH/PAS) transcription factor ARNT (aryl hydrocarbon receptor nuclear translocator) is a key component of various pathways which induce the transcription of cytochrome P450 and hypoxia response genes. ARNT can be alternatively spliced to express Alt ARNT, containing an additional 15 amino acids immediately N-terminal to the DNA-binding basic region. Here, we show that ARNT and Alt ARNT proteins are differentially phosphorylated by protein kinase CKII in vitro. Phosphorylation had an inhibitory effect on DNA-binding to an E-box probe by Alt ARNT, but not ARNT, homodimers. This inhibitory phosphorylation occurs through Ser77. Moreover, a point mutant, Alt ARNT S77A, shows increased activity on an E-box reporter gene, consistent with Ser77 being a regulatory site in vivo. In contrast, DNA binding by an Alt ARNT/dioxin receptor heterodimer to the xenobiotic response element is not inhibited by phosphorylation with CKII, nor does Alt ARNT S77A behave differently from wild type Alt ARNT in the context of a dioxin receptor heterodimer

  15. Purinergic receptor antagonists inhibit odorant-mediated CREB phosphorylation in sustentacular cells of mouse olfactory epithelium

    Directory of Open Access Journals (Sweden)

    Hatt Hanns

    2011-08-01

    Full Text Available Abstract Background Extracellular nucleotides have long been known to play neuromodulatory roles and to be involved in intercellular signalling. In the olfactory system, ATP is released by olfactory neurons, and exogenous ATP can evoke an increase in intracellular calcium concentration in sustentacular cells, the nonneuronal supporting cells of the olfactory epithelium. Here we investigate the hypothesis that olfactory neurons communicate with sustentacular cells via extracellular ATP and purinergic receptor activation. Results Here we show that exposure of mice to a mixture of odorants induced a significant increase in the levels of the transcription factor CREB phosphorylated at Ser-133 in the nuclei of both olfactory sensory neurons and sustentacular cells. This activation was dependent on adenylyl cyclase III-mediated olfactory signaling and on activation of P2Y purinergic receptors on sustentacular cells. Purinergic receptor antagonists inhibited odorant-dependent CREB phosphorylation specifically in the nuclei of the sustentacular cells. Conclusion Our results point to a possible role for extracellular nucleotides in mediating intercellular communication between the neurons and sustentacular cells of the olfactory epithelium in response to odorant exposure. Maintenance of extracellular ionic gradients and metabolism of noxious chemicals by sustentacular cells may therefore be regulated in an odorant-dependent manner by olfactory sensory neurons.

  16. Diurnal inhibition of NMDA-EPSCs at rat hippocampal mossy fibre synapses through orexin-2 receptors

    Science.gov (United States)

    Perin, Martina; Longordo, Fabio; Massonnet, Christine; Welker, Egbert; Lüthi, Anita

    2014-01-01

    Diurnal release of the orexin neuropeptides orexin-A (Ox-A, hypocretin-1) and orexin-B (Ox-B, hypocretin-2) stabilises arousal, regulates energy homeostasis and contributes to cognition and learning. However, whether cellular correlates of brain plasticity are regulated through orexins, and whether they do so in a time-of-day-dependent manner, has never been assessed. Immunohistochemically we found sparse but widespread innervation of hippocampal subfields through Ox-A- and Ox-B-containing fibres in young adult rats. The actions of Ox-A were studied on NMDA receptor (NMDAR)-mediated excitatory synaptic transmission in acute hippocampal slices prepared around the trough (Zeitgeber time (ZT) 4–8, corresponding to 4–8 h into the resting phase) and peak (ZT 23) of intracerebroventricular orexin levels. At ZT 4–8, exogenous Ox-A (100 nm in bath) inhibited NMDA receptor-mediated excitatory postsynaptic currents (NMDA-EPSCs) at mossy fibre (MF)–CA3 (to 55.6 ± 6.8% of control, P = 0.0003) and at Schaffer collateral–CA1 synapses (70.8 ± 6.3%, P = 0.013), whereas it remained ineffective at non-MF excitatory synapses in CA3. Ox-A actions were mediated postsynaptically and blocked by the orexin-2 receptor (OX2R) antagonist JNJ10397049 (1 μm), but not by orexin-1 receptor inhibition (SB334867, 1 μm) or by adrenergic and cholinergic antagonists. At ZT 23, inhibitory effects of exogenous Ox-A were absent (97.6 ± 2.9%, P = 0.42), but reinstated (87.2 ± 3.3%, P = 0.002) when endogenous orexin signalling was attenuated for 5 h through i.p. injections of almorexant (100 mg kg−1), a dual orexin receptor antagonist. In conclusion, endogenous orexins modulate hippocampal NMDAR function in a time-of-day-dependent manner, suggesting that they may influence cellular plasticity and consequent variations in memory performance across the sleep–wake cycle. PMID:25085886

  17. Cannabidiol restores intestinal barrier dysfunction and inhibits the apoptotic process induced by Clostridium difficile toxin A in Caco-2 cells.

    Science.gov (United States)

    Gigli, Stefano; Seguella, Luisa; Pesce, Marcella; Bruzzese, Eugenia; D'Alessandro, Alessandra; Cuomo, Rosario; Steardo, Luca; Sarnelli, Giovanni; Esposito, Giuseppe

    2017-12-01

    Clostridium difficile toxin A is responsible for colonic damage observed in infected patients. Drugs able to restore Clostridium difficile toxin A-induced toxicity have the potential to improve the recovery of infected patients. Cannabidiol is a non-psychotropic component of Cannabis sativa, which has been demonstrated to protect enterocytes against chemical and/or inflammatory damage and to restore intestinal mucosa integrity. The purpose of this study was to evaluate (a) the anti-apoptotic effect and (b) the mechanisms by which cannabidiol protects mucosal integrity in Caco-2 cells exposed to Clostridium difficile toxin A. Caco-2 cells were exposed to Clostridium difficile toxin A (30 ng/ml), with or without cannabidiol (10 -7 -10 -9  M), in the presence of the specific antagonist AM251 (10 -7  M). Cytotoxicity assay, transepithelial electrical resistence measurements, immunofluorescence analysis and immunoblot analysis were performed in the different experimental conditions. Clostridium difficile toxin A significantly decreased Caco-2 cells' viability and reduced transepithelial electrical resistence values and RhoA guanosine triphosphate (GTP), bax, zonula occludens-1 and occludin protein expression, respectively. All these effects were significantly and concentration-dependently inhibited by cannabidiol, whose effects were completely abolished in the presence of the cannabinoid receptor type 1 (CB1) antagonist, AM251. Cannabidiol improved Clostridium difficile toxin A-induced damage in Caco-2 cells, by inhibiting the apoptotic process and restoring the intestinal barrier integrity, through the involvement of the CB1 receptor.

  18. PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

    Science.gov (United States)

    Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang

    2015-02-01

    The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

  19. Acute inhibition of selected membrane-proximal mouse T cell receptor signaling by mitochondrial antagonists.

    Directory of Open Access Journals (Sweden)

    Kwangmi Kim

    2009-11-01

    Full Text Available T cells absorb nanometric membrane vesicles, prepared from plasma membrane of antigen presenting cells, via dual receptor/ligand interactions of T cell receptor (TCR with cognate peptide/major histocompatibility complex (MHC plus lymphocyte function-associated antigen 1 (LFA-1 with intercellular adhesion molecule 1. TCR-mediated signaling for LFA-1 activation is also required for the vesicle absorption. Exploiting those findings, we had established a high throughput screening (HTS platform and screened a library for isolation of small molecules inhibiting the vesicle absorption. Follow-up studies confirmed that treatments (1 hour with various mitochondrial antagonists, including a class of anti-diabetic drugs (i.e., Metformin and Phenformin, resulted in ubiquitous inhibition of the vesicle absorption without compromising viability of T cells. Further studies revealed that the mitochondrial drug treatments caused impairment of specific membrane-proximal TCR signaling event(s. Thus, activation of Akt and PLC-gamma1 and entry of extracellular Ca(2+ following TCR stimulation were attenuated while polymerization of monomeric actins upon TCR triggering progressed normally after the treatments. Dynamic F-actin rearrangement concurring with the vesicle absorption was also found to be impaired by the drug treatments, implying that the inhibition by the drug treatments of downstream signaling events (and the vesicle absorption could result from lack of directional relocation of signaling and cell surface molecules. We also assessed the potential application of mitochondrial antagonists as immune modulators by probing effects of the long-term drug treatments (24 hours on viability of resting primary T cells and cell cycle progression of antigen-stimulated T cells. This study unveils a novel regulatory mechanism for T cell immunity in response to environmental factors having effects on mitochondrial function.

  20. Inhibition of Protease-activated Receptor 1 Ameliorates Intestinal Radiation Mucositis in a Preclinical Rat Model

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Junru; Kulkarni, Ashwini [Division of Radiation Health, University of Arkansas for Medical Sciences, Little Rock, Arkansas (United States); Chintala, Madhu [Schering-Plough Research Institute, Kenilworth, New Jersey (United States); Fink, Louis M. [Nevada Cancer Institute, Las Vegas, Nevada (United States); Hauer-Jensen, Martin, E-mail: mhjensen@life.uams.edu [Division of Radiation Health, University of Arkansas for Medical Sciences, Little Rock, Arkansas (United States); Surgery Service, Central Arkansas Veterans Healthcare System, Little Rock, Arkansas (United States)

    2013-01-01

    Purpose: To determine, using a specific small-molecule inhibitor of protease-activated receptor 1 (PAR1) signaling, whether the beneficial effect of thrombin inhibition on radiation enteropathy development is due to inhibition of blood clotting or to cellular (PAR1-mediated) thrombin effects. Methods and Materials: Rats underwent fractionated X-irradiation (5 Gy Multiplication-Sign 9) of a 4-cm small-bowel segment. Early radiation toxicity was evaluated in rats receiving PAR1 inhibitor (SCH602539, 0, 10, or 15 mg/kg/d) from 1 day before to 2 weeks after the end of irradiation. The effect of PAR1 inhibition on development of chronic intestinal radiation fibrosis was evaluated in animals receiving SCH602539 (0, 15, or 30 mg/kg/d) until 2 weeks after irradiation, or continuously until termination of the experiment 26 weeks after irradiation. Results: Blockade of PAR1 ameliorated early intestinal toxicity, with reduced overall intestinal radiation injury (P=.002), number of myeloperoxidase-positive (P=.03) and proliferating cell nuclear antigen-positive (P=.04) cells, and collagen III accumulation (P=.005). In contrast, there was no difference in delayed radiation enteropathy in either the 2- or 26-week administration groups. Conclusion: Pharmacological blockade of PAR1 seems to reduce early radiation mucositis but does not affect the level of delayed intestinal radiation fibrosis. Early radiation enteropathy is related to activation of cellular thrombin receptors, whereas platelet activation or fibrin formation may play a greater role in the development of delayed toxicity. Because of the favorable side-effect profile, PAR1 blockade should be further explored as a method to ameliorate acute intestinal radiation toxicity in patients undergoing radiotherapy for cancer and to protect first responders and rescue personnel in radiologic/nuclear emergencies.

  1. Adrenergic receptors inhibit TRPV1 activity in the dorsal root ganglion neurons of rats.

    Science.gov (United States)

    Matsushita, Yumi; Manabe, Miki; Kitamura, Naoki; Shibuya, Izumi

    2018-01-01

    Transient receptor potential vanilloid type 1 (TRPV1) is a polymodal receptor channel that responds to multiple types of stimuli, such as heat, acid, mechanical pressure and some vanilloids. Capsaicin is the most commonly used vanilloid to stimulate TRPV1. TRPV1 channels are expressed in dorsal root ganglion neurons that extend to Aδ- and C-fibers and have a role in the transduction of noxious inputs to the skin into the electrical signals of the sensory nerve. Although noradrenergic nervous systems, including the descending antinociceptive system and the sympathetic nervous system, are known to modulate pain sensation, the functional association between TRPV1 and noradrenaline in primary sensory neurons has rarely been examined. In the present study, we examined the effects of noradrenaline on capsaicin-evoked currents in cultured dorsal root ganglion neurons of the rat by the whole-cell voltage clamp method. Noradrenaline at concentrations higher than 0.1 pM significantly reduced the amplitudes of the inward capsaicin currents recorded at -60 mV holding potential. This inhibitory action was reversed by either yohimbine (an α2 antagonist, 10 nM) or propranolol (a β antagonist, 10 nM). The α2 agonists, clonidine (1 pM) and dexmedetomidine (1 pM) inhibited capsaicin currents, and yohimbine (1 nM) reversed the effects of clonidine. The inhibitory action of noradrenaline was not seen in the neurons pretreated with pertussis toxin (100 μg/ml for 24 h) and the neurons dialyzed intracellularly with guanosine 5'- [β-thio] diphosphate (GDPβS, 200 μM), the catalytic subunit of protein kinase A (250 U/ml) or okadaic acid (1 μM). These results suggest that noradrenaline directly acts on dorsal root ganglion neurons to inhibit the activity of TRPV1 depending on the activation of α2-adrenoceptors followed by the inhibition of the adenylate cyclase/cAMP/protein kinase A pathway.

  2. Combined inhibition of monoacylglycerol lipase and cyclooxygenases synergistically reduces neuropathic pain in mice

    Science.gov (United States)

    Crowe, Molly S; Leishman, Emma; Banks, Matthew L; Gujjar, Ramesh; Mahadevan, Anu; Bradshaw, Heather B; Kinsey, Steven G

    2015-01-01

    Background and Purpose Neuropathic pain is commonly treated with GABA analogues, steroids or non-steroidal anti-inflammatory drugs (NSAIDs). NSAIDs inhibit one or more COX isozymes but chronic COX inhibition paradoxically increases gastrointestinal inflammation and risk of unwanted cardiovascular events. The cannabinoids also have analgesic and anti-inflammatory properties and reduce neuropathic pain in animal models. The present study investigated the analgesic effects of inhibiting both monoacylglycerol lipase (MAGL) and COX enzymes, using low doses of both inhibitors. Experimental Approach Mice subjected to chronic constriction injury (CCI) were tested for mechanical and cold allodynia after administration of the MAGL inhibitor, JZL184, or the non-selective COX inhibitor diclofenac. Then, both drugs were co-administered at fixed dose proportions of 1:3, 1:1 and 3:1, based on their ED50 values. PGs, endocannabinoids and related lipids were quantified in lumbar spinal cord. Key Results Combining low doses of JZL184 and diclofenac synergistically attenuated mechanical allodynia and additively reduced cold allodynia. The cannabinoid CB1 receptor antagonist, rimonabant, but not the CB2 receptor antagonist, SR144528, blocked the analgesic effects of the JZL184 and diclofenac combination on mechanical allodynia, implying that CB1 receptors were primarily responsible for the anti-allodynia. Diclofenac alone and with JZL184 significantly reduced PGE2 and PGF2α in lumbar spinal cord tissue, whereas JZL184 alone caused significant increases in the endocannabinoid metabolite, N-arachidonoyl glycine. Conclusions and Implications Combining COX and MAGL inhibition is a promising therapeutic approach for reducing neuropathic pain with minimal side effects. PMID:25393148

  3. Optimization of cAMP fluorescence dataset from ACTOne cannabinoid receptor 1 cell line

    Directory of Open Access Journals (Sweden)

    Chaela S. Presley

    2016-06-01

    Full Text Available The ACTOne cannabinoid receptor 1 functional system is comprised of transfected HEK cells with the parental cyclic nucleotide gated channel (CNG co-transfected with cannabinoid receptor 1 (CB1. The ACTOne CB1 cell line was evaluated for cAMP driven fluorescence by optimizing experimental conditions for sensitivity to forskolin and CP 55,940, reading time point, reliability of cell passage number, and pertussis inactivation of Gi/o.

  4. Inhibition of the Receptor Tyrosine Kinase AXL Restores Paclitaxel Chemosensitivity in Uterine Serous Cancer.

    Science.gov (United States)

    Palisoul, Marguerite L; Quinn, Jeanne M; Schepers, Emily; Hagemann, Ian S; Guo, Lei; Reger, Kelsey; Hagemann, Andrea R; McCourt, Carolyn K; Thaker, Premal H; Powell, Matthew A; Mutch, David G; Fuh, Katherine C

    2017-12-01

    Uterine serous cancer (USC) is aggressive, and the majority of recurrent cases are chemoresistant. Because the receptor tyrosine kinase AXL promotes invasion and metastasis of USC and is implicated in chemoresistance in other cancers, we assessed the role of AXL in paclitaxel resistance in USC, determined the mechanism of action, and sought to restore chemosensitivity by inhibiting AXL in vitro and in vivo We used short hairpin RNAs and BGB324 to knock down and inhibit AXL. We assessed sensitivity of USC cell lines to paclitaxel and measured paclitaxel intracellular accumulation in vitro in the presence or absence of AXL. We also examined the role of the epithelial-mesenchymal transition (EMT) in AXL-mediated paclitaxel resistance. Finally, we treated USC xenografts with paclitaxel, BGB324, or paclitaxel plus BGB324 and monitored tumor burden. AXL expression was higher in chemoresistant USC patient tumors and cell lines than in chemosensitive tumors and cell lines. Knockdown or inhibition of AXL increased sensitivity of USC cell lines to paclitaxel in vitro and increased cellular accumulation of paclitaxel. AXL promoted chemoresistance even in cells that underwent the EMT in vitro Finally, in vivo studies of combination treatment with BGB324 and paclitaxel showed a greater than 51% decrease in tumor volume after 2 weeks of treatment when compared with no treatment or single-agent treatments ( P USC. Mol Cancer Ther; 16(12); 2881-91. ©2017 AACR . ©2017 American Association for Cancer Research.

  5. Flavonoids Suppress Pseudomonas aeruginosa Virulence through Allosteric Inhibition of Quorum-sensing Receptors*

    Science.gov (United States)

    Paczkowski, Jon E.; Mukherjee, Sampriti; McCready, Amelia R.; Cong, Jian-Ping; Aquino, Christopher J.; Kim, Hahn; Henke, Brad R.; Smith, Chari D.; Bassler, Bonnie L.

    2017-01-01

    Quorum sensing is a process of cell-cell communication that bacteria use to regulate collective behaviors. Quorum sensing depends on the production, detection, and group-wide response to extracellular signal molecules called autoinducers. In many bacterial species, quorum sensing controls virulence factor production. Thus, disrupting quorum sensing is considered a promising strategy to combat bacterial pathogenicity. Several members of a family of naturally produced plant metabolites called flavonoids inhibit Pseudomonas aeruginosa biofilm formation by an unknown mechanism. Here, we explore this family of molecules further, and we demonstrate that flavonoids specifically inhibit quorum sensing via antagonism of the autoinducer-binding receptors, LasR and RhlR. Structure-activity relationship analyses demonstrate that the presence of two hydroxyl moieties in the flavone A-ring backbone are essential for potent inhibition of LasR/RhlR. Biochemical analyses reveal that the flavonoids function non-competitively to prevent LasR/RhlR DNA binding. Administration of the flavonoids to P. aeruginosa alters transcription of quorum sensing-controlled target promoters and suppresses virulence factor production, confirming their potential as anti-infectives that do not function by traditional bacteriocidal or bacteriostatic mechanisms. PMID:28119451

  6. Pax6 represses androgen receptor-mediated transactivation by inhibiting recruitment of the coactivator SPBP.

    Directory of Open Access Journals (Sweden)

    Julianne Elvenes

    Full Text Available The androgen receptor (AR has a central role in development and maintenance of the male reproductive system and in the etiology of prostate cancer. The transcription factor Pax6 has recently been reported to act as a repressor of AR and to be hypermethylated in prostate cancer cells. SPBP is a transcriptional regulator that previously has been shown to enhance the activity of Pax6. In this study we have identified SPBP to act as a transcriptional coactivator of AR. We also show that Pax6 inhibits SPBP-mediated enhancement of AR activity on the AR target gene probasin promoter, a repression that was partly reversed by increased expression of SPBP. Enhanced expression of Pax6 reduced the amount of SPBP associated with the probasin promoter when assayed by ChIP in HeLa cells. We mapped the interaction between both AR and SPBP, and AR and Pax6 to the DNA-binding domains of the involved proteins. Further binding studies revealed that Pax6 and SPBP compete for binding to AR. These results suggest that Pax6 represses AR activity by displacing and/or inhibiting recruitment of coactivators to AR target promoters. Understanding the mechanism for inhibition of AR coactivators can give rise to molecular targeted drugs for treatment of prostate cancer.

  7. Involvement of direct inhibition of NMDA receptors in the effects of sigma-receptor ligands on glutamate neurotoxicity in vitro.

    Science.gov (United States)

    Nishikawa, H; Hashino, A; Kume, T; Katsuki, H; Kaneko, S; Akaike, A

    2000-09-15

    This study was performed to examine the roles of the N-methyl-D-aspartate (NMDA) receptor/phencyclidine (PCP) channel complex in the protective effects of sigma-receptor ligands against glutamate neurotoxicity in cultured cortical neurons derived from fetal rats. A 1-h exposure of cultures to glutamate caused a marked loss of viability, as determined by Trypan blue exclusion. This acute neurotoxicity of glutamate was prevented by NMDA receptor antagonists. Expression of sigma(1) receptor mRNA in cortical cultures was confirmed by reverse transcription polymerase chain reaction (RT-PCR). sigma Receptor ligands with affinity for NMDA receptor channels including the PCP site, such as (+)-N-allylnormetazocine ((+)-SKF10,047), haloperidol, and R(-)-N-(3-phenyl-1-propyl)-1-phenyl-2-aminopropane ((-)-PPAP), prevented glutamate neurotoxicity in a concentration-dependent manner. In contrast, other sigma-receptor ligands without affinity for NMDA receptors, such as carbetapentane and R(+)-3-(3-hydroxyphenyl)-N-propylpiperidine ((+)-3-PPP), did not show neuroprotective effects. Putative endogenous sigma receptor ligands such as pregnenolone, progesterone, and dehydroepiandrosterone did not affect glutamate neurotoxicity. The protective effects of (+)-SKF10,047, haloperidol, and (-)-PPAP were not affected by the sigma(1) receptor antagonist rimcazole. These results suggested that a direct interaction with NMDA receptors but not with sigma receptors plays a crucial role in the neuroprotective effects of sigma receptor ligands with affinity for NMDA receptors.

  8. microRNA-150 inhibits the formation of macrophage foam cells through targeting adiponectin receptor 2

    Energy Technology Data Exchange (ETDEWEB)

    Li, Jing [Department of Geratory, Linzi District People’s Hospital of Zibo City, Zibo, Shandong (China); Zhang, Suhua, E-mail: drsuhuangzhang@qq.com [Department of HealthCare, Qilu Hospital of Shandong University (Qingdao), Qingdao City, Qingdao (China)

    2016-08-05

    Transformation of macrophages into foam cells plays a critical role in the pathogenesis of atherosclerosis. The aim of this study was to determine the expression and biological roles of microRNA (miR)-150 in the formation of macrophage foam cells and to identify its functional target(s). Exposure to 50 μg/ml oxidized low-density lipoprotein (oxLDL) led to a significant upregulation of miR-150 in THP-1 macrophages. Overexpression of miR-150 inhibited oxLDL-induced lipid accumulation in THP-1 macrophages, while knockdown of miR-150 enhanced lipid accumulation. apoA-I- and HDL-mediated cholesterol efflux was increased by 66% and 43%, respectively, in miR-150-overexpressing macrophages relative to control cells. In contrast, downregulation of miR-150 significantly reduced cholesterol efflux from oxLDL-laden macrophages. Bioinformatic analysis and luciferase reporter assay revealed adiponectin receptor 2 (AdipoR2) as a direct target of miR-150. Small interfering RNA-mediated downregulation of AdipoR2 phenocopied the effects of miR-150 overexpression, reducing lipid accumulation and facilitating cholesterol efflux in oxLDL-treated THP-1 macrophages. Knockdown of AdipoR2 induced the expression of proliferator-activated receptor gamma (PPARγ), liver X receptor alpha (LXRα), ABCA1, and ABCG1. Moreover, pharmacological inhibition of PPARγ or LXRα impaired AdipoR2 silencing-induced upregulation of ABCA1 and ABCG1. Taken together, our results indicate that miR-150 can attenuate oxLDL-induced lipid accumulation in macrophages via promotion of cholesterol efflux. The suppressive effects of miR-150 on macrophage foam cell formation are mediated through targeting of AdipoR2. Delivery of miR-150 may represent a potential approach to prevent macrophage foam cell formation in atherosclerosis. -- Highlights: •miR-150 inhibits macrophage foam cell formation. •miR-150 accelerates cholesterol efflux from oxLDL-laden macrophages. •miR-150 suppresses macrophage foam cell

  9. Cannabinoid 1 receptor knockout mice display cold allodynia, but enhanced recovery from spared-nerve injury-induced mechanical hypersensitivity.

    Science.gov (United States)

    Sideris, Alexandra; Piskoun, Boris; Russo, Lori; Norcini, Monica; Blanck, Thomas; Recio-Pinto, Esperanza

    2016-01-01

    The function of the Cannabinoid 1 receptor (CB1R) in the development of neuropathic pain is not clear. Mounting evidence suggest that CB1R expression and activation may contribute to pain. Cannabinoid 1 receptor knockout mice (CB1R-/-) generated on a C57Bl/6 background exhibit hypoalgesia in the hotplate assay and formalin test. These findings suggest that Cannabinoid 1 receptor expression mediates the responses to at least some types of painful stimuli. By using this mouse line, we sought to determine if the lack of Cannabinoid 1 receptor unveils a general hypoalgesic phenotype, including protection against the development of neuropathic pain. The acetone test was used to measure cold sensitivity, the electronic von Frey was used to measure mechanical thresholds before and after spared-nerve injury, and analysis of footprint patterns was conducted to determine if motor function is differentially affected after nerve-injury in mice with varying levels of Cannabinoid 1 receptor. At baseline, CB1R-/- mice were hypersensitive in the acetone test, and this phenotype was maintained after spared-nerve injury. Using calcium imaging of lumbar dorsal root ganglion (DRG) cultures, a higher percentage of neurons isolated from CB1R-/- mice were menthol sensitive relative to DRG isolated from wild-type (CB1R+/+) mice. Baseline mechanical thresholds did not differ among genotypes, and mechanical hypersensitivity developed similarly in the first two weeks following spared-nerve injury (SNI). At two weeks post-SNI, CB1R-/- mice recovered significantly from mechanical hypersensitivity, while the CB1R+/+ mice did not. Heterozygous knockouts (CB1R+/-) transiently developed cold allodynia only after injury, but recovered mechanical thresholds to a similar extent as the CB1R-/- mice. Sciatic functional indices, which reflect overall nerve health, and alternation coefficients, which indicate uniformity of strides, were not significantly different among genotypes. Cold allodynia and

  10. MECHANICAL VIBRATION INHIBITS OSTEOCLAST FORMATION BY REDUCING DC-STAMP RECEPTOR EXPRESSION IN OSTEOCLAST PRECURSOR CELLS

    Science.gov (United States)

    Kulkarni, R.N.; Voglewede, P.A.; Liu, D.

    2014-01-01

    It is well known that physical inactivity leads to loss of muscle mass, but it also causes bone loss. Mechanistically, osteoclastogenesis and bone resorption have recently been shown to be regulated by vibration. However, the underlying mechanism behind the inhibition of osteoclast formation is yet unknown. Therefore, we investigated whether mechanical vibration of osteoclast precursor cells affects osteoclast formation by the involvement of fusion-related molecules such as dendritic cell-specific transmembrane protein (DC-STAMP), and P2X7 receptor (P2X7R). RAW264.7 (a murine osteoclastic-like cell line) cells were treated with 20 ng/ml receptor activator of NF-κB ligand (RANKL). For 3 consecutive days, the cells were subjected to 1 hour of mechanical vibration with 20 µm displacement at a frequency of 4 Hz and compared to the control cells that were treated under the same condition but without the vibration. After 5 days of culture, osteoclast formation was determined. Gene expression of DC-STAMP and P2X7R by RAW264.7 cells were determined after 1 hour mechanical vibration, while protein production of the DC-STAMP was determined after 6 hours of post incubation after vibration. As a result, mechanical vibration of RAW264.7 cells inhibited the formation of osteoclasts. Vibration down-regulated DC-STAMP gene expression by 1.6-fold in the presence of RANKL and by 1.4-fold in the absence of RANKL. Additionally, DC-STAMP protein production was also down-regulated by 1.4-fold in the presence of RANKL and by 1.2-fold in the absence of RANKL in RAW264.7 cells in response to mechanical vibration. However, vibration did not affect P2X7R gene expression. Mouse anti-DC-STAMP antibody inhibited osteoclast formation in the absence of vibration. Our results suggest that mechanical vibration of osteoclast precursor cells reduce DC-STAMP expression in osteoclast precursor cells leading to the inhibition of osteoclast formation. PMID:23994170

  11. Mechanical vibration inhibits osteoclast formation by reducing DC-STAMP receptor expression in osteoclast precursor cells.

    Science.gov (United States)

    Kulkarni, Rishikesh N; Voglewede, Philip A; Liu, Dawei

    2013-12-01

    It is well known that physical inactivity leads to loss of muscle mass, but it also causes bone loss. Mechanistically, osteoclastogenesis and bone resorption have recently been shown to be regulated by vibration. However, the underlying mechanism behind the inhibition of osteoclast formation is yet unknown. Therefore, we investigated whether mechanical vibration of osteoclast precursor cells affects osteoclast formation by the involvement of fusion-related molecules such as dendritic cell-specific transmembrane protein (DC-STAMP) and P2X7 receptor (P2X7R). RAW264.7 (a murine osteoclastic-like cell line) cells were treated with 20ng/ml receptor activator of NF-κB ligand (RANKL). For 3 consecutive days, the cells were subjected to 1h of mechanical vibration with 20μm displacement at a frequency of 4Hz and compared to the control cells that were treated under the same condition but without the vibration. After 5days of culture, osteoclast formation was determined. Gene expression of DC-STAMP and P2X7R by RAW264.7 cells was determined after 1h of mechanical vibration, while protein production of the DC-STAMP was determined after 6h of postincubation after vibration. As a result, mechanical vibration of RAW264.7 cells inhibited the formation of osteoclasts. Vibration down-regulated DC-STAMP gene expression by 1.6-fold in the presence of RANKL and by 1.4-fold in the absence of RANKL. Additionally, DC-STAMP protein production was also down-regulated by 1.4-fold in the presence of RANKL and by 1.2-fold in the absence of RANKL in RAW264.7 cells in response to mechanical vibration. However, vibration did not affect P2X7R gene expression. Mouse anti-DC-STAMP antibody inhibited osteoclast formation in the absence of vibration. Our results suggest that mechanical vibration of osteoclast precursor cells reduces DC-STAMP expression in osteoclast precursor cells leading to the inhibition of osteoclast formation. © 2013 Elsevier Inc. All rights reserved.

  12. Inhibition of estrogen-responsive gene activation by the retinoid X receptor beta: evidence for multiple inhibitory pathways.

    Science.gov (United States)

    Segars, J H; Marks, M S; Hirschfeld, S; Driggers, P H; Martinez, E; Grippo, J F; Brown, M; Wahli, W; Ozato, K

    1993-04-01

    The retinoid X receptor beta (RXR beta; H-2RIIBP) forms heterodimers with various nuclear hormone receptors and binds multiple hormone response elements, including the estrogen response element (ERE). In this report, we show that endogenous RXR beta contributes to ERE binding activity in nuclear extracts of the human breast cancer cell line MCF-7. To define a possible regulatory role of RXR beta regarding estrogen-responsive transcription in breast cancer cells, RXR beta and a reporter gene driven by the vitellogenin A2 ERE were transfected into estrogen-treated MCF-7 cells. RXR beta inhibited ERE-driven reporter activity in a dose-dependent and element-specific fashion. This inhibition occurred in the absence of the RXR ligand 9-cis retinoic acid. The RXR beta-induced inhibition was specific for estrogen receptor (ER)-mediated ERE activation because inhibition was observed in ER-negative MDA-MB-231 cells only following transfection of the estrogen-activated ER. No inhibition of the basal reporter activity was observed. The inhibition was not caused by simple competition of RXR beta with the ER for ERE binding, since deletion mutants retaining DNA binding activity but lacking the N-terminal or C-terminal domain failed to inhibit reporter activity. In addition, cross-linking studies indicated the presence of an auxiliary nuclear factor present in MCF-7 cells that contributed to RXR beta binding of the ERE. Studies using known heterodimerization partners of RXR beta confirmed that RXR beta/triiodothyronine receptor alpha heterodimers avidly bind the ERE but revealed the existence of another triiodothyronine-independent pathway of ERE inhibition. These results indicate that estrogen-responsive genes may be negatively regulated by RXR beta through two distinct pathways.

  13. Scotopic vision in the monkey is modulated by the G protein-coupled receptor 55

    DEFF Research Database (Denmark)

    Bouskila, Joseph; Harrar, Vanessa; Javadi, Pasha

    2016-01-01

    The endogenous cannabinoid system plays important roles in the retina of mice and monkeys via their classic CB1 and CB2 receptors. We have previously reported that the G protein-coupled receptor 55 (GPR55), a putative cannabinoid receptor, is exclusively expressed in rod photoreceptors in the mon......The endogenous cannabinoid system plays important roles in the retina of mice and monkeys via their classic CB1 and CB2 receptors. We have previously reported that the G protein-coupled receptor 55 (GPR55), a putative cannabinoid receptor, is exclusively expressed in rod photoreceptors...

  14. Targeting Insulin-Like Growth Factor 1 Receptor Inhibits Pancreatic Cancer Growth and Metastasis

    Science.gov (United States)

    Subramani, Ramadevi; Lopez-Valdez, Rebecca; Arumugam, Arunkumar; Nandy, Sushmita; Boopalan, Thiyagarajan; Lakshmanaswamy, Rajkumar

    2014-01-01

    Pancreatic cancer is one of the most lethal cancers. Increasing incidence and mortality indicates that there is still much lacking in detection and management of the disease. This is partly due to a lack of specific symptoms during early stages of the disease. Several growth factor receptors have been associated with pancreatic cancer. Here, we have investigated if an RNA interference approach targeted to IGF-IR could be effective and efficient against pancreatic cancer growth and metastasis. For that, we evaluated the effects of IGF-1R inhibition using small interfering RNA (siRNAs) on tumor growth and metastasis in HPAC and PANC-1 pancreatic cancer cell lines. We found that silencing IGF-1R inhibits pancreatic cancer growth and metastasis by blocking key signaling pathways such AKT/PI3K, MAPK, JAK/STAT and EMT. Silencing IGF-1R resulted in an anti-proliferative effect in PANC-1 and HPAC pancreatic cancer cell lines. Matrigel invasion, transwell migration and wound healing assays also revealed a role for IGF-1R in metastatic properties of pancreatic cancer. These results were further confirmed using Western blotting analysis of key intermediates involved in proliferation, epithelial mesenchymal transition, migration, and invasion. In addition, soft agar assays showed that silencing IGF-1R also blocks the colony forming capabilities of pancreatic cancer cells in vitro. Western blots, as well as, flow cytometric analysis revealed the induction of apoptosis in IGF-1R silenced cells. Interestingly, silencing IGF-1R also suppressed the expression of insulin receptor β. All these effects together significantly control pancreatic cancer cell growth and metastasis. To conclude, our results demonstrate the significance of IGF-1R in pancreatic cancer. PMID:24809702

  15. Autoantibodies against Muscarinic Type 3 Receptor in Sjögren's Syndrome Inhibit Aquaporin 5 Trafficking

    Science.gov (United States)

    Lee, Byung Ha; Gauna, Adrienne E.; Perez, Geidys; Park, Yun-jong; Pauley, Kaleb M.; Kawai, Toshihisa; Cha, Seunghee

    2013-01-01

    Sjögren's syndrome (SjS) is a chronic autoimmune disease that mainly targets the salivary and lacrimal glands. It has been controversial whether anti-muscarinic type 3 receptor (α-M3R) autoantibodies in patients with SjS inhibit intracellular trafficking of aquaporin-5 (AQP5), water transport protein, leading to secretory dysfunction. To address this issue, GFP-tagged human AQP5 was overexpressed in human salivary gland cells (HSG-hAQP5) and monitored AQP5 trafficking to the plasma membrane following carbachol (CCh, M3R agonist) stimulation. AQP5 trafficking was indeed mediated by M3R stimulation, shown in partial blockage of trafficking by M3R-antagonist 4-DAMP. HSG-hAQP5 pre-incubated with SjS plasma for 24 hours significantly reduced AQP5 trafficking with CCh, compared with HSG-hAQP5 pre-incubated with healthy control (HC) plasma. This inhibition was confirmed by monoclonal α-M3R antibody and pre-absorbed plasma. Interestingly, HSG-hAQP5 pre-incubated with SjS plasma showed no change in cell volume, compared to the cells incubated with HC plasma showing shrinkage by twenty percent after CCh-stimulation. Our findings clearly indicate that binding of anti-M3R autoantibodies to the receptor, which was verified by immunoprecipitation, suppresses AQP5 trafficking to the membrane and contribute to impaired fluid secretion in SjS. Our current study urges further investigations of clinical associations between SjS symptoms, such as degree of secretory dysfunction, cognitive impairment, and/or bladder irritation, and different profiles (titers, isotypes, and/or specificity) of anti-M3R autoantibodies in individuals with SjS. PMID:23382834

  16. Liver X receptor activation inhibits PC-3 prostate cancer cells via the beta-catenin pathway.

    Science.gov (United States)

    Youlin, Kuang; Li, Zhang; Weiyang, He; Jian, Kang; Siming, Liang; Xin, Gou

    2017-03-01

    Liver X receptors (LXRs) are nuclear receptors family of ligand-dependent transcription factors that play a crucial role in regulating cholesterol metabolism and inflammation. Recent studies show that LXR agonists exhibit anti-cancer activities in a variety of cancer cell lines including prostate. To further identify the potential mechanisms of LXRα activation on prostate cancer, we investigated the effect of LXR agonist T0901317 on PC3 prostate cancer cell and in which activity of beta-catenin pathway involved. Prostate cancer PC3 cells were transfected with LXR-a siRNA and treated with LXR activator T0901317. qRT-PCR and western blot were used to detect the LXR-a expression. beta-catenin, cyclin D1 and c-MYC were analyzed by western blot. Cell apoptosis was examined by flow cytometry and Cell proliferation was assessed by Cell Counting Kit-8 assay. Cell migration was detected by Transwell chambers. Data showed that T0901317 significantly inhibited PC3 cell proliferation as well as invasion and increased apoptosis in vitro. Furthermore, we found that LXRα activation induced the reduction of beta-catenin expression in PC3 cells, and this inhibitory effect could be totally abolished when cells were treated with LXRα. Meanwhile, the expression of beta-catenin target gene cyclin D1 and c-MYC were also decreased. This study provided additional evidence that LXR activation inhibited PC-3 prostate cancer cells via suppressing beta-catenin pathway. Copyright © 2016 Elsevier GmbH. All rights reserved.

  17. Inhibition of fibroblast growth factor receptor with AZD4547 mitigates juvenile nasopharyngeal angiofibroma.

    Science.gov (United States)

    Le, Tran; New, Jacob; Jones, Joel W; Usman, Shireen; Yalamanchali, Sreeya; Tawfik, Ossama; Hoover, Larry; Bruegger, Dan E; Thomas, Sufi Mary

    2017-10-01

    Juvenile nasopharyngeal angiofibroma (JNA) is a benign tumor that presents in adolescent males. Although surgical excision is the mainstay of treatment, recurrences complicate treatment. There is a need to develop less invasive approaches for management. JNA tumors are composed of fibroblasts and vascular endothelial cells. We identified fibroblast growth factor receptor (FGFR) and vascular endothelial growth factor (VEGF) expression in JNA-derived fibroblasts. FGFR influences fibroblast proliferation and VEGF is necessary for angiogenesis. We hypothesized that targeting FGFR would mitigate JNA fibroblast proliferation, invasion, and migration, and that targeting the VEGF receptor would attenuate endothelial tubule formation. After informed consent, fibroblasts from JNA explants of 3 patients were isolated. Fibroblasts were treated with FGFR inhibitor AZD4547, 0 to 25 μg/mL for 72 hours and proliferation was quantified using CyQuant assay. Migration and invasion of JNA were assessed using 24-hour transwell assays with subsequent fixation and quantification. Mitigation of FGFR and downstream signaling was evaluated by immunoblotting. Tubule formation was assessed in human umbilical vein endothelial cells (HUVECs) treated with vehicle control (dimethylsulfoxide [DMSO]) or semaxanib (SU5416) as well as in serum-free media (SFM) or JNA conditioned media (CM). Tubule length was compared between treatment groups. Compared to control, AZD4547 inhibited JNA fibroblast proliferation, migration, and invasion through inhibition of FGFR and downstream signaling, specifically phosphorylation of - p44/42 mitogen activated protein kinase (p44/42 MAPK). JNA fibroblast CM significantly increased HUVEC tubule formation (p = 0.0039). AZD4547 effectively mitigates FGFR signaling and decreases JNA fibroblast proliferation, migration, and invasion. SU5416 attenuated JNA fibroblast-induced tubule formation. AZD4547 may have therapeutic potential in the treatment of JNA. © 2017 ARS

  18. Piperlongumine inhibits atherosclerotic plaque formation and vascular smooth muscle cell proliferation by suppressing PDGF receptor signaling

    Energy Technology Data Exchange (ETDEWEB)

    Son, Dong Ju [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Kim, Soo Yeon [Division of Life Science, Korea Basic Science Institute, Daejeon (Korea, Republic of); Han, Seong Su [University of Iowa Carver College of Medicine, Department of Pathology, Iowa City, IA (United States); Kim, Chan Woo [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Kumar, Sandeep [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Park, Byeoung Soo [Nanotoxtech Co., Ansan (Korea, Republic of); Lee, Sung Eun [Division of Applied Biology and Chemistry, Kyungpook National University, Daegu (Korea, Republic of); Yun, Yeo Pyo [College of Pharmacy, Chungbuk National University, Cheongju (Korea, Republic of); Jo, Hanjoong, E-mail: hjo@emory.edu [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Park, Young Hyun, E-mail: pyh012@sch.ac.kr [Department of Food Science and Nutrition, College of Natural Sciences, Soonchunhyang University, Asan (Korea, Republic of)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Anti-atherogenic effect of PL was examined using partial carotid ligation model in ApoE KO mice. Black-Right-Pointing-Pointer PL prevented atherosclerotic plaque development, VSMCs proliferation, and NF-{kappa}B activation. Black-Right-Pointing-Pointer Piperlongumine reduced vascular smooth muscle cell activation through PDGF-R{beta} and NF-{kappa}B-signaling. Black-Right-Pointing-Pointer PL may serve as a new therapeutic molecule for atherosclerosis treatment. -- Abstract: Piperlongumine (piplartine, PL) is an alkaloid found in the long pepper (Piper longum L.) and has well-documented anti-platelet aggregation, anti-inflammatory, and anti-cancer properties; however, the role of PL in prevention of atherosclerosis is unknown. We evaluated the anti-atherosclerotic potential of PL in an in vivo murine model of accelerated atherosclerosis and defined its mechanism of action in aortic vascular smooth muscle cells (VSMCs) in vitro. Local treatment with PL significantly reduced atherosclerotic plaque formation as well as proliferation and nuclear factor-kappa B (NF-{kappa}B) activation in an in vivo setting. PL treatment in VSMCs in vitro showed inhibition of migration and platelet-derived growth factor BB (PDGF-BB)-induced proliferation to the in vivo findings. We further identified that PL inhibited PDGF-BB-induced PDGF receptor beta activation and suppressed downstream signaling molecules such as phospholipase C{gamma}1, extracellular signal-regulated kinases 1 and 2 and Akt. Lastly, PL significantly attenuated activation of NF-{kappa}B-a downstream transcriptional regulator in PDGF receptor signaling, in response to PDGF-BB stimulation. In conclusion, our findings demonstrate a novel, therapeutic mechanism by which PL suppresses atherosclerosis plaque formation in vivo.

  19. The CB1 Neutral Antagonist Tetrahydrocannabivarin Reduces Default Mode Network and Increases Executive Control Network Resting State Functional Connectivity in Healthy Volunteers.

    Science.gov (United States)

    Rzepa, Ewelina; Tudge, Luke; McCabe, Ciara

    2015-09-10

    The cannabinoid cannabinoid type 1 (CB1) neutral antagonist tetrahydrocannabivarin (THCv) has been suggested as a possible treatment for obesity, but without the depressogenic side-effects of inverse antagonists such as Rimonabant. However, how THCv might affect the resting state functional connectivity of the human brain is as yet unknown. We examined the effects of a single 10mg oral dose of THCv and placebo in 20 healthy volunteers in a randomized, within-subject, double-blind design. Using resting state functional magnetic resonance imaging and seed-based connectivity analyses, we selected the amygdala, insula, orbitofrontal cortex, and dorsal medial prefrontal cortex (dmPFC) as regions of interest. Mood and subjective experience were also measured before and after drug administration using self-report scales. Our results revealed, as expected, no significant differences in the subjective experience with a single dose of THCv. However, we found reduced resting state functional connectivity between the amygdala seed region and the default mode network and increased resting state functional connectivity between the amygdala seed region and the dorsal anterior cingulate cortex and between the dmPFC seed region and the inferior frontal gyrus/medial frontal gyrus. We also found a positive correlation under placebo for the amygdala-precuneus connectivity with the body mass index, although this correlation was not apparent under THCv. Our findings are the first to show that treatment with the CB1 neutral antagonist THCv decreases resting state functional connectivity in the default mode network and increases connectivity in the cognitive control network and dorsal visual stream network. This effect profile suggests possible therapeutic activity of THCv for obesity, where functional connectivity has been found to be altered in these regions. © The Author 2015. Published by Oxford University Press on behalf of CINP.

  20. Soluble Form of Canine Transferrin Receptor Inhibits Canine Parvovirus Infection In Vitro and In Vivo

    Science.gov (United States)

    Wen, Jiexia; Pan, Sumin; Liang, Shuang; Zhong, Zhenyu; He, Ying; Lin, Hongyu; Li, Wenyan; Wang, Liyue; Li, Xiujin; Zhong, Fei

    2013-01-01

    Canine parvovirus (CPV) disease is an acute, highly infectious disease threatening the dog-raising industry. So far there are no effective therapeutic strategies to control this disease. Although the canine transferrin receptor (TfR) was identified as a receptor for CPV infection, whether extracellular domain of TfR (called soluble TfR (sTfR)) possesses anti-CPV activities remains elusive. Here, we used the recombinant sTfR prepared from HEK293T cells with codon-optimized gene structure to investigate its anti-CPV activity both in vitro and in vivo. Our results indicated that codon optimization could significantly improve sTfR expression in HEK293T cells. The prepared recombinant sTfR possessed a binding activity to both CPV and CPV VP2 capsid proteins and significantly inhibited CPV infection of cultured feline F81 cells and decreased the mortality of CPV-infected dogs, which indicates that the sTfR has the anti-CPV activity both in vitro and in vivo. PMID:24089666

  1. Soluble Form of Canine Transferrin Receptor Inhibits Canine Parvovirus Infection In Vitro and In Vivo

    Directory of Open Access Journals (Sweden)

    Jiexia Wen

    2013-01-01

    Full Text Available Canine parvovirus (CPV disease is an acute, highly infectious disease threatening the dog-raising industry. So far there are no effective therapeutic strategies to control this disease. Although the canine transferrin receptor (TfR was identified as a receptor for CPV infection, whether extracellular domain of TfR (called soluble TfR (sTfR possesses anti-CPV activities remains elusive. Here, we used the recombinant sTfR prepared from HEK293T cells with codon-optimized gene structure to investigate its anti-CPV activity both in vitro and in vivo. Our results indicated that codon optimization could significantly improve sTfR expression in HEK293T cells. The prepared recombinant sTfR possessed a binding activity to both CPV and CPV VP2 capsid proteins and significantly inhibited CPV infection of cultured feline F81 cells and decreased the mortality of CPV-infected dogs, which indicates that the sTfR has the anti-CPV activity both in vitro and in vivo.

  2. Blocking oxytocin receptors inhibits vaginal marking to male odors in female Syrian hamsters.

    Science.gov (United States)

    Martinez, Luis A; Albers, H Elliott; Petrulis, Aras

    2010-12-02

    In Syrian hamsters (Mesocricetus auratus), precopulatory behaviors such as vaginal scent marking are essential for attracting a suitable mate. Vaginal marking is dependent on forebrain areas implicated in the neural regulation of reproductive behaviors in rodents, including the medial preoptic/anterior hypothalamus (MPOA-AH). Within MPOA-AH, the neuropeptide oxytocin (OT) acts to facilitate copulation (lordosis), as well as ultrasonic vocalizations towards males. It is not known, however, if OT in this area also facilitates vaginal marking. In the present study, a specific oxytocin receptor antagonist (OTA) was injected into MPOA-AH of intact female Syrian hamsters to determine if oxytocin receptor-dependent signaling is critical for the normal expression of vaginal marking elicited by male, female, and clean odors. OTA injections significantly inhibited vaginal marking in response to male odors compared with vehicle injections. There was no effect of OTA on marking in response to either female or clean odors. When injected into the bed nucleus of the stria terminalis (BNST), a nearby region to MPOA-AH, OTA was equally effective in decreasing marking. Finally, the effects of OTA appear to be specific to vaginal marking, as OTA injections in MPOA-AH or BNST did not alter general locomotor activity, flank marking, or social odor investigation. Considered together, these results suggest that OT in MPOA-AH and/or BNST normally facilitates male odor-induced vaginal marking, providing further evidence that OT generally supports prosocial interactions among conspecifics. Copyright © 2010 Elsevier Inc. All rights reserved.

  3. Structural basis for subtype-specific inhibition of the P2X7 receptor

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    Karasawa, Akira; Kawate, Toshimitsu (Cornell)

    2016-12-09

    The P2X7 receptor is a non-selective cation channel activated by extracellular adenosine triphosphate (ATP). Chronic activation of P2X7 underlies many health problems such as pathologic pain, yet we lack effective antagonists due to poorly understood mechanisms of inhibition. Here we present crystal structures of a mammalian P2X7 receptor complexed with five structurally-unrelated antagonists. Unexpectedly, these drugs all bind to an allosteric site distinct from the ATP-binding pocket in a groove formed between two neighboring subunits. This novel drug-binding pocket accommodates a diversity of small molecules mainly through hydrophobic interactions. Functional assays propose that these compounds allosterically prevent narrowing of the drug-binding pocket and the turret-like architecture during channel opening, which is consistent with a site of action distal to the ATP-binding pocket. These novel mechanistic insights will facilitate the development of P2X7-specific drugs for treating human diseases.

  4. Desipramine inhibits histamine H1 receptor-induced Ca2+ signaling in rat hypothalamic cells.

    Directory of Open Access Journals (Sweden)

    Ji-Ah Kang

    Full Text Available The hypothalamus in the brain is the main center for appetite control and integrates signals from adipose tissue and the gastrointestinal tract. Antidepressants are known to modulate the activities of hypothalamic neurons and affect food intake, but the cellular and molecular mechanisms by which antidepressants modulate hypothalamic function remain unclear. Here we have investigated how hypothalamic neurons respond to treatment with antidepressants, including desipramine and sibutramine. In primary cultured rat hypothalamic cells, desipramine markedly suppressed the elevation of intracellular Ca(2+ evoked by histamine H1 receptor activation. Desipramine also inhibited the histamine-induced Ca(2+ increase and the expression of corticotrophin-releasing hormone in hypothalamic GT1-1 cells. The effect of desipramine was not affected by pretreatment with prazosin or propranolol, excluding catecholamine reuptake activity of desipramine as an underlying mechanism. Sibutramine which is also an antidepressant but decreases food intake, had little effect on the histamine-induced Ca(2+ increase or AMP-activated protein kinase activity. Our results reveal that desipramine and sibutramine have different effects on histamine H1 receptor signaling in hypothalamic cells and suggest that distinct regulation of hypothalamic histamine signaling might underlie the differential regulation of food intake between antidepressants.

  5. Inhibition of the MEK-1/p42 MAP kinase reduces aryl hydrocarbon receptor-DNA interactions

    International Nuclear Information System (INIS)

    Yim, Sujin; Oh, Myoungsuk; Choi, Su Mi; Park, Hyunsung

    2004-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induces expression of the cytochrome P450 1A1 gene, cyp1a1, by binding to its receptor, aryl hydrocarbon receptor (AhR). TCDD-bound AhR translocates to the nucleus and forms a heterodimer with its partner protein, AhR nuclear translocator (Arnt). The AhR/Arnt heterodimer then binds to the dioxin-response elements (DREs) in the cyp1a1 enhancer and stimulates transcription of cyp1a1. We tested whether kinase pathways are involved in this process by treating Hepa1c1c7 cells with kinase inhibitors. The MEK-1 inhibitor PD98059 reduced TCDD-induced transcription of cyp1a1. TCDD treatment results in phosphorylation of p44/p42 mitogen-activated protein kinase (MAPK), a substrate of MEK-1. Overexpression of dominant negative form of p42 MAPK suppressed TCDD-dependent transcription of a reporter gene controlled by dioxin-response elements (DREs), and pretreatment with PD98059 also blocked this transcription. PD98059 pretreatment also inhibited TCDD-induced DRE binding of the AhR/Arnt heterodimer. Together these results indicate that TCDD activates the MEK-1/p44/p42 MAPK pathway, which in turn activates AhR and so facilitates binding of AhR to the cyp1a1 DRE

  6. CINPA1 binds directly to constitutive androstane receptor and inhibits its activity.

    Science.gov (United States)

    Cherian, Milu T; Chai, Sergio C; Wright, William C; Singh, Aman; Alexandra Casal, Morgan; Zheng, Jie; Wu, Jing; Lee, Richard E; Griffin, Patrick R; Chen, Taosheng

    2018-03-31

    The constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are xenobiotic sensors that regulate the expression of drug-metabolizing enzymes and efflux transporters. CAR activation promotes drug elimination, thereby reducing therapeutic effectiveness, or causes adverse drug effects via toxic metabolites. CAR inhibitors could be used to attenuate these adverse drug effects. CAR and PXR share ligands and target genes, confounding the understanding of the regulation of receptor-specific activity. We previously identified a small-molecule inhibitor, CINPA1, that inhibits CAR (without activating PXR at lower concentrations) by altering CAR-coregulator interactions and reducing CAR recruitment to DNA response elements of regulated genes. However, solid evidence was not presented for the direct binding of CINPA1 to CAR. In this study, we demonstrate direct interaction of CINPA1 with the CAR ligand-binding domain (CAR-LBD) and identify key residues involved in such interactions through a combination of biophysical and computational methods. We found that CINPA1 resides in the ligand-binding pocket to stabilize the CAR-LBD in a more rigid, less fluid state. Molecular dynamics simulations, together with our previously reported docking model, enabled us to predict which CAR residues were critical for interactions with CINPA1. The importance of these residues for CINPA1 binding were then validated by directed mutations and testing the mutant CAR proteins in transcription reporter and coregulatory interaction assays. We demonstrated strong hydrogen bonding of CINPA1 with N165 and H203 and identified other residues involved in hydrophobic contacts with CINPA1. Overall, our data confirm that CINPA1 directly binds to CAR. Copyright © 2018 Elsevier Inc. All rights reserved.

  7. Haloperidol inhibits the development of atherosclerotic lesions in LDL receptor knockout mice.

    Science.gov (United States)

    van der Sluis, Ronald J; Nahon, Joya E; Reuwer, Anne Q; Van Eck, Miranda; Hoekstra, Menno

    2015-05-01

    Antipsychotic drugs have been shown to modulate the expression of ATP-binding cassette transporter A1 (ABCA1), a key factor in the anti-atherogenic reverse cholesterol transport process, in vitro. Here we evaluated the potential of the typical antipsychotic drug haloperidol to modulate the cholesterol efflux function of macrophages in vitro and their susceptibility to atherosclerosis in vivo. Thioglycollate-elicited peritoneal macrophages were used for in vitro studies. Hyperlipidaemic low-density lipoprotein (LDL) receptor knockout mice were implanted with a haloperidol-containing pellet and subsequently fed a Western-type diet for 5 weeks to induce the development of atherosclerotic lesions in vivo. Haloperidol induced a 54% decrease in the mRNA expression of ABCA1 in peritoneal macrophages. This coincided with a 30% decrease in the capacity of macrophages to efflux cholesterol to apolipoprotein A1. Haloperidol treatment stimulated the expression of ABCA1 (+51%) and other genes involved in reverse cholesterol transport, that is, CYP7A1 (+98%) in livers of LDL receptor knockout mice. No change in splenic ABCA1 expression was noted. However, the average size of the atherosclerotic size was significantly smaller (-31%) in the context of a mildly more atherogenic metabolic phenotype upon haloperidol treatment. More importantly, haloperidol markedly lowered MCP-1 expression (-70%) and secretion (-28%) by peritoneal macrophages. Haloperidol treatment lowered the susceptibility of hyperlipidaemic LDL receptor knockout mice to develop atherosclerotic lesions. Our findings suggest that the beneficial effect of haloperidol on atherosclerosis susceptibility can be attributed to its ability to inhibit macrophage chemotaxis. © 2015 The British Pharmacological Society.

  8. The Ly49E receptor inhibits the immune control of acute Trypanosoma cruzi infection

    Directory of Open Access Journals (Sweden)

    Jessica Filtjens

    2016-11-01

    Full Text Available The protozoan parasite Trypanosoma cruzi (T. cruzi circulates in the blood upon infection and invades a variety of cells. Parasites intensively multiply during the acute phase of infection and persist lifelong at low levels in tissues and blood during the chronic phase. Natural killer (NK and NKT cells play an important role in the immune control of T. cruzi infection, mainly by releasing the cytokine IFN-γ that activates the microbicidal action of macrophages and other cells and shapes a protective type 1 immune response. The mechanisms by which immune cells are regulated to produce IFN-γ during T. cruzi infection are still incompletely understood. Here, we show that urokinase plasminogen activator (uPA is induced early upon T. cruzi infection, and remains elevated until day 20 post inoculation. We previously demonstrated that the inhibitory receptor Ly49E, which is expressed, among others, on NK and NKT cells, is triggered by uPA. Therefore, we compared wild type (WT to Ly49E knockout (KO mice for their control of experimental T. cruzi infection. Our results show that young, i.e. 4- and 6-week-old, Ly49E KO mice control the infection better than WT mice, indicated by a lower parasite load and less cachexia. The beneficial effect of Ly49E depletion is more obvious in 4-week-old male than in female mice and weakens in 8-week-old mice. In young mice, the lower T. cruzi parasitemia in Ly49E KO mice is paralleled by higher IFN-γ production compared to their WT controls. Our data indicate that Ly49E receptor expression inhibits the immune control of T. cruzi infection. This is the first demonstration that the inhibitory Ly49E receptor can interfere with the immune response to a pathogen in vivo.

  9. An interspecies comparison of mercury inhibition on muscarinic acetylcholine receptor binding in the cerebral cortex and cerebellum

    International Nuclear Information System (INIS)

    Basu, Niladri; Stamler, Christopher J.; Loua, Kovana Marcel; Chan, H.M.

    2005-01-01

    Mercury (Hg) is a ubiquitous pollutant that can disrupt neurochemical signaling pathways in mammals. It is well documented that inorganic Hg (HgCl 2 ) and methyl Hg (MeHg) can inhibit the binding of radioligands to the muscarinic acetylcholine (mACh) receptor in rat brains. However, little is known concerning this relationship in specific anatomical regions of the brain or in other species, including humans. The purpose of this study was to explore the inhibitory effects of HgCl 2 and MeHg on [ 3 H]-quinuclidinyl benzilate ([ 3 H]-QNB) binding to the mACh receptor in the cerebellum and cerebral cortex regions from human, rat, mouse, mink, and river otter brain tissues. Saturation binding curves were obtained from each sample to calculate receptor density (B max ) and ligand affinity (K d ). Subsequently, samples were exposed to HgCl 2 or MeHg to derive IC50 values and inhibition constants (K i ). Results demonstrate that HgCl 2 is a more potent inhibitor of mACh receptor binding than MeHg, and the receptors in the cerebellum are more sensitive to Hg-mediated mACh receptor inhibition than those in the cerebral cortex. Species sensitivities, irrespective of Hg type and brain region, can be ranked from most to least sensitive: river otter > rat > mink > mouse > humans. In summary, our data demonstrate that Hg can inhibit the binding [ 3 H]-QNB to the mACh receptor in a range of mammalian species. This comparative study provides data on interspecies differences and a framework for interpreting results from human, murine, and wildlife studies

  10. Adenosine A₂A receptors inhibit delayed rectifier potassium currents and cell differentiation in primary purified oligodendrocyte cultures.

    Science.gov (United States)

    Coppi, Elisabetta; Cellai, Lucrezia; Maraula, Giovanna; Pugliese, Anna Maria; Pedata, Felicita

    2013-10-01

    Oligodendrocyte progenitor cells (OPCs) are a population of cycling cells which persist in the adult central nervous system (CNS) where, under opportune stimuli, they differentiate into mature myelinating oligodendrocytes. Adenosine A(2A) receptors are Gs-coupled P1 purinergic receptors which are widely distributed throughout the CNS. It has been demonstrated that OPCs express A(2A) receptors, but their functional role in these cells remains elusive. Oligodendrocytes express distinct voltage-gated ion channels depending on their maturation. Here, by electrophysiological recordings coupled with immunocytochemical labeling, we studied the effects of adenosine A(2A) receptors on membrane currents and differentiation of purified primary OPCs isolated from the rat cortex. We found that the selective A(2A) agonist, CGS21680, inhibits sustained, delayed rectifier, K(+) currents (I(K)) without modifying transient (I(A)) conductances. The effect was observed in all cells tested, independently from time in culture. CGS21680 inhibition of I(K) current was concentration-dependent (10-200 nM) and blocked in the presence of the selective A(2A) antagonist SCH58261 (100 nM). It is known that I(K) currents play an important role during OPC development since their block decreases cell proliferation and differentiation. In light of these data, our further aim was to investigate whether A(2A) receptors modulate these processes. CGS21680, applied at 100 nM in the culture medium of oligodendrocyte cultures, inhibits OPC differentiation (an effect prevented by SCH58261) without affecting cell proliferation. Data demonstrate that cultured OPCs express functional A(2A) receptors whose activation negatively modulate I(K) currents. We propose that, by this mechanism, A(2A) adenosine receptors inhibit OPC differentiation. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Midazolam inhibits chondrogenesis via peripheral benzodiazepine receptor in human mesenchymal stem cells.

    Science.gov (United States)

    Chen, Yung-Ching; Wu, King-Chuen; Huang, Bu-Miin; So, Edmund Cheung; Wang, Yang-Kao

    2018-05-01

    Midazolam, a benzodiazepine derivative, is widely used for sedation and surgery. However, previous studies have demonstrated that Midazolam is associated with increased risks of congenital malformations, such as dwarfism, when used during early pregnancy. Recent studies have also demonstrated that Midazolam suppresses osteogenesis of mesenchymal stem cells (MSCs). Given that hypertrophic chondrocytes can differentiate into osteoblast and osteocytes and contribute to endochondral bone formation, the effect of Midazolam on chondrogenesis remains unclear. In this study, we applied a human MSC line, the KP cell, to serve as an in vitro model to study the effect of Midazolam on chondrogenesis. We first successfully established an in vitro chondrogenic model in a micromass culture or a 2D high-density culture performed with TGF-β-driven chondrogenic induction medium. Treatment of the Midazolam dose-dependently inhibited chondrogenesis, examined using Alcian blue-stained glycosaminoglycans and the expression of chondrogenic markers, such as SOX9 and type II collagen. Inhibition of Midazolam by peripheral benzodiazepine receptor (PBR) antagonist PK11195 or small interfering RNA rescued the inhibitory effects of Midazolam on chondrogenesis. In addition, Midazolam suppressed transforming growth factor-β-induced Smad3 phosphorylation, and this inhibitory effect could be rescued using PBR antagonist PK11195. This study provides a possible explanation for Midazolam-induced congenital malformations of the musculoskeletal system through PBR. © 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  12. Emerging role of epidermal growth factor receptor inhibition in therapy for advanced malignancy: focus on NSCLC

    International Nuclear Information System (INIS)

    Langer, Corey J.

    2004-01-01

    Combination chemotherapy regimens have emerged as the standard approach in advanced non-small-cell lung cancer. Meta-analyses have demonstrated a 2-month increase in median survival after platinum-based therapy vs. best supportive care, and an absolute 10% improvement in the 1-year survival rate. Just as importantly, cytotoxic therapy has produced benefits in symptom control and quality of life. Newer agents, including the taxanes, vinorelbine, gemcitabine, and irinotecan, have expanded our therapeutic options in the treatment of advanced non-small-cell lung cancer. Despite their contributions, we have reached a therapeutic plateau, with response rates seldom exceeding 30-40% in cooperative group studies and 1-year survival rates stable between 30% and 40%. It is doubtful that substituting one agent for another in various combinations will lead to any further improvement in these rates. The thrust of current research has focused on targeted therapy, and epidermal growth factor receptor inhibition is one of the most promising clinical strategies. Epidermal growth factor receptor inhibitors currently under investigation include the small molecules gefitinib (Iressa, ZD1839) and erlotinib (Tarceva, OSI-774), as well as monoclonal antibodies such as cetuximab (IMC-225, Erbitux). Agents that have only begun to undergo clinical evaluation include CI-1033, an irreversible pan-erbB tyrosine kinase inhibitor, and PKI166 and GW572016, both examples of dual kinase inhibitors (inhibiting epidermal growth factor receptor and Her2). Preclinical models have demonstrated synergy for all these agents in combination with either chemotherapy or radiotherapy, leading to great enthusiasm regarding their ultimate contribution to lung cancer therapy. However, serious clinical challenges persist. These include the identification of the optimal dose(s); the proper integration of these agents into popular, established cytotoxic regimens; and the selection of the optimal setting(s) in which

  13. Valerian inhibits rat hepatocarcinogenesis by activating GABA(A receptor-mediated signaling.

    Directory of Open Access Journals (Sweden)

    Anna Kakehashi

    Full Text Available Valerian is widely used as a traditional medicine to improve the quality of sleep due to interaction of several active components with the γ-aminobutyric acid (GABA A receptor (GABA(AR system. Recently, activation of GABA signaling in stem cells has been reported to suppress cell cycle progression in vivo. Furthermore, possible inhibitory effects of GABA(AR agonists on hepatocarcinogenesis have been reported. The present study was performed to investigate modulating effects of Valerian on hepatocarcinogenesis using a medium-term rat liver bioassay. Male F344 rats were treated with one of the most powerful Valerian species (Valeriana sitchensis at doses of 0, 50, 500 and 5000 ppm in their drinking water after initiation of hepatocarcinogenesis with diethylnitrosamine (DEN. Formation of glutathione S-transferase placental form positive (GST-P(+ foci was significantly inhibited by Valerian at all applied doses compared with DEN initiation control rats. Generation of 8-hydroxy-2'-deoxyguanosine in the rat liver was significantly suppressed by all doses of Valerian, likely due to suppression of Nrf2, CYP7A1 and induction of catalase expression. Cell proliferation was significantly inhibited, while apoptosis was induced in areas of GST-P(+ foci of Valerian groups associated with suppression of c-myc, Mafb, cyclin D1 and induction of p21(Waf1/Cip1, p53 and Bax mRNA expression. Interestingly, expression of the GABA(AR alpha 1 subunit was observed in GST-P(+ foci of DEN control rats, with significant elevation associated with Valerian treatment. These results indicate that Valerian exhibits inhibitory effects on rat hepatocarcinogenesis by inhibiting oxidative DNA damage, suppressing cell proliferation and inducing apoptosis in GST-P(+ foci by activating GABA(AR-mediated signaling.

  14. Effects of epidermal growth factor receptor kinase inhibition on radiation response in canine osteosarcoma cells.

    Science.gov (United States)

    Mantovani, Fernanda B; Morrison, Jodi A; Mutsaers, Anthony J

    2016-05-31

    Radiation therapy is a palliative treatment modality for canine osteosarcoma, with transient improvement in analgesia observed in many cases. However there is room for improvement in outcome for these patients. It is possible that the addition of sensitizing agents may increase tumor response to radiation therapy and prolong quality of life. Epidermal growth factor receptor (EGFR) expression has been documented in canine osteosarcoma and higher EGFR levels have been correlated to a worse prognosis. However, effects of EGFR inhibition on radiation responsiveness in canine osteosarcoma have not been previously characterized. This study examined the effects of the small molecule EGFR inhibitor erlotinib on canine osteosarcoma radiation responses, target and downstream protein expression in vitro. Additionally, to assess the potential impact of treatment on tumor angiogenesis, vascular endothelial growth factor (VEGF) levels in conditioned media were measured. Erlotinib as a single agent reduced clonogenic survival in two canine osteosarcoma cell lines and enhanced the impact of radiation in one out of three cell lines investigated. In cell viability assays, erlotinib enhanced radiation effects and demonstrated single agent effects. Erlotinib did not alter total levels of EGFR, nor inhibit downstream protein kinase B (PKB/Akt) activation. On the contrary, erlotinib treatment increased phosphorylated Akt in these osteosarcoma cell lines. VEGF levels in conditioned media increased after erlotinib treatment as a single agent and in combination with radiation in two out of three cell lines investigated. However, VEGF levels decreased with erlotinib treatment in the third cell line. Erlotinib treatment promoted modest enhancement of radiation effects in canine osteosarcoma cells, and possessed activity as a single agent in some cell lines, indicating a potential role for EGFR inhibition in the treatment of a subset of osteosarcoma patients. The relative radioresistance of

  15. Enzalutamide inhibits proliferation of gemcitabine-resistant bladder cancer cells with increased androgen receptor expression.

    Science.gov (United States)

    Kameyama, Koji; Horie, Kengo; Mizutani, Kosuke; Kato, Taku; Fujita, Yasunori; Kawakami, Kyojiro; Kojima, Toshio; Miyazaki, Tatsuhiko; Deguchi, Takashi; Ito, Masafumi

    2017-01-01

    Advanced bladder cancer is treated mainly with gemcitabine and cisplatin, but most patients eventually become resistance. Androgen receptor (AR) signaling has been implicated in bladder cancer as well as other types of cancer including prostate cancer. In this study, we investigated the expression and role of AR in gemcitabine-resistant bladder cancer cells and also the potential of enzalutamide, an AR inhibitor, as a therapeutic for the chemoresistance. First of all, we established gemcitabine-resistant T24 cells (T24GR) from T24 bladder cancer cells and performed gene expression profiling. Microarray analysis revealed upregulation of AR expression in T24GR cells compared with T24 cells. AR mRNA and protein expression was confirmed to be increased in T24GR cells, respectively, by quantitative RT-PCR and western blot analysis, which was associated with more potent AR transcriptional activity as measured by luciferase reporter assay. The copy number of AR gene in T24GR cells determined by PCR was twice as many as that of T24 cells. AR silencing by siRNA transfection resulted in inhibition of proliferation of T24GR cells. Cell culture in charcoal-stripped serum and treatment with enzalutamide inhibited growth of T24GR cells, which was accompanied by cell cycle arrest. AR transcriptional activity was found to be reduced in T24GR cells by enzalutamide treatment. Lastly, enzalutamide also inhibited cell proliferation of HTB5 bladder cancer cells that express AR and possess intrinsic resistance to gemcitabine. Our results suggest that enzalutamide may have the potential to treat patients with advanced gemcitabine-resistant bladder cancer with increased AR expression.

  16. Estrogen receptor beta signaling inhibits PDGF induced human airway smooth muscle proliferation.

    Science.gov (United States)

    Ambhore, Nilesh Sudhakar; Katragadda, Rathnavali; Raju Kalidhindi, Rama Satyanarayana; Thompson, Michael A; Pabelick, Christina M; Prakash, Y S; Sathish, Venkatachalem

    2018-04-20

    Airway smooth muscle (ASM) cell hyperplasia driven by persistent inflammation is a hallmark feature of remodeling in asthma. Sex steroid signaling in the lungs is of considerable interest, given epidemiological data showing more asthma in pre-menopausal women and aging men. Our previous studies demonstrated that estrogen receptor (ER) expression increases in asthmatic human ASM; however, very limited data are available regarding differential roles of ERα vs. ERβ isoforms in human ASM cell proliferation. In this study, we evaluated the effect of selective ERα and ERβ modulators on platelet-derived growth factor (PDGF)-stimulated ASM proliferation and the mechanisms involved. Asthmatic and non-asthmatic primary human ASM cells were treated with PDGF, 17β-estradiol, ERα-agonist and/or ERβ-agonist and/or G-protein-coupled estrogen receptor 30 (GPR30/GPER) agonist and proliferation was measured using MTT and CyQuant assays followed by cell cycle analysis. Transfection of small interfering RNA (siRNA) ERα and ERβ significantly altered the human ASM proliferation. The specificity of siRNA transfection was confirmed by Western blot analysis. Gene and protein expression of cell cycle-related antigens (PCNA and Ki67) and C/EBP were measured by RT-PCR and Western analysis, along with cell signaling proteins. PDGF significantly increased ASM proliferation in non-asthmatic and asthmatic cells. Treatment with PPT showed no significant effect on PDGF-induced proliferation, whereas WAY interestingly suppressed proliferation via inhibition of ERK1/2, Akt, and p38 signaling. PDGF-induced gene expression of PCNA, Ki67 and C/EBP in human ASM was significantly lower in cells pre-treated with WAY. Furthermore, WAY also inhibited PDGF-activated PCNA, C/EBP, cyclin-D1, and cyclin-E. Overall, we demonstrate ER isoform-specific signaling in the context of ASM proliferation. Activation of ERβ can diminish remodeling in human ASM by inhibiting pro-proliferative signaling pathways

  17. β-Adrenergic receptor stimulation inhibits proarrhythmic alternans in postinfarction border zone cardiomyocytes: a computational analysis.

    Science.gov (United States)

    Tomek, Jakub; Rodriguez, Blanca; Bub, Gil; Heijman, Jordi

    2017-08-01

    The border zone (BZ) of the viable myocardium adjacent to an infarct undergoes extensive autonomic and electrical remodeling and is prone to repolarization alternans-induced cardiac arrhythmias. BZ remodeling processes may promote or inhibit Ca 2+ and/or repolarization alternans and may differentially affect ventricular arrhythmogenesis. Here, we used a detailed computational model of the canine ventricular cardiomyocyte to study the determinants of alternans in the BZ and their regulation by β-adrenergic receptor (β-AR) stimulation. The BZ model developed Ca 2+ transient alternans at slower pacing cycle lengths than the control model, suggesting that the BZ may promote spatially heterogeneous alternans formation in an infarcted heart. β-AR stimulation abolished alternans. By evaluating all combinations of downstream β-AR stimulation targets, we identified both direct (via ryanodine receptor channels) and indirect [via sarcoplasmic reticulum (SR) Ca 2+ load] modulation of SR Ca 2+ release as critical determinants of Ca 2+ transient alternans. These findings were confirmed in a human ventricular cardiomyocyte model. Cell-to-cell coupling indirectly modulated the likelihood of alternans by affecting the action potential upstroke, reducing the trigger for SR Ca 2+ release in one-dimensional strand simulations. However, β-AR stimulation inhibited alternans in both single and multicellular simulations. Taken together, these data highlight a potential antiarrhythmic role of sympathetic hyperinnervation in the BZ by reducing the likelihood of alternans and provide new insights into the underlying mechanisms controlling Ca 2+ transient and repolarization alternans. NEW & NOTEWORTHY We integrated, for the first time, postmyocardial infarction electrical and autonomic remodeling in a detailed, validated computer model of β-adrenergic stimulation in ventricular cardiomyocytes. Here, we show that β-adrenergic stimulation inhibits alternans and provide novel insights

  18. Lipids Derived from Virulent Francisella tularensis Broadly Inhibit Pulmonary Inflammation via Toll-Like Receptor 2 and Peroxisome Proliferator-Activated Receptor α

    Science.gov (United States)

    Crane, Deborah D.; Ireland, Robin; Alinger, Joshua B.; Small, Pamela

    2013-01-01

    Francisella tularensis is a Gram-negative facultative intracellular pathogen that causes an acute lethal respiratory disease in humans. The heightened virulence of the pathogen is linked to its unique ability to inhibit Toll-like receptor (TLR)-mediated inflammatory responses. The bacterial component and mechanism of this inhibition are unknown. Here we show that lipids isolated from virulent but not attenuated strains of F. tularensis are not detected by host cells, inhibit production of proinflammatory cytokines by primary macrophages in response to known TLR ligands, and suppress neutrophil recruitment in vivo. We further show that lipid-mediated inhibition of inflammation is dependent on TLR2, MyD88, and the nuclear hormone and fatty acid receptor peroxisome proliferator-activated receptor α (PPARα). Pathogen lipid-mediated interference with inflammatory responses through the engagement of TLR2 and PPARα represents a novel manipulation of host signaling pathways consistent with the ability of highly virulent F. tularensis to efficiently evade host immune responses. PMID:23925884

  19. Inhibition of cerebrovascular raf activation attenuates cerebral blood flow and prevents upregulation of contractile receptors after subarachnoid hemorrhage

    DEFF Research Database (Denmark)

    Ansar, Saema; Maddahi, Aida; Edvinsson, Lars

    2011-01-01

    of mitogen-activated protein kinase (MAPK) of the extracellular signal-regulated kinase (ERK)1/2 signal pathway. We hypothesize that SAH initiates cerebrovascular ERK1/2 activation, resulting in receptor upregulation. The raf inhibitor will inhibit the molecular events upstream ERK1/2 and may provide...

  20. Mannose receptor induces T-cell tolerance via inhibition of CD45 and up-regulation of CTLA-4.

    Science.gov (United States)

    Schuette, Verena; Embgenbroich, Maria; Ulas, Thomas; Welz, Meike; Schulte-Schrepping, Jonas; Draffehn, Astrid M; Quast, Thomas; Koch, Katharina; Nehring, Melanie; König, Jessica; Zweynert, Annegret; Harms, Frederike L; Steiner, Nancy; Limmer, Andreas; Förster, Irmgard; Berberich-Siebelt, Friederike; Knolle, Percy A; Wohlleber, Dirk; Kolanus, Waldemar; Beyer, Marc; Schultze, Joachim L; Burgdorf, Sven

    2016-09-20

    The mannose receptor (MR) is an endocytic receptor involved in serum homeostasis and antigen presentation. Here, we identify the MR as a direct regulator of CD8(+) T-cell activity. We demonstrate that MR expression on dendritic cells (DCs) impaired T-cell cytotoxicity in vitro and in vivo. This regulatory effect of the MR was mediated by a direct interaction with CD45 on the T cell, inhibiting its phosphatase activity, which resulted in up-regulation of cytotoxic T-lymphocyte-associated Protein 4 (CTLA-4) and the induction of T-cell tolerance. Inhibition of CD45 prevented expression of B-cell lymphoma 6 (Bcl-6), a transcriptional inhibitor that directly bound the CTLA-4 promoter and regulated its activity. These data demonstrate that endocytic receptors expressed on DCs contribute to the regulation of T-cell functionality.

  1. Genetic background can result in a marked or minimal effect of gene knockout (GPR55 and CB2 receptor in experimental autoimmune encephalomyelitis models of multiple sclerosis.

    Directory of Open Access Journals (Sweden)

    Sofia Sisay

    Full Text Available Endocannabinoids and some phytocannabinoids bind to CB1 and CB2 cannabinoid receptors, transient receptor potential vanilloid one (TRPV1 receptor and the orphan G protein receptor fifty-five (GPR55. Studies using C57BL/10 and C57BL/6 (Cnr2 (tm1Zim CB2 cannabinoid receptor knockout mice have demonstrated an immune-augmenting effect in experimental autoimmune encephalomyelitis (EAE models of multiple sclerosis. However, other EAE studies in Biozzi ABH mice often failed to show any treatment effect of either CB2 receptor agonism or antagonism on inhibition of T cell autoimmunity. The influence of genetic background on the induction of EAE in endocannabinoid system-related gene knockout mice was examined. It was found that C57BL/6.GPR55 knockout mice developed less severe disease, notably in female mice, following active induction with myelin oligodendrocyte glycoprotein 35-55 peptide. In contrast C57BL/6.CB2 (Cnr2 (Dgen receptor knockout mice developed augmented severity of disease consistent with the genetically and pharmacologically-distinct, Cnr2 (tm1Zim mice. However, when the knockout gene was bred into the ABH mouse background and EAE induced with spinal cord autoantigens the immune-enhancing effect of CB2 receptor deletion was lost. Likewise CB1 receptor and transient receptor potential vanilloid one knockout mice on the ABH background demonstrated no alteration in immune-susceptibility, in terms of disease incidence and severity of EAE, in contrast to that reported in some C57BL/6 mouse studies. Furthermore the immune-modulating influence of GPR55 was marginal on the ABH mouse background. Whilst sedative doses of tetrahydrocannabinol could induce immunosuppression, this was associated with a CB1 receptor rather than a CB2 receptor-mediated effect. These data support the fact that non-psychoactive doses of medicinal cannabis have a marginal influence on the immune response in MS. Importantly, it adds a note of caution for the translational

  2. Acutely increasing δGABA(A) receptor activity impairs memory and inhibits synaptic plasticity in the hippocampus.

    Science.gov (United States)

    Whissell, Paul D; Eng, Dave; Lecker, Irene; Martin, Loren J; Wang, Dian-Shi; Orser, Beverley A

    2013-01-01

    Extrasynaptic γ-aminobutyric acid type A (GABA(A)) receptors that contain the δ subunit (δGABA(A) receptors) are expressed in several brain regions including the dentate gyrus (DG) and CA1 subfields of the hippocampus. Drugs that increase δGABA(A) receptor activity have been proposed as treatments for a variety of disorders including insomnia, epilepsy and chronic pain. Also, long-term pretreatment with the δGABA(A) receptor-preferring agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) enhances discrimination memory and increases neurogenesis in the DG. Despite the potential therapeutic benefits of such treatments, the effects of acutely increasing δGABA(A) receptor activity on memory behaviors remain unknown. Here, we studied the effects of THIP (4 mg/kg, i.p.) on memory performance in wild-type (WT) and δGABA(A) receptor null mutant (Gabrd(-/-)) mice. Additionally, the effects of THIP on long-term potentiation (LTP), a molecular correlate of memory, were studied within the DG and CA1 subfields of the hippocampus using electrophysiological recordings of field potentials in hippocampal slices. The results showed that THIP impaired performance in the Morris water maze, contextual fear conditioning and object recognition tasks in WT mice but not Gabrd(-/-) mice. Furthermore, THIP inhibited LTP in hippocampal slices from WT but not Gabrd(-/-) mice, an effect that was blocked by GABA(A) receptor antagonist bicuculline. Thus, acutely increasing δGABA(A) receptor activity impairs memory behaviors and inhibits synaptic plasticity. These results have important implications for the development of therapies aimed at increasing δGABA(A) receptor activity.

  3. Unheated Cannabis sativa extracts and its major compound THC-acid have potential immuno-modulating properties not mediated bu CB1 en CB2 receptor coupled pathways

    NARCIS (Netherlands)

    Verhoeckx, K.C.M.; Korthout, H.A.A.J.; Meeteren-Kreikamp, van A.P.; Ehlert, K.A.; Wang, M.; Greef, de J.; Rodenburg, R.J.T.; Witkamp, R.F.

    2006-01-01

    There is a great interest in the pharmacological properties of cannabinoid like compounds that are not linked to the adverse effects of ¿9-tetrahydrocannabinol (THC), e.g. psychoactive properties. The present paper describes the potential immuno-modulating activity of unheated Cannabis sativa

  4. Unheated Cannabis sativa extracts and its major compound THC-acid have potential immuno-modulating properties not mediated by CB1 and CB2 receptor coupled pathways.

    NARCIS (Netherlands)

    Verhoeckx, K.C.; Korthout, H.A.; Meeteren-Kreikamp, A.P. van; Ehlert, K.A.; Wang, M.; Greef, J. van der; Rodenburg, R.J.T.; Witkamp, R.F.

    2006-01-01

    There is a great interest in the pharmacological properties of cannabinoid like compounds that are not linked to the adverse effects of Delta(9)-tetrahydrocannabinol (THC), e.g. psychoactive properties. The present paper describes the potential immuno-modulating activity of unheated Cannabis sativa

  5. Unheated Cannabis sativa extracts and its major compound THC-acid have potential immuno-modulating properties not mediated by CB1 and CB2 receptor coupled pathways

    NARCIS (Netherlands)

    Verhoeckx, K.C.M.; Korthout, H.A.A.J.; Meeteren-Kreikamp, A.P. van; Ehlert, K.A.; Wang, M.; Greef, J. van der; Rodenburg, R.J.T.; Witkamp, R.F.

    2006-01-01

    There is a great interest in the pharmacological properties of cannabinoid like compounds that are not linked to the adverse effects of Δ9-tetrahydrocannabinol (THC), e.g. psychoactive properties. The present paper describes the potential immuno-modulating activity of unheated Cannabis sativa

  6. Estudo da modelagem molecular do receptor canabinóide CB1 e suas interações com o ∆9 - THC

    OpenAIRE

    Emmanuela Ferreira de Lima

    2009-01-01

    Marihuana (Cannabis sativa) é uma planta amplamente usada pelo ser humano há séculos e suas várias aplicações têm benefícios importantes. A planta Cannabis sativa tem sido usada pelo homem como comida, em práticas medicinais e rituais religiosos. Seus efeitos incluem analgesia, alteração na percepção, cognição, memória e atividade psicomotora. Os compostos canabinoides têm sido usados na quimioterapia do câncer e AIDS. No entanto, o uso da marijuana é um problema devido aos seus efeitos indes...

  7. Effects of the cannabinoid CB1 receptor agonist CP55,940 and antagonist SR141716A on d-amphetamine-induced behaviours in Cebus monkeys

    DEFF Research Database (Denmark)

    Madsen, Morten V; Peacock, Linda; Werge, Thomas

    2006-01-01

    Several clinical studies have shown that alterations in the cannabinoid system in the brain may be associated with schizophrenia. Although evidence points towards an antipsychotic potential for cannabinoid antagonists, experimental studies have shown inconsistent behavioural effects of cannabinoi...

  8. Context-dependent modulation of alphabetagamma and alphabetadelta GABA A receptors by penicillin: implications for phasic and tonic inhibition.

    Science.gov (United States)

    Feng, Hua-Jun; Botzolakis, Emmanuel J; Macdonald, Robert L

    2009-01-01

    Penicillin, an open-channel blocker of GABA(A) receptors, was recently reported to inhibit phasic, but not tonic, currents in hippocampal neurons. To distinguish between isoform-specific and context-dependent modulation as possible explanations for this selectivity, the effects of penicillin were evaluated on recombinant GABA(A) receptors expressed in HEK293T cells. When co-applied with saturating GABA, penicillin decreased peak amplitude, induced rebound, and prolonged deactivation of currents evoked from both synaptic and extrasynaptic receptor isoforms. However, penicillin had isoform-specific effects on the extent of desensitization, reflecting its ability to differentially modulate peak (non-equilibrium) and residual (near-equilibrium) currents. This suggested that the context of activation could determine the apparent sensitivity of a given receptor isoform to penicillin. To test this hypothesis, we explored the ability of penicillin to modulate synaptic and extrasynaptic isoform currents that were activated under more physiologically relevant conditions. Interestingly, while currents evoked from synaptic isoforms under phasic conditions (transient activation by a saturating concentration of GABA) were substantially inhibited by penicillin, currents evoked from extrasynaptic isoforms under tonic conditions (prolonged application by a sub-saturating concentration of GABA) were minimally affected. We therefore concluded that the reported inability of penicillin to modulate tonic currents could not simply be attributed to insensitivity of extrasynaptic receptors, but rather, reflected an inability to modulate these receptors in their native context of activation.

  9. EP3 receptors inhibit antidiuretic-hormone-dependent sodium transport across frog skin epithelium

    DEFF Research Database (Denmark)

    Rytved, Klaus A.; Nielsen, Robert

    1999-01-01

    Antidiuretic hormone; tight epithelium; prostaglandin receptors; sulprostone; misoprostol; cAMP; cellular Ca2+......Antidiuretic hormone; tight epithelium; prostaglandin receptors; sulprostone; misoprostol; cAMP; cellular Ca2+...

  10. Fasitibant chloride, a kinin B2 receptor antagonist, and dexamethasone interact to inhibit carrageenan-induced inflammatory arthritis in rats

    Science.gov (United States)

    Valenti, Claudio; Giuliani, Sandro; Cialdai, Cecilia; Tramontana, Manuela; Maggi, Carlo Alberto

    2012-01-01

    BACKGROUND AND PURPOSE Bradykinin, through the kinin B2 receptor, is involved in inflammatory processes related to arthropathies. B2 receptor antagonists inhibited carrageenan-induced arthritis in rats in synergy with anti-inflammatory steroids. The mechanism(s) underlying this drug interaction was investigated. EXPERIMENTAL APPROACH Drugs inhibiting inflammatory mediators released by carrageenan were injected, alone or in combination, into the knee joint of pentobarbital anaesthetized rats 30 min before intra-articular administration of carrageenan. Their effects on the carrageenan-induced inflammatory responses (joint pain, oedema and neutrophil recruitment) and release of inflammatory mediators (prostaglandins, IL-1β, IL-6 and the chemokine GRO/CINC-1), were assessed after 6 h. KEY RESULTS The combination of fasitibant chloride (MEN16132) and dexamethasone was more effective than each drug administered alone in inhibiting knee joint inflammation and release of inflammatory mediators. Fasitibant chloride, MK571, atenolol, des-Arg9-[Leu8]-bradykinin (B2 receptor, leukotriene, catecholamine and B1 receptor antagonists, respectively) and dexketoprofen (COX inhibitor), reduced joint pain and, except for the latter, also diminished joint oedema. A combination of drugs inhibiting joint pain (fasitibant chloride, des-Arg9-[Leu8]-bradykinin, dexketoprofen, MK571 and atenolol) and oedema (fasitibant chloride, des-Arg9-[Leu8]-bradykinin, MK571 and atenolol) abolished the respective inflammatory response, producing inhibition comparable with that achieved with the combination of fasitibant chloride and dexamethasone. MK571 alone was able to block neutrophil recruitment. CONCLUSIONS AND IMPLICATIONS Bradykinin-mediated inflammatory responses to intra-articular carrageenan were not controlled by steroids, which were not capable of preventing bradykinin effects either by direct activation of the B2 receptor, or through the indirect effects mediated by release of eicosanoids

  11. Inhibition of iodine-125-labeled human follitropin binding to testicular receptor by epidermal growth factor and synthetic peptides

    International Nuclear Information System (INIS)

    Sluss, P.M.; Krystek, S.R. Jr.; Andersen, T.T.; Melson, B.E.; Huston, J.S.; Ridge, R.; Reichert, L.E. Jr.

    1986-01-01

    Two tetrapeptide sequence homologies between mouse epidermal growth factor precursor (mEGFP) and human follitropin (FSH) were revealed by a computer program that identifies identical residues among polypeptide sequences. The two tetrapeptides, Lys-Thr-Cys-Thr (KTCT) and Thr-Arg-Asp-Leu (TRDL), are present in the hormone-specific beta subunit of FSH from all species studied. These tetrapeptides are not present in the alpha subunit, which is common to all pituitary glycoprotein hormones. Both tetrapeptides are also found in mEGFP, and one tetrapeptide, TRDL, is located within the 53-residue form of mEGF purified from mouse submaxillary glands. Computer-generated hydropathy profiles predicted that both tetrapeptides are located in hydrophilic portions of the FSH beta subunit and that TRDL is in a hydrophilic portion of commercially available mEGF. Therefore, the tetrapeptides might be accessible to receptor binding sites for FSH. We report that mEGF inhibits binding of 125 I-labeled human FSH to receptors in testis by 50% (I50) at a concentration of 1.8 X 10(-5) M. No binding inhibition was observed by GnRH or arginine-vasopressin at 10(-4) M, neither of which contain the tetrapeptide sequences. FSH beta subunit, which contains both tetrapeptides, also inhibited binding (I50 = 9 X 10(-8) M) of 125 I-labeled human FSH to testis receptor. Thus, it appears that FSH beta subunit and mEGF are capable of inhibiting binding of FSH to testicular FSH receptors, presumably through interactions that include the homologous tetrapeptides. This presumption was supported by the observation that the synthetic tetrapeptides (KTCT or TRDL) were also active in inhibiting binding of 125 I-labeled human FSH to testis receptor

  12. Hyperosmotic stress inhibits insulin receptor substrate-1 function by distinct mechanisms in 3T3-L1 adipocytes

    DEFF Research Database (Denmark)

    Gual, Philippe; Gonzalez, Teresa; Grémeaux, Thierry

    2003-01-01

    . Furthermore, the mammalian target of rapamycin (mTOR) inhibitor rapamycin prevented the osmotic shock-induced phosphorylation of IRS-1 on Ser307. The inhibition of mTOR completely reversed the inhibitory effect of hyperosmotic stress on insulin-induced IRS-1 tyrosine phosphorylation and PI 3-kinase activation......In 3T3-L1 adipocytes, hyperosmotic stress was found to inhibit insulin signaling, leading to an insulin-resistant state. We show here that, despite normal activation of insulin receptor, hyperosmotic stress inhibits both tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) and IRS-1....... In addition, prolonged osmotic stress enhanced the degradation of IRS proteins through a rapamycin-insensitive pathway and a proteasome-independent process. These data support evidence of new mechanisms involved in osmotic stress-induced cellular insulin resistance. Short-term osmotic stress induces...

  13. Ketamine-induced inhibition of glycogen synthase kinase-3 contributes to the augmentation of AMPA receptor signaling

    Science.gov (United States)

    Beurel, Eléonore; Grieco, Steven F; Amadei, Celeste; Downey, Kimberlee; Jope, Richard S

    2016-01-01

    Objectives Sub-anesthetic doses of ketamine have been found to provide rapid antidepressant actions, indicating that the cellular signaling systems targeted by ketamine are potential sites for therapeutic intervention. Ketamine acts as an antagonist of N-methyl-D-aspartate (NMDA) receptors, and animal studies indicate that subsequent augmentation of signaling by α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors is critical for the antidepressant outcome. Methods In this study, we tested if the inhibitory effect of ketamine on glycogen synthase kinase-3 (GSK3) affected hippocampal cell-surface AMPA receptors using immunoblotting of membrane and synaptosomal extracts from wild-type and GSK3 knockin mice. Results Treatment with an antidepressant dose of ketamine increased the hippocampal membrane level of the AMPA glutamate receptor (GluA)1 subunit, but did not alter the localization of GluA2, GluA3, or GluA4. This effect of ketamine was abrogated in GSK3 knockin mice expressing mutant GSK3 that cannot be inhibited by ketamine, demonstrating that ketamine-induced inhibition of GSK3 is necessary for up-regulation of cell surface AMPA GluA1 subunits. AMPA receptor trafficking is regulated by post-synaptic density-95 (PSD-95), a substrate for GSK3. Ketamine treatment decreased the hippocampal membrane level of phosphorylated PSD-95 on Thr-19, the target of GSK3 that promotes AMPA receptor internalization. Conclusions These results demonstrate that ketamine-induced inhibition of GSK3 causes reduced phosphorylation of PSD-95, diminishing the internalization of AMPA GluA1 subunits to allow for augmented signaling through AMPA receptors following ketamine treatment. PMID:27687706

  14. Antinociceptive Action of Isolated Mitragynine from Mitragyna Speciosa through Activation of Opioid Receptor System

    Directory of Open Access Journals (Sweden)

    Mohamad Aris Mohd Moklas

    2012-09-01

    Full Text Available Cannabinoids and opioids systems share numerous pharmacological properties and antinociception is one of them. Previous findings have shown that mitragynine (MG, a major indole alkaloid found in Mitragyna speciosa (MS can exert its antinociceptive effects through the opioids system. In the present study, the action of MG was investigated as the antinociceptive agent acting on Cannabinoid receptor type 1 (CB1 and effects on the opioids receptor. The latency time was recorded until the mice showed pain responses such as shaking, licking or jumping and the duration of latency was measured for 2 h at every 15 min interval by hot plate analysis. To investigate the beneficial effects of MG as antinociceptive agent, it was administered intraperitoneally 15 min prior to pain induction with a single dosage (3, 10, 15, 30, and 35 mg/kg b.wt. In this investigation, 35 mg/kg of MG showed significant increase in the latency time and this dosage was used in the antagonist receptor study. The treated groups were administered with AM251 (cannabinoid receptor-1 antagonist, naloxone (non-selective opioid antagonist, naltrindole (δ-opioid antagonist naloxonazine (µ1-receptor antagonist and norbinaltorpimine (κ-opioid antagonist respectively, prior to administration of MG (35 mg/kg. The results showed that the antinociceptive effect of MG was not antagonized by AM251; naloxone and naltrindole were effectively blocked; and norbinaltorpimine partially blocked the antinociceptive effect of MG. Naloxonazine did inhibit the effect of MG, but it was not statistically significant. These results demonstrate that CB1 does not directly have a role in the antinociceptive action of MG where the effect was observed with the activation of opioid receptor.

  15. PTP1B Inhibition Causes Rac1 Activation by Enhancing Receptor Tyrosine Kinase Signaling

    Directory of Open Access Journals (Sweden)

    Ayako Tsuchiya

    2014-04-01

    Full Text Available Background/Aims: The present study investigated the signaling pathway underlying Rac1 activation induced by the linoleic acid derivative 8-[2-(2-pentyl-cyclopropylmethyl-cyclopropyl]-octanoic acid (DCP-LA. Methods: Activity of protein tyrosine phosphatase 1B (PTP1B was assayed under cell-free conditions. Western blot was carried out to quantify phosphorylation of insulin receptor substrate-1 (IRS-1 and Akt in PC-12 cells. Rac1 activity was monitored in the föerster resonance energy transfer (FRET analysis using living and fixed PC-12 cells. Results: DCP-LA markedly suppressed PTP1B activity in a concentration (100 pM-100 µM-dependent manner. In the DCP-LA binding assay, fluorescein-conjugated DCP-LA produced a single fluorescent signal band at 60 kDa, corresponding to the molecule of PTP1B, and the signal was attenuated or abolished by co-treatment or pretreatment with non-conjugated DCP-LA. DCP-LA significantly enhanced nerve growth factor (NGF-stimulated phosphorylation of IRS-1 at Tyr1222 and Akt1/2 at Thr308/309 and Ser473/474 in PC-12 cells. In the FRET analysis, DCP-LA significantly enhanced NGF-stimulated Rac1 activation, which is abrogated by the phosphatidylinositol 3 kinase (PI3K inhibitor wortmannin, the 3-phosphoinositide-dependent protein kinase-1 (PDK1 inhibitor BX912, or the Akt inhibitor MK2206. Conclusion: The results of the present study show that DCP-LA-induced PTP1B inhibition, possibly through its direct binding, causes Rac1 activation by enhancing a pathway along a receptor tyrosine kinase (RTK/IRS-1/PI3K/Akt/Rac1 axis.

  16. Targeting non-small cell lung cancer cells by dual inhibition of the insulin receptor and the insulin-like growth factor-1 receptor.

    Directory of Open Access Journals (Sweden)

    Emma E Vincent

    Full Text Available Phase III trials of the anti-insulin-like growth factor-1 receptor (IGF1R antibody figitumumab in non-small cell lung cancer (NSCLC patients have been discontinued owing to lack of survival benefit. We investigated whether inhibition of the highly homologous insulin receptor (IR in addition to the IGF1R would be more effective than inhibition of the IGF1R alone at preventing the proliferation of NSCLC cells. Signalling through IGF1R and IR in the NSCLC cell lines A549 and Hcc193 was stimulated by a combination of IGF1, IGF2 and insulin. It was inhibited by antibodies that block ligand binding, αIR3 (IGF1R and IR47-9 (IR, and by the ATP-competitive small molecule tyrosine kinase inhibitors AZ12253801 and NVPAWD742 which inhibit both IGF1R and IR tyrosine kinases. The effect of inhibitors was determined by an anchorage-independent proliferation assay and by analysis of Akt phosphorylation. In Hcc193 cells the reduction in cell proliferation and Akt phosphorylation due to anti-IGF1R antibody was enhanced by antibody-mediated inhibition of the IR whereas in A549 cells, with a relatively low IR:IGF1R expression ratio, it was not. In each cell line proliferation and Akt phosphorylation were more effectively inhibited by AZ12253801 and NVPAWD742 than by combined αIR3 and IR47-9. When the IGF1R alone is inhibited, unencumbered signalling through the IR can contribute to continued NSCLC cell proliferation. We conclude that small molecule inhibitors targeting both the IR and IGF1R more effectively reduce NSCLC cell proliferation in a manner independent of the IR:IGF1R expression ratio, providing a therapeutic rationale for the treatment of this disease.

  17. Dopamine inhibits reproduction in female zebrafish (Danio rerio) via three pituitary D2 receptor subtypes.

    Science.gov (United States)

    Fontaine, Romain; Affaticati, Pierre; Yamamoto, Kei; Jolly, Cécile; Bureau, Charlotte; Baloche, Sylvie; Gonnet, Françoise; Vernier, Philippe; Dufour, Sylvie; Pasqualini, Catherine

    2013-02-01

    In many teleosts, the stimulatory control of gonadotrope axis by GnRH is opposed by an inhibitory control by dopamine (DA). The functional importance of this inhibitory pathway differs widely from one teleostean species to another. The zebrafish (Danio rerio) is a teleost fish that has become increasingly popular as an experimental vertebrate model. However, the role of DA in the neuroendocrine control of its reproduction has never been studied. Here the authors evaluated in sexually regressed female zebrafish the effects of in vivo treatments with a DA D2 receptor (D2-R) antagonist domperidone, or a GnRH agonist, alone and in combination, on the pituitary level of FSHβ and LHβ transcripts, the gonadosomatic index, and the ovarian histology. Only the double treatment with GnRH agonist and domperidone could induce an increase in the expression of LHβ, in the gonadosomatic index, and a stimulation of ovarian vitellogenesis, indicating that removal of dopaminergic inhibition is required for the stimulatory action of GnRH and reactivation of ovarian function to occur. Using double immunofluorescent staining on pituitary, the authors showed in this species the innervation of LH cells by tyrosine-hydroxylase immunoreactive fibers. Finally, using in situ hybridization and immunofluorescence, the authors showed that the three subtypes of zebrafish DA D2-R (D2a, D2b, and D2c) were expressed in LH-producing cells, suggesting that they all may be involved in mediating this inhibition. These results show for the first time that, in zebrafish, DA has a direct and potent inhibitory action capable of opposing the stimulatory effect of GnRH in the neuroendocrine control of reproduction.

  18. Nifedipine inhibits advanced glycation end products (AGEs) and their receptor (RAGE) interaction-mediated proximal tubular cell injury via peroxisome proliferator-activated receptor-gamma activation

    International Nuclear Information System (INIS)

    Matsui, Takanori; Yamagishi, Sho-ichi; Takeuchi, Masayoshi; Ueda, Seiji; Fukami, Kei; Okuda, Seiya

    2010-01-01

    Research highlights: → Nifedipine inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by GW9662, an inhibitor of peroxisome proliferator-activated receptor-γ. → GW9662 treatment alone increased RAGE mRNA levels in tubular cells. → Nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-κB activation and increases in intercellular adhesion molecule-1 and transforming growth factor-β gene expression in tubular cells, all of which were blocked by GW9662. -- Abstract: There is a growing body of evidence that advanced glycation end products (AGEs) and their receptor (RAGE) interaction evokes oxidative stress generation and subsequently elicits inflammatory and fibrogenic reactions, thereby contributing to the development and progression of diabetic nephropathy. We have previously found that nifedipine, a calcium-channel blocker (CCB), inhibits the AGE-induced mesangial cell damage in vitro. However, effects of nifedipine on proximal tubular cell injury remain unknown. We examined here whether and how nifedipine blocked the AGE-induced tubular cell damage. Nifedipine, but not amlodipine, a control CCB, inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-γ (PPARγ). GW9662 treatment alone was found to increase RAGE mRNA levels in tubular cells. Further, nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-κB activation and increases in intercellular adhesion molecule-1 and transforming growth factor-beta gene expression in tubular cells, all of which were blocked by GW9662. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-oxidative and anti-inflammatory agent against AGEs in tubular cells by suppressing RAGE expression via PPARγ activation.

  19. Nifedipine inhibits advanced glycation end products (AGEs) and their receptor (RAGE) interaction-mediated proximal tubular cell injury via peroxisome proliferator-activated receptor-gamma activation

    Energy Technology Data Exchange (ETDEWEB)

    Matsui, Takanori [Department of Pathophysiology and Therapeutics of Diabetic Vascular Complications, Kurume University School of Medicine, Kurume (Japan); Yamagishi, Sho-ichi, E-mail: shoichi@med.kurume-u.ac.jp [Department of Pathophysiology and Therapeutics of Diabetic Vascular Complications, Kurume University School of Medicine, Kurume (Japan); Takeuchi, Masayoshi [Department of Pathophysiological Science, Faculty of Pharmaceutical Science, Hokuriku University, Kanazawa (Japan); Ueda, Seiji; Fukami, Kei; Okuda, Seiya [Department of Medicine, Kurume University School of Medicine, Kurume (Japan)

    2010-07-23

    Research highlights: {yields} Nifedipine inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma}. {yields} GW9662 treatment alone increased RAGE mRNA levels in tubular cells. {yields} Nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-{kappa}B activation and increases in intercellular adhesion molecule-1 and transforming growth factor-{beta} gene expression in tubular cells, all of which were blocked by GW9662. -- Abstract: There is a growing body of evidence that advanced glycation end products (AGEs) and their receptor (RAGE) interaction evokes oxidative stress generation and subsequently elicits inflammatory and fibrogenic reactions, thereby contributing to the development and progression of diabetic nephropathy. We have previously found that nifedipine, a calcium-channel blocker (CCB), inhibits the AGE-induced mesangial cell damage in vitro. However, effects of nifedipine on proximal tubular cell injury remain unknown. We examined here whether and how nifedipine blocked the AGE-induced tubular cell damage. Nifedipine, but not amlodipine, a control CCB, inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}). GW9662 treatment alone was found to increase RAGE mRNA levels in tubular cells. Further, nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-{kappa}B activation and increases in intercellular adhesion molecule-1 and transforming growth factor-beta gene expression in tubular cells, all of which were blocked by GW9662. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-oxidative and anti-inflammatory agent against AGEs in tubular cells by suppressing RAGE expression

  20. Plant lectin can target receptors containing sialic acid, exemplified by podoplanin, to inhibit transformed cell growth and migration.

    Directory of Open Access Journals (Sweden)

    Jhon Alberto Ochoa-Alvarez

    Full Text Available Cancer is a leading cause of death of men and women worldwide. Tumor cell motility contributes to metastatic invasion that causes the vast majority of cancer deaths. Extracellular receptors modified by α2,3-sialic acids that promote this motility can serve as ideal chemotherapeutic targets. For example, the extracellular domain of the mucin receptor podoplanin (PDPN is highly O-glycosylated with α2,3-sialic acid linked to galactose. PDPN is activated by endogenous ligands to induce tumor cell motility and metastasis. Dietary lectins that target proteins containing α2,3-sialic acid inhibit tumor cell growth. However, anti-cancer lectins that have been examined thus far target receptors that have not been identified. We report here that a lectin from the seeds of Maackia amurensis (MASL with affinity for O-linked carbohydrate chains containing sialic acid targets PDPN to inhibit transformed cell growth and motility at nanomolar concentrations. Interestingly, the biological activity of this lectin survives gastrointestinal proteolysis and enters the cardiovascular system to inhibit melanoma cell growth, migration, and tumorigenesis. These studies demonstrate how lectins may be used to help develop dietary agents that target specific receptors to combat malignant cell growth.

  1. Midazolam inhibits hippocampal long-term potentiation and learning through dual central and peripheral benzodiazepine receptor activation and neurosteroidogenesis.

    Science.gov (United States)

    Tokuda, Kazuhiro; O'Dell, Kazuko A; Izumi, Yukitoshi; Zorumski, Charles F

    2010-12-15

    Benzodiazepines (BDZs) enhance GABA(A) receptor inhibition by direct actions on central BDZ receptors (CBRs). Although some BDZs also bind mitochondrial receptors [translocator protein (18 kDa) (TSPO)] and promote the synthesis of GABA-enhancing neurosteroids, the role of neurosteroids in the clinical effects of BDZs is unknown. In rat hippocampal slices, we compared midazolam, an anesthetic BDZ, with clonazepam, an anticonvulsant/anxiolytic BDZ that activates CBRs selectively. Midazolam, but not clonazepam, increased neurosteroid levels in CA1 pyramidal neurons without changing TSPO immunostaining. Midazolam, but not clonazepam, also augmented a form of spike inhibition after stimulation adjacent to the pyramidal cell layer and inhibited induction of long-term potentiation. These effects were prevented by finasteride, an inhibitor of neurosteroid synthesis, or 17PA [17-phenyl-(3α,5α)-androst-16-en-3-ol], a blocker of neurosteroid effects on GABA(A) receptors. Moreover, the synaptic effects were mimicked by a combination of clonazepam with FGIN (2-[2-(4-fluorophenyl)-1H-indol-3-yl]-N,N-dihexylacetamide), a selective TSPO agonist, or a combination of clonazepam with exogenous allopregnanolone. Consistent with these in vitro results, finasteride abolished the effects of midazolam on contextual fear learning when administrated 1 d before midazolam injection. Thus, dual activation of CBRs and TSPO appears to result in unique actions of clinically important BDZs. Furthermore, endogenous neurosteroids are shown to be important regulators of pyramidal neuron function and synaptic plasticity.

  2. Adenosine inhibits neutrophil vascular endothelial growth factor release and transendothelial migration via A2B receptor activation.

    LENUS (Irish Health Repository)

    Wakai, A

    2012-02-03

    The effects of adenosine on neutrophil (polymorphonuclear neutrophils; PMN)-directed changes in vascular permeability are poorly characterized. This study investigated whether adenosine modulates activated PMN vascular endothelial growth factor (vascular permeability factor; VEGF) release and transendothelial migration. PMN activated with tumour necrosis factor-alpha (TNF-alpha, 10 ng\\/mL) were incubated with adenosine and its receptor-specific analogues. Culture supernatants were assayed for VEGF. PMN transendothelial migration across human umbilical vein endothelial cell (HUVEC) monolayers was assessed in vitro. Adhesion molecule receptor expression was assessed flow cytometrically. Adenosine and some of its receptor-specific analogues dose-dependently inhibited activated PMN VEGF release. The rank order of potency was consistent with the affinity profile of human A2B receptors. The inhibitory effect of adenosine was reversed by 3,7-dimethyl-1-propargylxanthine, an A2 receptor antagonist. Adenosine (100 microM) or the A2B receptor agonist 5\\'-N-ethylcarboxamidoadenosine (NECA, 100 microM) significantly reduced PMN transendothelial migration. However, expression of activated PMN beta2 integrins and HUVEC ICAM-1 were not significantly altered by adenosine or NECA. Adenosine attenuates human PMN VEGF release and transendothelial migration via the A2B receptor. This provides a novel target for the modulation of PMN-directed vascular hyperpermeability in conditions such as the capillary leak syndrome.

  3. Inhibition of Androgen Receptor Nuclear Localization and Castration-Resistant Prostate Tumor Growth by Pyrroloimidazole-based Small Molecules.

    Science.gov (United States)

    Masoodi, Khalid Z; Xu, Yadong; Dar, Javid A; Eisermann, Kurtis; Pascal, Laura E; Parrinello, Erica; Ai, Junkui; Johnston, Paul A; Nelson, Joel B; Wipf, Peter; Wang, Zhou

    2017-10-01

    The androgen receptor (AR) is a ligand-dependent transcription factor that controls the expression of androgen-responsive genes. A key step in androgen action, which is amplified in castration-resistant prostate cancer (CRPC), is AR nuclear translocation. Small molecules capable of inhibiting AR nuclear localization could be developed as novel therapeutics for CRPC. We developed a high-throughput screen and identified two structurally-related pyrroloimidazoles that could block AR nuclear localization in CRPC cells. We show that these two small molecules, 3-(4-ethoxyphenyl)-6,7-dihydro-5 H -pyrrolo[1,2- a ]imidazole (EPPI) and 3-(4-chlorophenyl)-6,7-dihydro-5 H -pyrrolo[1,2- a ]imidazole (CPPI) can inhibit the nuclear localization and transcriptional activity of AR and reduce the proliferation of AR-positive but not AR-negative prostate cancer cell lines. EPPI and CPPI did not inhibit nuclear localization of the glucocorticoid receptor or the estrogen receptor, suggesting they selectively target AR. In LNCaP tumor xenografts, CPPI inhibited the proliferation of relapsed LNCaP tumors. These findings suggest that EPPI and CPPI could serve as lead structures for the development of therapeutic agents for CRPC. Mol Cancer Ther; 16(10); 2120-9. ©2017 AACR . ©2017 American Association for Cancer Research.

  4. Pu-erh Tea Protects the Nervous System by Inhibiting the Expression of Metabotropic Glutamate Receptor 5.

    Science.gov (United States)

    Li, Chunjie; Chai, Shaomeng; Ju, Yongzhi; Hou, Lu; Zhao, Hang; Ma, Wei; Li, Tian; Sheng, Jun; Shi, Wei

    2017-09-01

    Glutamate is one of the major excitatory neurotransmitters of the CNS and is essential for numerous key neuronal functions. However, excess glutamate causes massive neuronal death and brain damage owing to excitotoxicity via the glutamate receptors. Metabotropic glutamate receptor 5 (mGluR5) is one of the glutamate receptors and represents a promising target for studying neuroprotective agents of potential application in neurodegenerative diseases. Pu-erh tea, a fermented tea, mainly produced in Yunnan province, China, has beneficial effects, including the accommodation of the CNS. In this study, pu-erh tea markedly decreased the transcription and translation of mGluR5 compared to those by black and green teas. Pu-erh tea also inhibited the expression of Homer, one of the synaptic scaffolding proteins binding to mGluR5. Pu-erh tea protected neural cells from necrosis via blocked Ca 2+ influx and inhibited protein kinase C (PKC) activation induced by excess glutamate. Pu-erh tea relieved rat epilepsy induced by LiCl-pilocarpine in behavioural and physiological assays. Pu-erh tea also decreased the expression of mGluR5 in the hippocampus. These results show that the inhibition of mGluR5 plays a role in protecting neural cells from glutamate. The results also indicate that pu-erh tea contains biological compounds binding transcription factors and inhibiting the expression of mGluR5 and identify pu-erh tea as a novel natural neuroprotective agent.

  5. Effect of Delta-tetrahydrocannabinol, a cannabinoid receptor agonist, on the triggering of transient lower oesophageal sphincter relaxations in dogs and humans

    NARCIS (Netherlands)

    Beaumont, H.; Jensen, J.; Carlsson, A.; Ruth, M.; Lehmann, A.; Boeckxstaens, G. E.

    2009-01-01

    Background and purpose: Transient lower oesophageal sphincter relaxations (TLESRs) are the main mechanism underlying gastro-oesophageal reflux and are a potential pharmacological treatment target. We evaluated the effect of the CB(1)/CB(2) receptor agonist Delta(9)-tetrahydrocannabinol

  6. The insecticide fipronil and its metabolite fipronil sulphone inhibit the rat α1β2γ2L GABAA receptor

    Science.gov (United States)

    Li, P; Akk, G

    2008-01-01

    Background and purpose: Fipronil is the active ingredient in a number of widely used insecticides. Human exposure to fipronil leads to symptoms (headache, nausea and seizures) typically associated with the antagonism of GABAA receptors in the brain. In this study, we have examined the modulation of the common brain GABAA receptor subtype by fipronil and its major metabolite, fipronil sulphone. Experimental approach: Whole-cell and single-channel recordings were made from HEK 293 cells transiently expressing rat α1β2γ2L GABAA receptors. Key results: The major effect of fipronil was to increase the rate of current decay in macroscopic recordings. In single-channel recordings, the presence of fipronil resulted in shorter cluster durations without affecting the intracluster open and closed time distributions or the single-channel conductance. The α1V256S mutation, previously shown alleviate channel inhibition by inhibitory steroids and several insecticides, had a relatively small effect on channel block by fipronil. The mode of action of fipronil sulphone was similar to that of its parent compound but the metabolite was less potent at inhibiting the α1β2γ2L receptor. Conclusions and implications: We conclude that exposure to fipronil induces accumulation of receptors in a novel, long-lived blocked state. This process proceeds in parallel with and independently of, channel desensitization. The lower potency of fipronil sulphone indicates that the conversion serves as a detoxifying process in mammalian brain. PMID:18660823

  7. E-cadherin homophilic ligation inhibits cell growth and epidermal growth factor receptor signaling independently of other cell interactions

    DEFF Research Database (Denmark)

    Perrais, Michaël; Chen, Xiao; Perez-Moreno, Mirna

    2007-01-01

    growth inhibitory signals. To address this question, we have selectively formed E-cadherin homophilic bonds at the cell surface of isolated epithelial cells by using functionally active recombinant E-cadherin protein attached to microspheres. We find that E-cadherin ligation alone reduces the frequency...... of cells entering the S phase, demonstrating that E-cadherin ligation directly transduces growth inhibitory signals. E-cadherin binding to beta-catenin is required for cell growth inhibition, but beta-catenin/T-cell factor transcriptional activity is not involved in growth inhibition resulting from...... homophilic binding. Neither E-cadherin binding to p120-catenin nor beta-catenin binding to alpha-catenin, and thereby the actin cytoskeleton, is required for growth inhibition. E-cadherin ligation also inhibits epidermal growth factor (EGF) receptor-mediated growth signaling by a beta...

  8. Arctigenin antagonizes mineralocorticoid receptor to inhibit the transcription of Na/K-ATPase.

    Science.gov (United States)

    Cheng, Ye; Zhou, Meili; Wang, Yan

    2016-01-01

    Hypertension is one of the most important risk factors in cardiovascular disease and is the most common chronic disease. Mineralocorticoid receptor (MR) antagonists have been successfully used in clinic for the treatment of hypertension. Our study aims to investigate whether Arctigenin can antagonize MR and inhibit the transcription of Na/K-ATPase. The yeast two-hybrid assay was used to screen natural products and Arctigenin was identified as an MR antagonist. The direct binding of Arctigenin to MR was determined using assays based on surface plasmon resonance, differential scanning calorimetry and fluorescence quenching. Furthermore, results from mammalian one-hybrid and transcriptional activation experiments also confirmed that Arctigenin can potently antagonize MR in cells. We demonstrated that Arctigenin can decrease the level of Na/K-ATPase mRNA by antagonizing MR in HK-2 cells. Our findings show that Arctigenin can effectively decrease Na/K-ATPase transcription; thus highlight its potential as an anti-hypertensive drug lead compound. Our current findings demonstrate that Arctigenin is an antagonist of MR and effectively decreases the Na/K-ATPase 1 gene expression. Our work provides a hint for the drug discovery against cardiovascular disease.

  9. Nanoparticles containing a liver X receptor agonist inhibit inflammation and atherosclerosis.

    Science.gov (United States)

    Zhang, Xue-Qing; Even-Or, Orli; Xu, Xiaoyang; van Rosmalen, Mariska; Lim, Lucas; Gadde, Suresh; Farokhzad, Omid C; Fisher, Edward A

    2015-01-28

    Liver X receptor (LXR) signaling pathways regulate lipid metabolism and inflammation, which has generated widespread interest in developing synthetic LXR agonists as potential therapeutics for the management of atherosclerosis. In this study, it is demonstrated that nanoparticles (NPs) containing the synthetic LXR agonist GW3965 (NP-LXR) exert anti-inflammatory effects and inhibit the development of atherosclerosis without causing hepatic steatosis. These NPs are engineered through self-assembly of a biodegradable diblock poly(lactide-co-glycolide)-b-poly(ethylene glycol) (PLGA-b-PEG) copolymer. NP-LXR is significantly more effective than free GW3965 at inducing LXR-target gene expression and suppressing inflammatory factors in macrophages in vitro and in vivo. Additionally, the NPs elicit negligible lipogenic gene stimulation in the liver. Using the Ldlr (-/-) mouse model of atherosclerosis, abundant colocalization of fluorescently labeled NPs within plaque macrophages following systemic administration is seen. Notably, six intravenous injections of NP-LXR over 2 weeks markedly reduce the CD68-positive cell (macrophage) content of plaques (by 50%) without increasing total cholesterol or triglycerides in the liver and plasma. Together, these findings identify GW3965-encapsulated PLGA-b-PEG NPs as a promising nanotherapeutic approach to combat atherosclerosis, providing the benefits of LXR agonists without their adverse effects on hepatic and plasma lipid metabolism. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. A Novel Dietary Flavonoid Fisetin Inhibits Androgen Receptor Signaling and Tumor Growth in Athymic Nude Mice

    Science.gov (United States)

    Khan, Naghma; Asim, Mohammad; Afaq, Farrukh; Zaid, Mohammad Abu; Mukhtar, Hasan

    2010-01-01

    Androgen receptor (AR)–mediated signaling plays an important role in the development and progression of prostate cancer (PCa). Hormonal therapies, mainly with combinations of antiandrogens and androgen deprivation, are the mainstay treatment for advanced disease. However, emergence of androgen resistance largely due to inefficient antihormone action limits their therapeutic usefulness. Here, we report that fisetin, a novel dietary flavonoid, acts as a novel AR ligand by competing with the high-affinity androgen to interact with the ligand binding domain of AR. We show that this physical interaction results in substantial decrease in AR stability and decrease in amino-terminal/carboxyl-terminal (N-C) interaction of AR. This results in blunting of AR-mediated transactivation of target genes including prostate-specific antigen (PSA). In addition, treatment of LNCaP cells with fisetin decreased AR protein levels, in part, by decreasing its promoter activity and by accelerating its degradation. Fisetin also synergized with Casodex in inducing apoptosis in LNCaP cells. Treatment with fisetin in athymic nude mice implanted with AR-positive CWR22Rυ1 human PCa cells resulted in inhibition of tumor growth and reduction in serum PSA levels. These data identify fisetin as an inhibitor of AR signaling axis and suggest that it could be a useful chemopreventive and chemotherapeutic agent to delay progression of PCa. PMID:18922931

  11. Activation of adenosine receptors and inhibition of cyclooxygenases: two recent pharmacological approaches to modulation of radiation suppressed hematopoiesis

    International Nuclear Information System (INIS)

    Hofer, M.; Pospisil, M.; Vacek, A.; Hola, J.; Weiterova, L.; Streitova, D.; Znojil, V.

    2008-01-01

    Searching for drugs conforming to requirements for protection and/or treatment of radiation-induced damage belongs to the most important tasks of current radiobiology. In the Laboratory of Experimental Hematology, Institute of Biophysics, v.v.i., Academy of Sciences of the Czech Republic, Brno, Czech Republic, two original approaches for stimulation of radiation-suppressed hematopoiesis have been tested in recent years, namely activation of adenosine receptors and inhibition of cyclooxygenases. Non-selective activation of adenosine receptors, induced by combined administration of dipyridamole, a drug preventing adenosine uptake and supporting thus its extracellular receptor-mediated action, and adenosine monophosphate, an adenosine prodrug, has been found to stimulate hematopoiesis when the drugs were given either pre- or post-irradiation. When synthetic adenosine receptor agonists selective for individual adenosine receptor subtypes were tested, stimulatory effects in myelosuppressed mice have been found after administration of IB-MECA, a selective adenosine A3 receptor agonist. Non-selective cyclooxygenase inhibitors, inhibiting both cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2), indomethacin, diclofenac, or flurbiprofen, have been observed to act positively on radiation-perturbed hematopoiesis in sublethally irradiated mice. However, their undesirable gastrointestinal side effects have been found to negatively influence survival of lethally irradiated animals. Recently tested selective COX-2 inhibitor meloxicam, preserving protective action of COX-1-synthesized prostaglandins in the gastrointestinal tissues, has been observed to retain the hematopoiesis-stimulating effects of non-selective cyclooxygenase inhibitors and to improve the survival of animals exposed to lethal radiation doses. These findings bear evidence for the possibility to use selective adenosine A3 receptor agonists and selective COX-2 inhibitors in human practice for treatment of

  12. Ergopeptines bromocriptine and ergovaline and the dopamine type-2 receptor inhibitor domperidone inhibit bovine equilibrative nucleoside transporter 1-like activity.

    Science.gov (United States)

    Miles, Edwena D; Xue, Yan; Strickland, James R; Boling, James A; Matthews, James C

    2011-09-14

    Neotyphodium coenophialum-infected tall fescue contains ergopeptines. Except for interactions with biogenic amine receptors (e.g., dopamine type-2 receptor, D2R), little is known about how ergopeptines affect animal metabolism. The effect of ergopeptines on bovine nucleoside transporters (NT) was evaluated using Madin-Darby bovine kidney (MDBK) cells. Equilibrative NT1 (ENT1)-like activity accounted for 94% of total NT activity. Inhibitory competition (IC(50)) experiments found that this activity was inhibited by both bromocriptine (a synthetic model ergopeptine and D2R agonist) and ergovaline (a predominant ergopeptine of tall fescue). Kinetic inhibition analysis indicated that bromocriptine inhibited ENT1-like activity through a competitive and noncompetitive mechanism. Domperidone (a D2R antagonist) inhibited ENT1 activity more in the presence than in the absence of bromocriptine and displayed an IC(50) value lower than that of bromocriptine or ergovaline, suggesting that inhibition was not through D2R-mediated events. These novel mechanistic findings imply that cattle consuming endophyte-infected tall fescue have reduced ENT1 activity and, thus, impaired nucleoside metabolism.

  13. In vitro nuclear receptor inhibition and cytotoxicity of hydraulic fracturing chemicals and their binary mixtures.

    Science.gov (United States)

    Bain, Peter A; Kumar, Anu

    2018-05-01

    The widespread use of hydraulic fracturing (HF) in oil and gas extraction operations has led to concern over environmental risks posed by chemicals used in HF fluids. Here we employed a suite of stable luciferase reporter gene assays to investigate the potential for selected HF chemicals or geogenics to activate or antagonise nuclear receptor signalling. We screened three biocides (bronopol [BP], glutaraldehyde [GA], and tetrakis(hydroxymethyl)phosphonium sulfate [THPS]), a surfactant (2-butoxyethanol), a friction reducer (polyacrylamide), and a coal seam geogenic (o-cresol) for their potential to act as agonists or antagonists of the estrogen receptor, androgen receptor, progesterone receptor (PR), glucocorticoid receptor or peroxisome proliferator-activated receptor gamma (PPARγ). None of the chemicals induced luciferase activity in any of assays used in the study. In antagonistic mode, BP, GA and THPS caused reductions in luciferase activity in the reporter assays at higher concentrations (50-100 μM), while at low concentrations (2-10 μM) GA and THPS enhanced luciferase activity in some assays relative to controls. None of the other tested chemicals exhibited antagonism in the selected assays. In most cases, altered receptor signalling only occurred at concentrations exhibiting cytotoxicity. However, PPARγ activity, and to a lesser extent PR activity, were inhibited by THPS at sub-cytotoxic concentrations. The majority of binary combinations tested exhibited significantly less-than-additive cytotoxicity, and none of the combinations exhibited synergistic cytotoxicity. In summary, the results of the present study indicate that the selected chemicals are not likely to function as direct agonists of the nuclear receptors tested, and only one chemical, THPS was an apparent partial antagonist of two nuclear receptors. Copyright © 2017. Published by Elsevier Ltd.

  14. Mechanisms of Inhibition of the Epidermal Growth Factor Receptor: Implications for Novel Anti-Cancer Therapies

    National Research Council Canada - National Science Library

    Klein, Daryl E

    2005-01-01

    .... No secreted or extracellular ErbB receptor inhibitors have been reported in mammals. However, two natural inhibitors of the highly homologous Drosophila EGF receptor are found in Drosophila melanogaster...

  15. Presynaptic G Protein-Coupled Receptors: Gatekeepers of Addiction?

    Directory of Open Access Journals (Sweden)

    Kari A Johnson

    2016-11-01

    Full Text Available Drug abuse and addiction cause widespread social and public health problems, and the neurobiology underlying drug actions and drug use and abuse is an area of intensive research. Drugs of abuse alter synaptic transmission, and these actions contribute to acute intoxication as well as the chronic effects of abused substances. Transmission at most mammalian synapses involves neurotransmitter activation of two receptor subtypes, ligand-gated ion channels that mediate fast synaptic responses, and G protein-coupled receptors (GPCRs that have slower neuromodulatory actions. The GPCRs represent a large proportion of neurotransmitter receptors involved in almost all facets of nervous system function. In addition, these receptors are targets for many pharmacotherapeutic agents. Drugs of abuse directly or indirectly affect neuromodulation mediated by GPCRs, with important consequences for intoxication, drug taking and responses to prolonged drug exposure, withdrawal and addiction. Among the GPCRs are several subtypes involved in presynaptic inhibition, most of which are coupled to the Gi/o class of G protein. There is increasing evidence that these presynaptic Gi/o-coupled GPCRs have important roles in the actions of drugs of abuse, as well as behaviors related to these drugs. This topic will be reviewed, with particular emphasis on receptors for three neurotransmitters, dopamine (D1- and D2-like receptors, endocannabinoids (CB1 receptors and glutamate (group II metabotropic glutamate (mGlu receptors. The focus is on recent evidence from laboratory animal models (and some evidence in humans implicating these receptors in the acute and chronic effects of numerous abused drugs, as well as in the control of drug seeking and taking. The ability of drugs targeting these receptors to modify drug seeking behavior has raised the possibility of using compounds targeting these receptors for addiction pharmacotherapy. This topic is also discussed, with emphasis on

  16. ACE inhibition is superior to angiotensin receptor blockade for renography in renal artery stenosis

    International Nuclear Information System (INIS)

    Karanikas, Georgios; Becherer, Alexander; Wiesner, Karoline; Dudczak, Robert; Kletter, Kurt

    2002-01-01

    Angiotensin converting enzyme (ACE) inhibitors as well as angiotensin II receptor antagonists are able to prevent the vasoconstrictive effect of angiotensin II on the efferent renal vessels, which is believed to play an important role in renovascular hypertension. This effect is assumed to be essential for the demonstration of renovascular hypertension by captopril renography. In this study, renographic changes induced by captopril and the AT1 receptor antagonist valsartan were compared in patients with a high probability for renovascular hypertension. Twenty-five patients with 33 stenosed renal arteries (grade of stenosis >50%) and hypertension were studied. Captopril, valsartan and baseline renography were performed within 48 h using technetium-99m mercaptoacetyltriglycine. Blood pressure was monitored, plasma renin concentration before and after intervention was determined and urinary flow was estimated from the urinary output of the hydrated patients. Alterations in renographic curves after intervention were evaluated according to the Santa Fe consensus on ACE inhibitor renography. Captopril renography was positive, indicating renovascular hypertension, in 25 of the 33 stenosed vessels, whereas valsartan renography was positive in only ten. Blood pressure during captopril and valsartan renography was not different; reduction in blood pressure was the same after valsartan and captopril. Plasma renin concentration was comparable for valsartan and captopril studies, showing suppressed values after intervention in as many as 12 of the 25 patients. Urinary flow after valsartan was higher than after captopril (P<0.05). However, this difference could not explain the markedly higher sensitivity of captopril compared with valsartan in demonstrating renal artery stenosis. In 14 of the 25 patients, blood pressure response to revascularisation was monitored, showing a much better predictive value for captopril renography. It is concluded that captopril renography is much

  17. ACE inhibition is superior to angiotensin receptor blockade for renography in renal artery stenosis

    Energy Technology Data Exchange (ETDEWEB)

    Karanikas, Georgios; Becherer, Alexander; Wiesner, Karoline; Dudczak, Robert; Kletter, Kurt [Department of Nuclear Medicine, University of Vienna (Austria)

    2002-03-01

    Angiotensin converting enzyme (ACE) inhibitors as well as angiotensin II receptor antagonists are able to prevent the vasoconstrictive effect of angiotensin II on the efferent renal vessels, which is believed to play an important role in renovascular hypertension. This effect is assumed to be essential for the demonstration of renovascular hypertension by captopril renography. In this study, renographic changes induced by captopril and the AT1 receptor antagonist valsartan were compared in patients with a high probability for renovascular hypertension. Twenty-five patients with 33 stenosed renal arteries (grade of stenosis >50%) and hypertension were studied. Captopril, valsartan and baseline renography were performed within 48 h using technetium-99m mercaptoacetyltriglycine. Blood pressure was monitored, plasma renin concentration before and after intervention was determined and urinary flow was estimated from the urinary output of the hydrated patients. Alterations in renographic curves after intervention were evaluated according to the Santa Fe consensus on ACE inhibitor renography. Captopril renography was positive, indicating renovascular hypertension, in 25 of the 33 stenosed vessels, whereas valsartan renography was positive in only ten. Blood pressure during captopril and valsartan renography was not different; reduction in blood pressure was the same after valsartan and captopril. Plasma renin concentration was comparable for valsartan and captopril studies, showing suppressed values after intervention in as many as 12 of the 25 patients. Urinary flow after valsartan was higher than after captopril (P<0.05). However, this difference could not explain the markedly higher sensitivity of captopril compared with valsartan in demonstrating renal artery stenosis. In 14 of the 25 patients, blood pressure response to revascularisation was monitored, showing a much better predictive value for captopril renography. It is concluded that captopril renography is much

  18. Leptin receptor signaling inhibits ovarian follicle development and egg laying in chicken hens

    Science.gov (United States)

    2014-01-01

    Background Nutrition intake during growth strongly influences ovarian follicle development and egg laying in chicken hens, yet the underlying endocrine regulatory mechanism is still poorly understood. The relevant research progress is hindered by difficulties in detection of leptin gene and its expression in the chicken. However, a functional leptin receptor (LEPR) is present in the chicken which has been implicated to play a regulatory role in ovarian follicle development and egg laying. The present study targeted LEPR by immunizing against its extracellular domain (ECD), and examined the resultant ovarian follicle development and egg-laying rate in chicken hens. Methods Hens that have been immunized four times with chicken LEPR ECD were assessed for their egg laying rate and feed intake, numbers of ovarian follicles, gene expression profiles, serum lipid parameters, as well as STAT3 signaling pathway. Results Administrations of cLEPR ECD antigen resulted in marked reductions in laying rate that over time eventually recovered to the levels exhibited by the Control hens. Together with the decrease in egg laying rate, cLEPR-immunized hens also exhibited significant reductions in feed intake, plasma concentrations of glucose, triglyceride, high-density lipoprotein, and low-density lipoprotein. Parallelled by reductions in feed intake, mRNA gene expression levels of AgRP, orexin, and NPY were down regulated, but of POMC, MC4R and lepR up-regulated in Immunized hen hypothalamus. cLEPR-immunization also promoted expressions of apoptotic genes such as caspase3 in theca and fas in granulosa layer, but severely depressed IGF-I expression in both theca and granulosa layers. Conclusions Immunization against cLEPR ECD in egg-laying hens generated antibodies that mimic leptin bioactivity by enhancing leptin receptor transduction. This up-regulated apoptotic gene expression in ovarian follicles, negatively regulated the expression of genes that promote follicular development

  19. Inhibition of Epithelial TNF-α Receptors by Purified Fruit Bromelain Ameliorates Intestinal Inflammation and Barrier Dysfunction in Colitis

    OpenAIRE

    Zhou, Zijuan; Wang, Liang; Feng, Panpan; Yin, Lianhong; Wang, Chen; Zhi, Shengxu; Dong, Jianyi; Wang, Jingyu; Lin, Yuan; Chen, Dapeng; Xiong, Yongjian; Peng, Jinyong

    2017-01-01

    Activation of the TNF-α receptor (TNFR) leads to an inflammatory response, and anti-TNF therapy has been administered to reduce inflammation symptoms and heal mucosal ulcers in inflammatory bowel disease (IBD). Bromelain, a complex natural mixture of proteolytic enzymes, has been shown to exert anti-inflammatory effects. This study aimed to investigate the effect of purified fruit bromelain (PFB)-induced inhibition of epithelial TNFR in a rat colitis model. Colitis was established by intracol...

  20. Inhibition of alpha oscillations through serotonin-2A receptor activation underlies the visual effects of ayahuasca in humans

    OpenAIRE

    Valle, Marta; Ana Elda, Maqueda; Rabella, Mireia; Rodríguez Pujadas, Aina; Antonijoan Arbós, Rosa Maria; Romero Lafuente, Sergio; Alonso López, Joan Francesc; Mañanas Villanueva, Miguel Ángel; Barker, Steven; Friedlander, Pablo; Feilding, Amanda; Riba, Jordi

    2016-01-01

    Ayahuasca is an Amazonian psychotropic plant tea typically obtained from two plants, Banisteriopsis caapi and Psychotria viridis. It contains the psychedelic 5-HT2A and sigma-1 agonist N,N-dimethyltryptamine (DMT) plus ß-carboline alkaloids with monoamine-oxidase (MAO)-inhibiting properties. Although the psychoactive effects of ayahuasca have commonly been attributed solely to agonism at the 5-HT2A receptor, the molecular target of classical psychedelics, this has not been tested experimental...

  1. Selective inhibition of prostaglandin E2 receptors EP2 and EP4 inhibits adhesion of human endometriotic epithelial and stromal cells through suppression of integrin-mediated mechanisms.

    Science.gov (United States)

    Lee, JeHoon; Banu, Sakhila K; Burghardt, Robert C; Starzinski-Powitz, Anna; Arosh, Joe A

    2013-03-01

    Endometriosis is a chronic gynecological disease of reproductive age women characterized by the presence of functional endometrial tissues outside the uterine cavity. Interactions between the endometriotic cells and the peritoneal extracellular matrix proteins (ECM) are crucial mechanisms that allow adhesion of the endometriotic cells into peritoneal mesothelia. Prostaglandin E2 (PGE2) plays an important role in the pathogenesis of endometriosis. In previous studies, we have reported that selective inhibition of PGE2 receptors PTGER2 and PTGER4 decreases survival and invasion of human endometriotic epithelial and stromal cells through multiple mechanisms. Results of the present study indicates that selective inhibition of PTGER2- and PTGER4-mediated PGE2 signaling 1) decreases the expression and/or activity of specific integrin receptor subunits Itgb1 (beta1) and Itgb3 (beta3) but not Itgb5 (beta5), Itga1 (alpha1), Itga2 (alpha2), Itga5 (alpha5), and Itgav (alphav); 2) decreases integrin-signaling components focal adhesion kinase or protein kinase 2 (PTK2) and talin proteins; 3) inhibits interactions between Itgb1/Itgb3 subunits, PTK2, and talin and PTGER2/PTGER4 proteins through beta-arrestin-1 and Src kinase protein complex in human endometriotic epithelial cells 12Z and stromal cells 22B; and 4) decreases adhesion of 12Z and 22B cells to ECM collagen I, collagen IV, fibronectin, and vitronectin in a substrate-specific manner. These novel findings provide an important molecular framework for further evaluation of selective inhibition of PTGER2 and PTGER4 as potential nonsteroidal therapy to expand the spectrum of currently available treatment options for endometriosis in child-bearing age women.

  2. Arctigenin reduces neuronal responses in the somatosensory cortex via the inhibition of non-NMDA glutamate receptors.

    Science.gov (United States)

    Borbély, Sándor; Jócsák, Gergely; Moldován, Kinga; Sedlák, Éva; Preininger, Éva; Boldizsár, Imre; Tóth, Attila; Atlason, Palmi T; Molnár, Elek; Világi, Ildikó

    2016-07-01

    Lignans are biologically active phenolic compounds related to lignin, produced in different plants. Arctigenin, a dibenzylbutyrolactone-type lignan, has been used as a neuroprotective agent for the treatment of encephalitis. Previous studies of cultured rat cerebral cortical neurones raised the possibility that arctigenin inhibits kainate-induced excitotoxicity. The aims of the present study were: 1) to analyse the effect of arctigenin on normal synaptic activity in ex vivo brain slices, 2) to determine its receptor binding properties and test the effect of arctigenin on AMPA/kainate receptor activation and 3) to establish its effects on neuronal activity in vivo. Arctigenin inhibited glutamatergic transmission and reduced the evoked field responses. The inhibitory effect of arctigenin on the evoked field responses proved to be substantially dose dependent. Our results indicate that arctigenin exerts its effects under physiological conditions and not only on hyper-excited neurons. Furthermore, arctigenin can cross the blood-brain barrier and in the brain it interacts with kainate sensitive ionotropic glutamate receptors. These results indicate that arctigenin is a potentially useful new pharmacological tool for the inhibition of glutamate-evoked responses in the central nervous system in vivo. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Anticonvulsants Based on the α-Substituted Amide Group Pharmacophore Bind to and Inhibit Function of Neuronal Nicotinic Acetylcholine Receptors.

    Science.gov (United States)

    Krivoshein, Arcadius V

    2016-03-16

    Although the antiepileptic properties of α-substituted lactams, acetamides, and cyclic imides have been known for over 60 years, the mechanism by which they act remains unclear. I report here that these compounds bind to the nicotinic acetylcholine receptor (nAChR) and inhibit its function. Using transient kinetic measurements with functionally active, nondesensitized receptors, I have discovered that (i) α-substituted lactams and cyclic imides are noncompetitive inhibitors of heteromeric subtypes (such as α4β2 and α3β4) of neuronal nAChRs and (ii) the binding affinity of these compounds toward the nAChR correlates with their potency in preventing maximal electroshock (MES)-induced convulsions in mice. Based on the hypothesis that α-substituted amide group is the essential pharmacophore of these drugs, I found and tested a simple compound, 2-phenylbutyramide. This compound indeed inhibits nAChR and shows good anticonvulsant activity in mice. Molecular docking simulations suggest that α-substituted lactams, acetamides, and cyclic imides bind to the same sites on the extracellular domain of the receptor. These new findings indicate that inhibition of brain nAChRs may play an important role in the action of these antiepileptic drugs, a role that has not been previously recognized.

  4. Glioblastoma chemotherapy adjunct via potent serotonin receptor-7 inhibition using currently marketed high-affinity antipsychotic medicines

    Science.gov (United States)

    Kast, RE

    2010-01-01

    Glioblastoma treatment as now constituted offers increased survival measured in months over untreated patients. Because glioblastomas are active in synthesizing a bewildering variety of growth factors, a systematic approach to inhibiting these is being undertaken as treatment adjunct. The serotonin 7 receptor is commonly overexpressed in glioblastoma. Research documentation showing agonists at serotonin receptor 7 cause increased extracellular regulated kinase 1/2 activation, increased interleukin-6 synthesis, increased signal transducer and activator of transcription-3 activation, increased resistance to apoptosis and other growth enhancing changes in glioblastoma is reviewed in this paper. Because three drugs in wide use to treat thought disorders – paliperidone, pimozide and risperidone – are also potent and well-tolerated inhibitors at serotonin receptor 7, these drugs should be studied for growth factor deprivation in an adjunctive role in glioblastoma treatment. PMID:20880389

  5. Inverse agonism at the P2Y12 receptor and ENT1 transporter blockade contribute to platelet inhibition by ticagrelor.

    Science.gov (United States)

    Aungraheeta, Riyaad; Conibear, Alexandra; Butler, Mark; Kelly, Eamonn; Nylander, Sven; Mumford, Andrew; Mundell, Stuart J

    2016-12-08

    Ticagrelor is a potent antagonist of the P2Y 12 receptor (P2Y 12 R) and consequently an inhibitor of platelet activity effective in the treatment of atherothrombosis. Here, we sought to further characterize its molecular mechanism of action. Initial studies showed that ticagrelor promoted a greater inhibition of adenosine 5'-diphosphate (ADP)-induced Ca 2+ release in washed platelets vs other P2Y 12 R antagonists. This additional effect of ticagrelor beyond P2Y 12 R antagonism was in part as a consequence of ticagrelor inhibiting the equilibrative nucleoside transporter 1 (ENT1) on platelets, leading to accumulation of extracellular adenosine and activation of G s -coupled adenosine A 2A receptors. This contributed to an increase in basal cyclic adenosine monophosphate (cAMP) and vasodilator-stimulated phosphoprotein phosphorylation (VASP-P). In addition, ticagrelor increased platelet cAMP and VASP-P in the absence of ADP in an adenosine receptor-independent manner. We hypothesized that this increase originated from a direct effect on basal agonist-independent P2Y 12 R signaling, and this was validated in 1321N1 cells stably transfected with human P2Y 12 R. In these cells, ticagrelor blocked the constitutive agonist-independent activity of the P2Y 12 R, limiting basal G i -coupled signaling and thereby increasing cAMP levels. These data suggest that ticagrelor has the pharmacological profile of an inverse agonist. Based on our results showing insurmountable inhibition of ADP-induced Ca 2+ release and forskolin-induced cAMP, the mode of antagonism of ticagrelor also appears noncompetitive, at least functionally. In summary, our studies describe 2 novel modes of action of ticagrelor, inhibition of platelet ENT1 and inverse agonism at the P2Y 12 R that contribute to its effective inhibition of platelet activation. © 2016 by The American Society of Hematology.

  6. Inhibition of aryl hydrocarbon receptor-dependent transcription by resveratrol or kaempferol is independent of estrogen receptor α expression in human breast cancer cells

    Science.gov (United States)

    MacPherson, Laura; Matthews, Jason

    2016-01-01

    Resveratrol and kaempferol are natural chemopreventative agents that are also aryl hydrocarbon receptor (AHR) antagonists and estrogen receptor (ER) agonists. In this study we evaluated the role of ERα in resveratrol- and kaempferol-mediated inhibition of AHR-dependent transcription. Kaempferol or resveratrol inhibited dioxin-induced cytochrome P450 1A1 (CYP1A1) and CYP1B1 expression levels and recruitment of AHR, ERα and co-activators to CYP1A1 and CYP1B1. Both phytochemicals induced the expression and recruitment of ERα to gene amplified in breast cancer 1 (GREB1). RNAi-mediated knockdown of ERα in T-47D cells did not affect the inhibitory action of either phytochemical on AHR activity. Both compounds also inhibited AHR-dependent transcription in ERα-negative MDA-MB-231 and BT-549 breast cancer cells. These data show that ERα does not contribute to the AHR-inhibitory activities of resveratrol and kaempferol. PMID:20846786

  7. Intestinal Serotonin Transporter Inhibition by Toll-Like Receptor 2 Activation. A Feedback Modulation.

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    Eva Latorre

    Full Text Available TLR2 is a microbiota recognition receptor that has been described to contribute to intestinal homeostasis and to ameliorate inflammatory intestinal injury. In this context, serotonin (5-HT has shown to be an essential intestinal physiological neuromodulator that is also involved in intestinal inflammatory diseases. Since the interaction between TLR2 activation and the intestinal serotoninergic system remains non-investigated, our main aim was to analyze the effect of TLR2 on intestinal serotonin transporter (SERT activity and expression and the intracellular pathways involved. Caco-2/TC7 cells were used to analyze SERT and TLR2 molecular expression and SERT activity by measuring 5-HT uptake. The results showed that apical TLR2 activation inhibits SERT activity in Caco-2/TC7 cells mainly by reducing SERT protein level either in the plasma membrane, after short-term TLR2 activation or in both the plasma membrane and cell lysate, after long-term activation. cAMP/PKA pathway appears to mediate short-term inhibitory effect of TLR2 on SERT; however, p38 MAPK pathway has been shown to be involved in both short- and long-term TLR2 effect. Reciprocally, 5-HT long-term treatment yielded TLR2 down regulation in Caco-2/TC7 cells. Finally, results from in vivo showed an augmented intestinal SERT expression in mice Tlr2-/-, thus confirming our inhibitory effect of TLR2 on intestinal SERT in vitro. The present work infers that TLR2 may act in intestinal pathophysiology, not only by its inherent innate immune role, but also by regulating the intestinal serotoninergic system.

  8. Epidermal growth factor receptor inhibition by anti-CD147 therapy in cutaneous squamous cell carcinoma.

    Science.gov (United States)

    Frederick, John W; Sweeny, Larissa; Hartman, Yolanda; Zhou, Tong; Rosenthal, Eben L

    2016-02-01

    Advanced cutaneous squamous cell carcinoma (SCC) is an uncommon and aggressive malignancy. As a result, there is limited understanding of its biology and pathogenesis. CD147 and epidermal growth factor receptor (EGFR) have been identified as oncologically important targets, but their relationship remains undefined in cutaneous SCC. Multiple cutaneous SCC cell lines (Colo-16, SRB-1, and SRB-12), were treated in vitro with a range of chimeric anti-CD147 monoclonal antibody (mAb) (0, 50, 100, and 200 µg/mL) or transfected with a small interfering RNA against CD147 (SiCD147). Cell proliferation, migration (scratch wound healing assay), and protein expression was then assessed. In vivo, Colo-16 flank xenografts were treated anti-CD147 mAb (150 µg i.p. triweekly). After treatment with anti-CD147 (200 µg/mL), there was a significant decrease in proliferation for all cell lines relative to controls (p CD147 (200 µg/mL) resulted in decreased cell migration for all cell lines, with an average of 43% reduction in closure compared to controls (p CD147 antibody therapy and siRNA mediated reduction in CD147 expression were both found to decrease protein expression of EGFR, which correlated with a reduction in downstream total and phosphorylated protein kinase B (pAKT). Tumor growth in vivo was reduced for both the anti-CD147 treatment group and the SiCD147 group relative to controls. Inhibition and downregulation of CD147 in cutaneous SCC resulted in suppression of the malignant phenotype in vitro and in vivo, which may be mediated in part by an alteration in EGFR expression. As a result, CD147 may serve as a potential therapeutic target for advanced cutaneous SCC. © 2014 Wiley Periodicals, Inc.

  9. Clofibric acid, a peroxisome proliferator-activated receptor alpha ligand, inhibits growth of human ovarian cancer.

    Science.gov (United States)

    Yokoyama, Yoshihito; Xin, Bing; Shigeto, Tatsuhiko; Umemoto, Mika; Kasai-Sakamoto, Akiko; Futagami, Masayuki; Tsuchida, Shigeki; Al-Mulla, Fahd; Mizunuma, Hideki

    2007-04-01

    Recent reports have shown that peroxisome proliferator-activated receptor (PPAR)alpha ligands reduce growth of some types of malignant tumors and prevent carcinogenesis. In this study, we investigated the inhibitory effect of clofibric acid (CA), a ligand for PPARalpha on growth of ovarian malignancy, in in vivo and in vitro experiments using OVCAR-3 and DISS cells derived from human ovarian cancer and aimed to elucidate the molecular mechanism of its antitumor effect. CA treatment significantly suppressed the growth of OVCAR-3 tumors xenotransplanted s.c. and significantly prolonged the survival of mice with malignant ascites derived from DISS cells as compared with control. CA also dose-dependently inhibited cell proliferation of cultured cell lines. CA treatment increased the expression of carbonyl reductase (CR), which promotes the conversion of prostaglandin E(2) (PGE(2)) to PGF(2alpha), in implanted OVCAR-3 tumors as well as cultured cells. CA treatment decreased PGE(2) level as well as vascular endothelial growth factor (VEGF) amount in both of OVCAR-3-tumor and DISS-derived ascites. Reduced microvessel density and induced apoptosis were found in solid OVCAR-3 tumors treated by CA. Transfection of CR expression vector into mouse ovarian cancer cells showed significant reduction of PGE(2) level as well as VEGF expression. These results indicate that CA produces potent antitumor effects against ovarian cancer in conjunction with a reduction of angiogenesis and induction of apoptosis. We conclude that CA could be an effective agent in ovarian cancer and should be tested alone and in combination with other anticancer drugs.

  10. Covalent Targeting of Fibroblast Growth Factor Receptor Inhibits Metastatic Breast Cancer.

    Science.gov (United States)

    Brown, Wells S; Tan, Li; Smith, Andrew; Gray, Nathanael S; Wendt, Michael K

    2016-09-01

    Therapeutic targeting of late-stage breast cancer is limited by an inadequate understanding of how tumor cell signaling evolves during metastatic progression and by the currently available small molecule inhibitors capable of targeting these processes. Herein, we demonstrate that both β3 integrin and fibroblast growth factor receptor-1 (FGFR1) are part of an epithelial-mesenchymal transition (EMT) program that is required to facilitate metastatic outgrowth in response to fibroblast growth factor-2 (FGF2). Mechanistically, β3 integrin physically disrupts an interaction between FGFR1 and E-cadherin, leading to a dramatic redistribution of FGFR1 subcellular localization, enhanced FGF2 signaling and increased three-dimensional (3D) outgrowth of metastatic breast cancer cells. This ability of β3 integrin to drive FGFR signaling requires the enzymatic activity of focal adhesion kinase (FAK). Consistent with these mechanistic data, we demonstrate that FGFR, β3 integrin, and FAK constitute a molecular signature capable of predicting decreased survival of patients with the basal-like subtype of breast cancer. Importantly, covalent targeting of a conserved cysteine in the P-loop of FGFR1-4 with our newly developed small molecule, FIIN-4, more effectively blocks 3D metastatic outgrowth as compared with currently available FGFR inhibitors. In vivo application of FIIN-4 potently inhibited the growth of metastatic, patient-derived breast cancer xenografts and murine-derived metastases growing within the pulmonary microenvironment. Overall, the current studies demonstrate that FGFR1 works in concert with other EMT effector molecules to drive aberrant downstream signaling, and that these events can be effectively targeted using our novel therapeutics for the treatment of the most aggressive forms of breast cancer. Mol Cancer Ther; 15(9); 2096-106. ©2016 AACR. ©2016 American Association for Cancer Research.

  11. Voltage-Dependent Inhibition of Glycine Receptor Channels by Niflumic Acid

    Directory of Open Access Journals (Sweden)

    Galyna Maleeva

    2017-05-01

    Full Text Available Niflumic acid (NFA is a member of the fenamate class of nonsteroidal anti-inflammatory drugs. This compound and its derivatives are used worldwide clinically for the relief of chronic and acute pain. NFA is also a commonly used blocker of voltage-gated chloride channels. Here we present evidence that NFA is an efficient blocker of chloride-permeable glycine receptors (GlyRs with subunit heterogeneity of action. Using the whole-cell configuration of patch-clamp recordings and molecular modeling, we analyzed the action of NFA on homomeric α1ΔIns, α2B, α3L, and heteromeric α1β and α2β GlyRs expressed in CHO cells. NFA inhibited glycine-induced currents in a voltage-dependent manner and its blocking potency in α2 and α3 GlyRs was higher than that in α1 GlyR. The Woodhull analysis suggests that NFA blocks α1 and α2 GlyRs at the fractional electrical distances of 0.16 and 0.65 from the external membrane surface, respectively. Thus, NFA binding site in α1 GlyR is closer to the external part of the membrane, while in α2 GlyR it is significantly deeper in the pore. Mutation G254A at the cytoplasmic part of the α1 GlyR pore-lining TM2 helix (level 2′ increased the NFA blocking potency, while incorporation of the β subunit did not have a significant effect. The Hill plot analysis suggests that α1 and α2 GlyRs are preferably blocked by two and one NFA molecules, respectively. Molecular modeling using Monte Carlo energy minimizations provides the structural rationale for the experimental data and proposes more than one interaction site along the pore where NFA can suppress the ion permeation.

  12. Prenatal Alcohol Exposure Increases Histamine H3 Receptor-Mediated Inhibition of Glutamatergic Neurotransmission in Rat Dentate Gyrus.

    Science.gov (United States)

    Varaschin, Rafael K; Allen, Nyika A; Rosenberg, Martina J; Valenzuela, C Fernando; Savage, Daniel D

    2018-02-01

    We have reported that prenatal alcohol exposure (PAE)-induced deficits in dentate gyrus, long-term potentiation (LTP), and memory are ameliorated by the histamine H 3 receptor inverse agonist ABT-239. Curiously, ABT-239 did not enhance LTP or memory in control offspring. Here, we initiated an investigation of how PAE alters histaminergic neurotransmission in the dentate gyrus and other brain regions employing combined radiohistochemical and electrophysiological approaches in vitro to examine histamine H 3 receptor number and function. Long-Evans rat dams voluntarily consumed either a 0% or 5% ethanol solution 4 hours each day throughout gestation. This pattern of drinking, which produces a mean peak maternal serum ethanol concentration of 60.8 ± 5.8 mg/dl, did not affect maternal weight gain, litter size, or offspring birthweight. Radiohistochemical studies in adult offspring revealed that specific [ 3 H]-A349821 binding to histamine H 3 receptors was not different in PAE rats compared to controls. However, H 3 receptor-mediated G i /G o protein-effector coupling, as measured by methimepip-stimulated [ 35 S]-GTPγS binding, was significantly increased in cerebral cortex, cerebellum, and dentate gyrus of PAE rats compared to control. A LIGAND analysis of detailed methimepip concentration-response curves in dentate gyrus indicated that PAE significantly elevates receptor-effector coupling by a lower affinity H 3 receptor population without significantly altering the affinities of H 3 receptor subpopulations. In agreement with the [ 35 S]-GTPγS studies, a similar range of methimepip concentrations also inhibited electrically evoked field excitatory postsynaptic potential responses and increased paired-pulse ratio, a measure of decreased glutamate release, to a significantly greater extent in dentate gyrus slices from PAE rats than in controls. These results suggest that a PAE-induced elevation in H 3 receptor-mediated inhibition of glutamate release from

  13. Colony stimulating factor 1 receptor inhibition delays recurrence of glioblastoma after radiation by altering myeloid cell recruitment and polarization

    Science.gov (United States)

    Stafford, Jason H.; Hirai, Takahisa; Deng, Lei; Chernikova, Sophia B.; Urata, Kimiko; West, Brian L.; Brown, J. Martin

    2016-01-01

    Background Glioblastoma (GBM) may initially respond to treatment with ionizing radiation (IR), but the prognosis remains extremely poor because the tumors invariably recur. Using animal models, we previously showed that inhibiting stromal cell–derived factor 1 signaling can prevent or delay GBM recurrence by blocking IR-induced recruitment of myeloid cells, specifically monocytes that give rise to tumor-associated macrophages. The present study was aimed at determining if inhibiting colony stimulating factor 1 (CSF-1) signaling could be used as an alternative strategy to target pro-tumorigenic myeloid cells recruited to irradiated GBM. Methods To inhibit CSF-1 signaling in myeloid cells, we used PLX3397, a small molecule that potently inhibits the tyrosine kinase activity of the CSF-1 receptor (CSF-1R). Combined IR and PLX3397 therapy was compared with IR alone using 2 different human GBM intracranial xenograft models. Results GBM xenografts treated with IR upregulated CSF-1R ligand expression and increased the number of CD11b+ myeloid-derived cells in the tumors. Treatment with PLX3397 both depleted CD11b+ cells and potentiated the response of the intracranial tumors to IR. Median survival was significantly longer for mice receiving combined therapy versus IR alone. Analysis of myeloid cell differentiation markers indicated that CSF-1R inhibition prevented IR-recruited monocyte cells from differentiating into immunosuppressive, pro-angiogenic tumor-associated macrophages. Conclusion CSF-1R inhibition may be a promising strategy to improve GBM response to radiotherapy. PMID:26538619

  14. Evaluation of the In Vivo and Ex Vivo Binding of Novel BC1 Cannabinoid Receptor Radiotracers

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    Miller, A.; Gatley, J.; Gifford, A.

    2002-01-01

    The primary active ingredient of marijuana, 9-tetrahydrocannabinol, exerts its psychoactive effects by binding to cannabinoid CB1 receptors. These receptors are found throughout the brain with high concentrations in the hippocampus and cerebellum. The current study was conducted to evaluate the binding of a newly developed putative cannabinoid antagonist, AM630, and a classical cannabinoid 8-tetrahydrocannabinol as potential PET and/or SPECT imaging agents for brain CB1 receptors. For both of these ligands in vivo and ex vivo studies in mice were conducted. AM630 showed good overall brain uptake (as measure by %IA/g) and a moderately rapid clearance from the brain with a half-clearance time of approximately 30 minutes. However, AM630 did not show selective binding to CB1 cannabinoid receptors. Ex vivo autoradiography supported the lack of selective binding seen in the in vivo study. Similar to AM630, 8-tetrahydrocanibol also failed to show selective binding to CB1 receptor rich brain areas. The 8-tetrahydrocanibol showed moderate overall brain uptake and relatively slow brain clearance as compared to AM630. Further studies were done with AM2233, a cannabinoid ligand with a similar structure as AM630. These studies were done to develop an ex vivo binding assay to quantify the displacement of [131I]AM2233 binding by other ligands in Swiss-Webster and CB1 receptor knockout mice. By developing this assay we hoped to determine the identity of an unknown binding site for AM2233 present in the hippocampus of CB1 knockout mice. Using an approach based on incubation of brain slices prepared from mice given intravenous [131I]AM2233 in either the presence or absence of AM2233 (unlabelled) it was possible to demonstrate a significant AM2233-displacable binding in the Swiss-Webster mice. Future studies will determine if this assay is appropriate for identifying the unknown binding site for AM2233 in the CB1 knockout mice.

  15. Group I mGlu receptor stimulation inhibits activation-induced cell death of human T lymphocytes

    Science.gov (United States)

    Chiocchetti, Annalisa; Miglio, Gianluca; Mesturini, Riccardo; Varsaldi, Federica; Mocellin, Marco; Orilieri, Elisabetta; Dianzani, Chiara; Fantozzi, Roberto; Dianzani, Umberto; Lombardi, Grazia

    2006-01-01

    The effects of L-glutamate on activation-induced cell death (AICD) of human activated (1 μg ml−1 phytohemagglutinin plus 2 U ml−1 interleukin-2; 8 days) T lymphocytes were studied by measuring anti-CD3 monoclonal antibody (10 μg ml−1; 18 h)-induced cell apoptosis (Annexin V and propidium iodide staining). L-Glutamate (1 × 10−8–1 × 10−4 M) significantly (P⩽0.01) inhibited AICD in a concentration-dependent manner (EC50=6.3 × 10−8 M; maximum inhibition 54.8±6.3% at 1 × 10−6 M). The L-glutamate inhibitory effect was pharmacologically characterized as mediated by group I mGlu receptors, since mGlu receptor agonists reproduced this effect. The EC50 values were: 3.2 × 10−7 M for (1S,3R)-ACPD; 4.5 × 10−8 M for quisqualate; 1.0 × 10−6 M for (S)-3,5-DHPG; 2.0 × 10−5 M for CHPG. Group I mGlu receptor antagonists inhibited the effects of quisqualate 1.0 × 10−6 M. The IC50 values calculated were: 8.7 × 10−5, 4.3 × 10−6 and 6.3 × 10−7 M for AIDA, LY 367385 and MPEP, respectively. L-Glutamate (1 × 10−6 M; 18 h) significantly (P⩽0.05) inhibited FasL expression (40.8±11.3%) (cytofluorimetric analysis), whereas it did not affect Fas signalling. Expression of both mGlu1 and mGlu5 receptor mRNA by T lymphocytes and T-cell lines, as demonstrated by reverse transcriptase–PCR analysis, suggests that L-glutamate-mediated inhibition of AICD was exerted on T cells. These data depict a novel role for L-glutamate in the regulation of the immune response through group I mGlu receptor-mediated mechanisms. PMID:16751798

  16. Small-animal PET imaging of the type 1 and type 2 cannabinoid receptors in a photothrombotic stroke model

    International Nuclear Information System (INIS)

    Vandeputte, Caroline; Casteels, Cindy; Koole, Michel; Gerits, Anneleen; Struys, Tom; Veghel, Daisy van; Evens, Nele; Bormans, Guy; Dresselaers, Tom; Himmelreich, Uwe; Lambrichts, Ivo; Laere, Koen van

    2012-01-01

    Recent ex vivo and pharmacological evidence suggests involvement of the endocannabinoid system in the pathophysiology of stroke, but conflicting roles for type 1 and 2 cannabinoid receptors (CB 1 and CB 2 ) have been suggested. The purpose of this study was to evaluate CB 1 and CB 2 receptor binding over time in vivo in a rat photothrombotic stroke model using PET. CB 1 and CB 2 microPET imaging was performed at regular time-points up to 2 weeks after stroke using [ 18 F]MK-9470 and [ 11 C]NE40. Stroke size was measured using MRI at 9.4 T. Ex vivo validation was performed via immunostaining for CB 1 and CB 2 . Immunofluorescent double stainings were also performed with markers for astrocytes (GFAP) and macrophages/microglia (CD68). [ 18 F]MK-9470 PET showed a strong increase in CB 1 binding 24 h and 72 h after stroke in the cortex surrounding the lesion, extending to the insular cortex 24 h after surgery. These alterations were consistently confirmed by CB 1 immunohistochemical staining. [ 11 C]NE40 did not show any significant differences between stroke and sham-operated animals, although staining for CB 2 revealed minor immunoreactivity at 1 and 2 weeks after stroke in this model. Both CB 1 + and CB 2 + cells showed minor immunoreactivity for CD68. Time-dependent and regionally strongly increased CB 1 , but not CB 2 , binding are early consequences of photothrombotic stroke. Pharmacological interventions should primarily aim at CB 1 signalling as the role of CB 2 seems minor in the acute and subacute phases of stroke. (orig.)

  17. Co-inhibition of epidermal growth factor receptor and insulin-like growth factor receptor 1 enhances radiosensitivity in human breast cancer cells

    International Nuclear Information System (INIS)

    Li, Ping; Veldwijk, Marlon R; Zhang, Qing; Li, Zhao-bin; Xu, Wen-cai; Fu, Shen

    2013-01-01

    Over-expression of epidermal growth factor receptor (EGFR) or insulin-like growth factor-1 receptor (IGF-1R) have been shown to closely correlate with radioresistance of breast cancer cells. This study aimed to investigate the impact of co-inhibition of EGFR and IGF-1R on the radiosensitivity of two breast cancer cells with different profiles of EGFR and IGF-1R expression. The MCF-7 (EGFR +/−, IGF-1R +++) and MDA-MB-468 (EGFR +++, IGF-1R +++) breast cancer cell lines were used. Radiosensitizing effects were determined by colony formation assay. Apoptosis and cell cycle distribution were measured by flow cytometry. Phospho-Akt and phospho-Erk1/2 were quantified by western blot. In vivo studies were conducted using MDA-MB-468 cells xenografted in nu/nu mice. In MDA-MB-468 cells, the inhibition of IGF-1R upregulated the p-EGFR expression. Either EGFR (AG1478) or IGF-1R inhibitor (AG1024) radiosensitized MDA-MB-468 cells. In MCF-7 cells, radiosensitivity was enhanced by AG1024, but not by AG1478. Synergistical radiosensitizing effect was observed by co-inhibition of EGFR and IGF-1R only in MDA-MB-468 cells with a DMF 10% of 1.90. The co-inhibition plus irradiation significantly induced more apoptosis and arrested the cells at G0/G1 phase in MDA-MB-468 cells. Only co-inhibition of EGFR and IGF-1R synergistically diminished the expression of p-Akt and p-Erk1/2 in MDA-MB-468 cells. In vivo studies further verified the radiosensitizing effects by co-inhibition of both pathways in a MDA-MB-468 xenograft model. Our data suggested that co-inhibition of EGFR and IGF-1R synergistically radiosensitized breast cancer cells with both EGFR and IGF-1R high expression. The approach may have an important therapeutic implication in the treatment of breast cancer patients with high expression of EGFR and IGF-1R

  18. Glycine and GABAA receptors mediate tonic and phasic inhibitory processes that contribute to prepulse inhibition in the goldfish startle network

    Directory of Open Access Journals (Sweden)

    Paul C.P. Curtin

    2015-03-01

    Full Text Available Prepulse inhibition (PPI is understood as an inhibitory process that attenuates sensory flow during early stages (20-1000ms of information processing. Here, we applied in vivo electrophysiology and pharmacology to determine if prepulse inhibition (PPI is mediated by glycine receptors (GlyRs and/or GABAA receptors (GABAARs in the goldfish auditory startle circuit. Specifically, we used selective antagonists to dissect the contributions of target receptors on sound-evoked postsynaptic potentials (PSPs recorded in the neurons that initiate startle, the Mauthner-cells (M-cell. We found that strychnine, a GlyR antagonist, disrupted a fast-activated (5 ms and rapidly (< 50ms decaying (feed-forward inhibitory process that disrupts PPI at 20 ms prepulse/pulse inter-stimulus intervals (ISI. Additionally we observed increases of the evoked postsynaptic potential (PSP peak amplitude (+87.43 ± 21.53%; N=9 and duration (+204 ± 48.91%, N=9. In contrast, treatment with bicuculline, a GABAAR antagonist, caused a general reduction in PPI across all tested ISIs (20-500 ms, essentially eliminating PPI at ISIs from 20-100 ms. Bicuculline also increased PSP peak amplitude (+133.8 ± 10.3%, N=5 and PSP duration (+284.95 ± 65.64%, N=5. Treatment with either antagonist also tonically increased post-synaptic excitability in the M-cells, reflected by an increase in the magnitude of antidromically-evoked action potentials (APs by 15.07 ± 3.21%, N=7 and 16.23 ± 7.08%, N=5 for strychnine and bicuculline, respectively. These results suggest that GABAARs and GlyRs are functionally segregated to short- and longer-lasting sound-evoked (phasic inhibitory processes that contribute to PPI, with the mediation of tonic inhibition by both receptor systems being critical for gain control within the M-cell startle circuit.

  19. Edaravone inhibits pressure overload-induced cardiac fibrosis and dysfunction by reducing expression of angiotensin II AT1 receptor

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    Zhang WW

    2017-10-01

    Full Text Available Wei-Wei Zhang,1,2 Feng Bai,1 Jin Wang,1 Rong-Hua Zheng,1 Li-Wang Yang,1 Erskine A James,3 Zhi-Qing Zhao1,4 1Department of Physiology, Shanxi Medical University, 2Department of Anesthesiology, Shanxi Provincial People’s Hospital, Taiyuan, Shanxi, China; 3Department of Internal Medicine, Navicent Health, Macon, 4Department of Basic Biomedical Sciences, Mercer University School of Medicine, Savannah, GA, USA Abstract: Angiotensin II (Ang II is known to be involved in the progression of ventricular dysfunction and heart failure by eliciting cardiac fibrosis. The purpose of this study was to demonstrate whether treatment with an antioxidant compound, edaravone, reduces cardiac fibrosis and improves ventricular function by inhibiting Ang II AT1 receptor. The study was conducted in a rat model of transverse aortic constriction (TAC. In control, rats were subjected to 8 weeks of TAC. In treated rats, edaravone (10 mg/kg/day or Ang II AT1 receptor blocker, telmisartan (10 mg/kg/day was administered by intraperitoneal injection or gastric gavage, respectively, during TAC. Relative to the animals with TAC, edaravone reduced myocardial malonaldehyde level and increased superoxide dismutase activity. Protein level of the AT1 receptor was reduced and the AT2 receptor was upregulated, as evidenced by the reduced ratio of AT1 over AT2 receptor (0.57±0.2 vs 3.16±0.39, p<0.05 and less locally expressed AT1 receptor in the myocardium. Furthermore, the protein level of angiotensin converting enzyme 2 was upregulated. In coincidence with these changes, edaravone significantly decreased the populations of macrophages and myofibroblasts in the myocardium, which were accompanied by reduced levels of transforming growth factor beta 1 and Smad2/3. Collagen I synthesis was inhibited and collagen-rich fibrosis was attenuated. Relative to the TAC group, cardiac systolic function was preserved, as shown by increased left ventricular systolic pressure (204±51 vs 110±19

  20. Tissue-type plasminogen activator-binding RNA aptamers inhibiting low-density lipoprotein receptor family-mediated internalisation.

    Science.gov (United States)

    Bjerregaard, Nils; Bøtkjær, Kenneth A; Helsen, Nicky; Andreasen, Peter A; Dupont, Daniel M

    2015-07-01

    Recombinant tissue-type plasminogen activator (tPA, trade name Alteplase), currently the only drug approved by the US Food and Drug Administration and the European Medicines Agency for the treatment of cerebral ischaemic stroke, has been implicated in a number of adverse effects reportedly mediated by interactions with the low-density lipoprotein (LDL) family receptors, including neuronal cell death and an increased risk of cerebral haemorrhage. The tissue-type plasminogen activator is the principal initiator of thrombolysis in human physiology, an effect that is mediated directly via localised activation of the plasmin zymogen plasminogen at the surface of fibrin clots in the vascular lumen. Here, we sought to identify a ligand to tPA capable of inhibiting the relevant LDL family receptors without interfering with the fibrinolytic activity of tPA. Systematic evolution of ligands by exponential enrichment (SELEX) was employed to isolate tPA-binding RNA aptamers, which were characterised in biochemical assays of tPA association to low density lipoprotein receptor-related protein-1 (LRP-1, an LDL receptor family member); tPA-mediated in vitro and ex vivo clot lysis; and tPA-mediated plasminogen activation in the absence and presence of a stimulating soluble fibrin fragment. Two aptamers, K18 and K32, had minimal effects on clot lysis, but were able to efficiently inhibit tPA-LRP-1 association and LDL receptor family-mediated endocytosis in human vascular endothelial cells and astrocytes. These observations suggest that coadministration alongside tPA may be a viable strategy to improve the safety of thrombolytic treatment of cerebral ischaemic stroke by restricting tPA activity to the vascular lumen.

  1. Oral haloperidol or olanzapine intake produces distinct and region-specific increase in cannabinoid receptor levels that is prevented by high fat diet.

    Science.gov (United States)

    Delis, Foteini; Rosko, Lauren; Shroff, Aditya; Leonard, Kenneth E; Thanos, Panayotis K

    2017-10-03

    Clinical studies show higher levels of cannabinoid CB1 receptors (CB1R) in the brain of schizophrenic patients while preclinical studies report a significant functional interaction between dopamine D2 receptors and CB1Rs as well as an upregulation of CB1Rs after antipsychotic treatment. These findings prompted us to study the effects of chronic oral intake of a first and a second generation antipsychotic, haloperidol and olanzapine, on the levels and distribution of CB1Rs in the rat brain. Rats consumed either regular chow or high-fat food and drank water, haloperidol drinking solution (1.5mg/kg), or olanzapine drinking solution (10mg/kg) for four weeks. Motor and cognitive functions were tested at the end of treatment week 3 and upon drug discontinuation. Two days after drug discontinuation, rats were euthanized and brains were processed for in vitro receptor autoradiography. In chow-fed animals, haloperidol and olanzapine increased CB1R levels in the basal ganglia and the hippocampus, in a similar, but not identical pattern. In addition, olanzapine had unique effects in CB1R upregulation in higher order cognitive areas, in the secondary somatosensory cortex, in the visual and auditory cortices and the geniculate nuclei, as well as in the hypothalamus. High fat food consumption prevented antipsychotic-induced increase in CB1R levels in all regions examined, with one exception, the globus pallidus, in which they were higher in haloperidol-treated rats. The results point towards the hypothesis that increased CB1R levels could be a confounding effect of antipsychotic medication in schizophrenia that is circumveneted by high fat feeding. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Prostaglandin D2 inhibits airway dendritic cell migration and function in steady state conditions by selective activation of the D prostanoid receptor 1

    NARCIS (Netherlands)

    H. Hammad (Hamida); H.J. de Heer; T. Soullié (Thomas); H.C. Hoogsteden (Henk); F. Trottein; B.N.M. Lambrecht (Bart)

    2003-01-01

    textabstractPGD(2) is the major mediator released by mast cells during allergic responses, and it acts through two different receptors, the D prostanoid receptor 1 (DP1) and DP2, also known as CRTH2. Recently, it has been shown that PGD(2) inhibits the migration of epidermal

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

    Science.gov (United States)

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

    2017-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Catherine A Vulfius

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

  5. Gefitinib Radiosensitizes Stem-Like Glioma Cells: Inhibition