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Sample records for gamma-hydroxybutyrate receptors stimulation

  1. Gamma-hydroxybutyrate (GHB) induces cognitive deficits and affects GABAB receptors and IGF-1 receptors in male rats.

    PubMed

    Johansson, Jenny; Grönbladh, Alfhild; Hallberg, Mathias

    2014-08-01

    In recent years, the abuse of the club drug gamma-hydroxybutyrate (GHB) has become increasingly popular among adolescents. The drug induces euphoria but can also result in sedation, anaesthesia as well as short-term amnesia. In addition, the abuse of GHB causes cognitive impairments and the mechanism by which GHB induces these impairments is not clarified. The present study investigates the impact of GHB treatment on spatial learning and memory using a water maze (WM) test in rats. Furthermore, the behavioural data is combined with an autoradiographic analysis of the GABAB and the IGF-1 receptor systems. The results demonstrate that the animals administered with GHB display an impaired performance in the WM test as compared to controls. In addition, significant alterations in GABAB and IGF-1 receptor density as well as GABAB receptor functionality, were observed in several brain regions associated with cognitive functions e.g. hippocampus. To conclude, our findings suggest that GHB treatment can affect spatial learning and memory, and that this outcome at least to some extent is likely to involve both GABAB and IGF-1 receptors.

  2. gamma-Hydroxybutyrate (GHB) induces GABA(B) receptor independent intracellular Ca2+ transients in astrocytes, but has no effect on GHB or GABA(B) receptors of medium spiny neurons in the nucleus accumbens.

    PubMed

    Molnár, T; Antal, K; Nyitrai, G; Emri, Z

    2009-08-18

    We report on cellular actions of the illicit recreational drug gamma-hydroxybutyrate (GHB) in the brain reward area nucleus accumbens. First, we compared the effects of GHB and the GABA(B) receptor agonist baclofen. Neither of them affected the membrane currents of medium spiny neurons in rat nucleus accumbens slices. GABAergic and glutamatergic synaptic potentials of medium spiny neurons, however, were reduced by baclofen but not GHB. These results indicate the lack of GHB as well as postsynaptic GABA(B) receptors, and the presence of GHB insensitive presynaptic GABA(B) receptors in medium spiny neurons. In astrocytes GHB induced intracellular Ca(2+) transients, preserved in slices from GABA(B) receptor type 1 subunit knockout mice. The effects of tetrodotoxin, zero added Ca(2+) with/without intracellular Ca(2+) store depletor cyclopiazonic acid or vacuolar H-ATPase inhibitor bafilomycin A1 indicate that GHB-evoked Ca(2+) transients depend on external Ca(2+) and intracellular Ca(2+) stores, but not on vesicular transmitter release. GHB-induced astrocytic Ca(2+) transients were not affected by the GHB receptor-specific antagonist NCS-382, suggesting the presence of a novel NCS-382-insensitive target for GHB in astrocytes. The activation of astrocytes by GHB implies their involvement in physiological actions of GHB. Our findings disclose a novel profile of GHB action in the nucleus accumbens. Here, unlike in other brain areas, GHB does not act on GABA(B) receptors, but activates an NCS-382 insensitive GHB-specific target in a subpopulation of astrocytes. The lack of either post- or presynaptic effects on medium spiny neurons in the nucleus accumbens distinguishes GHB from many drugs and natural rewards with addictive properties and might explain why GHB has only a weak reinforcing capacity.

  3. Gamma-butyrolactone (GBL) disruption of passive avoidance learning in the day-old chick appears to be due to its effect on GABAB not gamma-hydroxybutyric [corrected] acid (GHB) receptors.

    PubMed

    Sherry, Joanne M; Hazi, Agnes; Hale, Mathew W; Milsome, Sarah L; Crowe, Simon F

    2009-02-11

    Gamma-butyrolactone (GBL) is a prodrug to gamma-hydroxybutyric acid (GHB) and metabolises to GHB when ingested. Discrimination stimulus studies report generalisation of effects of GHB to GBL. While amnesia is one of the most commonly reported symptoms of GHB's ingestion in human users, as yet few studies have examined this effect. Although an endogenous GHB specific receptor is present in the brain, several studies have indicated that the clinical effects of exogenous doses of GBL/GHB are due to its action on GABA(B) receptors rather than on the GHB receptor. In this series of studies, New Hampshire x White leghorn cockerels were trained using a modified version of the passive avoidance learning task. Subcutaneous injections of GBL induced a memory deficit by 10 min post-training, which persisted for at least 24 h. No effect on memory was seen with administration of the specific GHB agonist NCS-356 (gamma-p-chlorophenyl-trans-4-hydroxycrotonate). The GBL-induced memory deficit appeared similar to the deficit produced by baclofen, where the antagonist facilitated learning. Additionally, GBL-induced memory deficit was ameliorated by application of a GABA(B) antagonist. The results support the hypothesis that GBL exerts its influence on memory via the GABA(B) receptor rather than by the specific GHB receptor.

  4. Gamma-hydroxybutyrate enhances mood and prosocial behavior without affecting plasma oxytocin and testosterone.

    PubMed

    Bosch, Oliver G; Eisenegger, Christoph; Gertsch, Jürg; von Rotz, Robin; Dornbierer, Dario; Gachet, M Salomé; Heinrichs, Markus; Wetter, Thomas C; Seifritz, Erich; Quednow, Boris B

    2015-12-01

    Gamma-hydroxybutyrate (GHB) is a GHB-/GABAB-receptor agonist. Reports from GHB abusers indicate euphoric, prosocial, and empathogenic effects of the drug. We measured the effects of GHB on mood, prosocial behavior, social and non-social cognition and assessed potential underlying neuroendocrine mechanisms. GHB (20mg/kg) was tested in 16 healthy males, using a randomized, placebo-controlled, cross-over design. Subjective effects on mood were assessed by visual-analogue-scales and the GHB-Specific-Questionnaire. Prosocial behavior was examined by the Charity Donation Task, the Social Value Orientation test, and the Reciprocity Task. Reaction time, memory, empathy, and theory-of-mind were also tested. Blood plasma levels of GHB, oxytocin, testosterone, progesterone, dehydroepiandrosterone (DHEA), cortisol, aldosterone, and adrenocorticotropic-hormone (ACTH) were determined. GHB showed stimulating and sedating effects, and elicited euphoria, disinhibition, and enhanced vitality. In participants with low prosociality, the drug increased donations and prosocial money distributions. In contrast, social cognitive abilities such as emotion recognition, empathy, and theory-of-mind, and basal cognitive functions were not affected. GHB increased plasma progesterone, while oxytocin and testosterone, cortisol, aldosterone, DHEA, and ACTH levels remained unaffected. GHB has mood-enhancing and prosocial effects without affecting social hormones such as oxytocin and testosterone. These data suggest a potential involvement of GHB-/GABAB-receptors and progesterone in mood and prosocial behavior. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. BEHAVIORAL EFFECTS OF GAMMA-HYDROXYBUTYRATE (GHB) IN HUMANS

    PubMed Central

    Oliveto, Alison; Gentry, W. Brooks; Pruzinsky, Rhonda; Gonsai, Kishorchandra; Kosten, Thomas R.; Martell, Bridget; Poling, James

    2010-01-01

    Despite the therapeutic use and abuse potential of gamma-hydroxybutyrate (GHB or Xyrem), relatively few studies have examined the behavioral effects of GHB in humans under controlled laboratory conditions. Thus, this eight-session study examined in 10 non substance-abusing volunteers the behavioral effects of GHB at each of the following doses: 0, 0.32, 0.56, 0.75, 1.0, 1.8, 2.4, 3.2 g/70 kg, p.o.. Order of dose testing was random, except that the first two participants received active doses in ascending order and 2.4 g/70 kg was always tested before 3.2 g/70 kg. Prior to drug administration and at several post-drug time points, self-report, observer-report, physiological, and psychomotor performance measures were obtained. Analyses based on area under the curve showed that GHB produced dose-related increases in subjective ratings of sedative-like, stimulant-like, positive mood, and dissociative effects, but no changes in psychomotor performance measures or blood pressure. Analyses based on peak effects generally showed dose-related increases in ratings indicating sedative-like, dissociative, and drug liking, although some measures showed U-shaped dose-related changes. These initial findings suggest that GHB at doses of 0.32–3.2 g/70 kg produces dissociative, sedating and some stimulant-like effects in humans without a history of sedative abuse. PMID:20526195

  6. Significance of gamma-hydroxybutyric acid in the brain.

    PubMed

    Tunnicliff, G

    1992-11-01

    1. Administration of the endogenous compound gamma-hydroxybutyric acid (GHB) can induce a sleep-like state in experimental animals and, indeed, it has been used as a general anaesthetic in clinical medicine. 2. Although GHB appears to be a CNS depressant, there is evidence it possesses epileptiform activity resembling petit mal epilepsy. In the brain GHB is evidently derived from GABA, the final step being catalyzed by succinic semialdehyde reductase, a cytosolic NADP(+)-dependent enzyme. 3. Two different oxidoreductases, GHB dehydrogenase and hydroxyacid-ketoacid dehydrogenase, acting independently, are responsible for the reverse reaction when GHB is being metabolically inactivated. 4. Brain contains a Na(+)-dependent GHB uptake system which exhibits two components, one with a Km of 46 microM and the other with a Km of 325 microM. GHB also binds to receptor sites in brain homogenates and exhibits two distinct affinities. One binding site displays a Kd of 95 nM whereas the second site has a Kd of 16 microM. Binding to both sites is inhibited in the presence of NCS-382, a GHB receptor antagonist. 5. GHB might play a role as a neurotransmitter, particularly being involved in influencing dopamine release in the substantia nigra.

  7. Gamma-Hydroxybutyrate inhibits excitatory postsynaptic potentials in rat hippocampal slices.

    PubMed

    Berton, F; Brancucci, A; Beghè, F; Cammalleri, M; Demuro, A; Francesconi, W; Gessa, G L

    1999-09-10

    Gamma-Hydroxybutyrate (GHB) has been shown to mimic different central actions of ethanol, to suppress alcohol withdrawal syndrome, and to reduce alcohol consumption both in rats and in humans. The aim of the present study was to determine if GHB shared with alcohol the ability to inhibit glutamate action at both NMDA and AMPA/kainate receptors. The NMDA or the AMPA/kainate receptors-mediated postsynaptic potentials were evoked in CA1 pyramidal neurons by stimulation of Schaffer-collateral commissural fibers in the presence of CGP 35348, bicuculline to block the GABA(B) and GABA(A) receptors, and 10 microM 6,7-dinitroquinoxaline-2,3-dione (DNQX) or 30 microM DL-2-amino-5-phosphonovalerate (d-APV) to block AMPA/kainate or NMDA receptors, respectively. GHB (600 microM) produced a depression of both NMDA and AMPA/kainate receptors-mediated excitatory postsynaptic potentials with recovery on washout. The GHB receptors antagonist, NCS-382, at the concentration of 500 microM had no effect per se on these responses but prevented the depressant effect of GHB (600 microM) on the NMDA and AMPA/kainate-mediated responses. In the paired-pulse experiments, GHB (600 microM) depressed the amplitude of the first and the second evoked AMPA/kainate excitatory postsynaptic potentials, and significantly increased the paired-pulse facilitation (PPF). These results suggest that GHB inhibits excitatory synaptic transmission at Schaffer-collateral commissural-pyramidal neurons synapses by decreasing the probability of release of glutamate.

  8. Abuse and therapeutic potential of gamma-hydroxybutyric acid.

    PubMed

    Galloway, G P; Frederick-Osborne, S L; Seymour, R; Contini, S E; Smith, D E

    2000-04-01

    Gamma-hydroxbutyric acid is a compound found in mammalian brain that is structurally related to the neurotransmitters gamma-aminobutyric acid and glutamic acid. Gamma-hydroxybutyric acid effects dopaminergic systems in the brain and may be a neurotransmitter. Gamma-hydroxybutyric acid was first reported as a drug of abuse in 1990 and continues to be abused by bodybuilders, participants of "rave" dance parties, and polydrug abusers. Physical dependence can develop after prolonged, high-dose use, and overdoses have been widely reported. Its use in sexual assaults as a "date rape" drug and availability on the internet have recently emerged. Gamma-hydroxybutyric acid has established efficacy as an anesthetic agent, and preliminary evidence supports its utility in the treatment of alcohol dependence, opiate dependence, and narcolepsy.

  9. 21 CFR 1304.26 - Additional recordkeeping requirements applicable to drug products containing gamma-hydroxybutyric...

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... to drug products containing gamma-hydroxybutyric acid. 1304.26 Section 1304.26 Food and Drugs DRUG....26 Additional recordkeeping requirements applicable to drug products containing gamma-hydroxybutyric....22, practitioners dispensing gamma-hydroxybutyric acid that is manufactured or distributed in...

  10. 21 CFR 1304.26 - Additional recordkeeping requirements applicable to drug products containing gamma-hydroxybutyric...

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... to drug products containing gamma-hydroxybutyric acid. 1304.26 Section 1304.26 Food and Drugs DRUG....26 Additional recordkeeping requirements applicable to drug products containing gamma-hydroxybutyric....22, practitioners dispensing gamma-hydroxybutyric acid that is manufactured or distributed in...

  11. Preference for Gamma-Hydroxybutyrate (GHB) in Current Users

    ERIC Educational Resources Information Center

    Roll, John M.; Newton, Thomas; Chudzynski, Joy; Cameron, Jennifer M.; McPherson, Sterling; Fong, Tim; Torrington, Matt

    2012-01-01

    Gamma-hydroxybutyrate (GHB) is a drug with significant abuse potential. The present study aimed to assess the relative value of escalating doses of GHB to current GHB users via the Multiple Choice Procedure (MCP), and to validate that the dose rated highest with the MCP would be self-administered at a greater rate than placebo. Participants were 5…

  12. Preference for Gamma-Hydroxybutyrate (GHB) in Current Users

    ERIC Educational Resources Information Center

    Roll, John M.; Newton, Thomas; Chudzynski, Joy; Cameron, Jennifer M.; McPherson, Sterling; Fong, Tim; Torrington, Matt

    2012-01-01

    Gamma-hydroxybutyrate (GHB) is a drug with significant abuse potential. The present study aimed to assess the relative value of escalating doses of GHB to current GHB users via the Multiple Choice Procedure (MCP), and to validate that the dose rated highest with the MCP would be self-administered at a greater rate than placebo. Participants were 5…

  13. Central and peripheral metabolic changes induced by gamma-hydroxybutyrate.

    PubMed

    Luca, Gianina; Vienne, Julie; Vaucher, Angélique; Jimenez, Sonia; Tafti, Mehdi

    2015-02-01

    Gamma-hydroxybutyrate (GHB) was originally introduced as an anesthetic but was first abused by bodybuilders and then became a recreational or club drug.1 Sodium salt of GHB is currently used for the treatment of cataplexy in patients with narcolepsy. The mode of action and metabolism of GHB is not well understood. GHB stimulates growth hormone release in humans and induces weight loss in treated patients, suggesting an unexplored metabolic effect. In different experiments the effect of GHB administration on central (cerebral cortex) and peripheral (liver) biochemical processes involved in the metabolism of the drug, as well as the effects of the drug on metabolism, were evaluated in mice. C57BL/6J, gamma-aminobutyric acid B (GABAB) knockout and obese (ob/ob) mice were acutely or chronically treated with GHB at 300 mg/kg. Respiratory ratio decreased under GHB treatment, independent of food intake, suggesting a shift in energy substrate from carbohydrates to lipids. GHB-treated C57BL/6J and GABAB null mice but not ob/ob mice gained less weight than matched controls. GHB dramatically increased the corticosterone level but did not affect growth hormone or prolactin. Metabolome profiling showed that an acute high dose of GHB did not increase the brain GABA level. In the brain and the liver, GHB was metabolized into succinic semialdehyde by hydroxyacid-oxoacid transhydrogenase. Chronic administration decreased glutamate, s-adenosylhomocysteine, and oxidized gluthathione, and increased omega-3 fatty acids. Our findings indicate large central and peripheral metabolic changes induced by GHB with important relevance to its therapeutic use. © 2015 Associated Professional Sleep Societies, LLC.

  14. Baclofen antagonises intravenous self-administration of gamma-hydroxybutyric acid in mice.

    PubMed

    Fattore, L; Cossu, G; Martellotta, M C; Deiana, S; Fratta, W

    2001-07-20

    gamma-Hydroxybutyric acid (GHB) is a widely used recreational drug known to exert positive reinforcing effects in animals and humans. The GABA(B) receptor agonist baclofen has been proved to possess antimotivational effect and to inhibit alcohol, cocaine, heroin and nicotine intake. In the present study we evaluated the effect of baclofen on i.v. self-administration of GHB in drug-naive mice under a fixed-ratio (FR-1) schedule of reinforcement and nose-poking-like response as operandum. Results show that baclofen was able to completely prevent GHB seeking behaviour, decreasing the rate of responding to basal values, without showing any reinforcing properties when made contingent on nose-poking response. Our findings demonstrate that baclofen antagonises GHB i.v. self-administration, supporting an important role for the GABA(B) receptor in reward-related mechanisms underlying addictive behaviour.

  15. Therapeutic intervention in mice deficient for succinate semialdehyde dehydrogenase (gamma-hydroxybutyric aciduria).

    PubMed

    Gupta, Maneesh; Greven, Rachel; Jansen, Erwin E W; Jakobs, Cornelis; Hogema, Boris M; Froestl, Wolfgang; Snead, O Carter; Bartels, Hilke; Grompe, Markus; Gibson, K Michael

    2002-07-01

    Therapeutic intervention for human succinic semialdehyde dehydrogenase (SSADH) deficiency (gamma-hydroxybutyric aciduria) has been limited to vigabatrin (VGB). Pharmacologically, VGB should be highly effective due to 4-aminobutyrate-transaminase (GABA-transaminase) inhibition, lowering succinic semialdehyde and, thereby, gamma-hydroxybutyric acid (GHB) levels. Unfortunately, clinical efficacy has been limited. Because GHB possesses a number of potential receptor interactions, we addressed the hypothesis that antagonism of these interactions in mice with SSADH deficiency could lead to the development of novel treatment strategies for human patients. SSADH-deficient mice have significantly elevated tissue GHB levels, are neurologically impaired, and die within 4 weeks postnatally. In the current report, we compared oral versus intraperitoneal administration of VGB, CGP 35348 [3-aminopropyl(diethoxymethyl)phosphinic acid, a GABA(B) receptor antagonist], and the nonprotein amino acid taurine in rescue of SSADH-deficient mice from early death. In addition, we assessed the efficacy of the specific GHB receptor antagonist NCS-382 (6,7,8,9-tetrahydro-5-[H]benzocycloheptene-5-ol-6-ylideneacetic acid) using i.p. administration. All interventions led to significant lifespan extension (22-61%), with NCS-382 being most effective (50-61% survival). To explore the limited human clinical efficacy of VGB, we measured brain GHB and gamma-aminobutyric acid (GABA) levels in SSADH-deficient mice receiving VGB. Whereas high-dose VGB led to the expected elevation of brain GABA, we found no parallel decrease in GHB levels. Our data indicate that, at a minimum, GHB and GABA(B) receptors are involved in the pathophysiology of SSADH deficiency. We conclude that taurine and NCS-382 may have therapeutic relevance in human SSADH deficiency and that the poor clinical efficacy of VGB in this disease may relate to an inability to decrease brain GHB concentrations.

  16. Drug-facilitated sexual assault involving gamma-hydroxybutyric acid.

    PubMed

    Stillwell, Matthew E

    2002-09-01

    The first case involving an alleged sexual assault linked to the use of gamma-hydroxybutyric acid (GHB) in Oklahoma is reported. A-48-year-old Caucasian woman taking amitriptyline was known to have voluntarily ingested a sports drink containing a relaxing health product. She purportedly experienced unconsciousness that persisted for approximately 4 h. The toxicological testing on urine identified GHB, amitriptyline, and nortriptyline using a capillary Hewlett-Packard 6890 gas chromatograph coupled to a Hewlett-Packard 5973 mass selective detector (MSD). The GHB concentration in urine was 26.9 microg/mL. Urine concentrations of amitriptyline and nortriptyline were not determined. The analytical method used for identifying and quantitating GHB can be applied to matters of forensic interests.

  17. Mechanisms underlying the sympathomimetic cardiovascular responses elicited by gamma-hydroxybutyrate.

    PubMed

    Hicks, Alissa R; Kapusta, Daniel R; Varner, Kurt J

    2004-12-01

    Gamma-hydroxybutyrate (GHB) is generally thought to be a central nervous system depressant; however, GHB also has sympathomimetic cardiovascular actions. Radio telemetry was used to record the cardiovascular responses elicited by GHB (180-1000 mg/kg IV) in conscious rats. GHB elicited increases in mean arterial pressure (MAP) (24 +/- 3 to 60 +/- 5 mm Hg) lasting from 28 +/- 8 to 227 +/- 37 minutes. GHB (560 and 1000 mg/kg IV) also elicited a prolonged tachycardic response (85 +/- 23 and 95 +/- 22 bpm). The hypertension and tachycardia elicited by GHB (560 mg/kg) were reversed by the intravenous and intracerebroventricular administration of the GABAb receptor antagonist CGP 35348. CGP 35348 also reversed GHB-mediated increases in renal sympathetic nerve activity (RSNA). Administration of the purported GHB receptor antagonist NCS-382 reversed the increase in heart rate but not the pressor response elicited by GHB in telemetered rats. These data indicate that the intravenous administration of GHB markedly increases MAP, heart rate, and RSNA in conscious rats via activation of central GABAb receptors. In addition, GHB receptors appear to selectively mediate the increase in heart rate elicited by large doses of GHB.

  18. Enhancing sexual desire and experience: an investigation of the sexual correlates of gamma-hydroxybutyrate (GHB) use.

    PubMed

    Kapitány-Fövény, Máté; Mervó, Barbara; Corazza, Ornella; Kökönyei, Gyöngyi; Farkas, Judit; Urbán, Róbert; Zacher, Gábor; Demetrovics, Zsolt

    2015-07-01

    Various studies have dealt with gamma-hydroxybutyrate's (GHB) potential role in sexual assaults, while the sexual correlates of intentional recreational GHB use have not well been highlighted. Our study aims to explore GHB's sexual effects, the patterns of choice of sexual partners, the frequency of experienced blackouts, and endured sexual or acquisitory crimes as a result of GHB use. Sixty recreational GHB users filled out a questionnaire on experienced subjective, somatic, and sexual effects of GHB, the frequency of blackouts due to their GHB use, and items on their sexual experiences in relation to GHB use. Of the sample, 25.9% reported increased sexual arousal as well as more intense attraction towards their sexual partners and increased sexual openness when using GHB; 34.8% had sexual intercourse with strangers, or with others, but not with their partners when using GHB; and 8.6% were victims of acquisitory crimes, whereas 3.4% were victims of a sexual assault. Furthermore, 24.6% typically experienced blackouts when using GHB. Gamma-hydroxybutyrate seems to be a potential substitute for both stimulant and depressant substances. Increased sexual desire and disinhibition may lead to a more frequent and potentially more riskful sexual activity. Experienced blackouts need to be considered as risk factors for suffering sexual or acquisitory crimes. Copyright © 2015 John Wiley & Sons, Ltd.

  19. EXPERIENCES OF GAMMA HYDROXYBUTYRATE (GHB) INGESTION: A FOCUS GROUP STUDY

    PubMed Central

    Barker, Judith C.; Harris, Shana L.; Dyer, Jo E.

    2008-01-01

    GHB (gamma hydroxybutyrate) is a significant new drug of abuse added to the United States Controlled Substance Act in 2000. The majority of the published literature on GHB consists of clinical case reports, mainly from emergency departments, and a collection of laboratory-based studies, focused mainly on anesthesia. While comments about the various experiences and behaviors of human users are often included in such studies or reports, these aspects of GHB are only just beginning to be systematically investigated or detailed. Reported here are data from a qualitative study using focus group methods on the consumption habits, experiences, and beliefs of GHB users. A total of 51 people, 30 men and 21 women, mean age of 31.1±7.6 years (range 18 – 52 years), who report having used GHB for an average of 4.3±2.5 years (range 1–11 years), were interviewed in 10 separate groups held in 2004. This paper discusses broadly the general experience of the GHB ‘high,’ major perceived benefits including sexual responses to the drug, perceived risks and dangers of ingestion, co-ingestion, and various contexts of use. The paper concludes with a discussion of the implications drawn from this information for clinicians treating patients who use GHB. PMID:17703706

  20. Gamma hydroxybutyrate: an ethnographic study of recreational use and abuse.

    PubMed

    Lee, Steven J; Levounis, Petros

    2008-09-01

    Gamma hydroxybutyrate (GHB) is a psychoactive substance with complex neurophysiological activity and significant potential for abuse, addiction, and dangerous toxicity. In this study, a semistructured interview was administered to 17 subjects to investigate GHB use, including: manner of use; setting; positive and negative consequences; other drug history; and sexual practices. Respondents were overwhelmingly male, but otherwise had a broad demographic background. Settings varied from nightclubs to private use at home. There was significant variability in the drug obtained, which subjects found problematic because of the narrow therapeutic window and ease of accidental overdose. Common positive experiences included increased sexual desire, decreased sexual inhibitions, and decreased anxiety. Common negative consequences included oversedation, loss of consciousness, motor incoordination, and mental confusion. Nine subjects reported that they would use GHB again, some despite severe negative consequences. Although most subjects reported negative experiences, only three felt their use was problematic, and none sought treatment for GHB abuse or addiction. Subjects were highly drug-experienced, most commonly using MDMA, ketamine, cocaine, alcohol, and methamphetamine. Some reported that GHB could cause poor decision making in sexual situations. This effect has significant ramifications for issues such as date rape and control of sexually transmitted diseases, such as HIV.

  1. Preference for gamma-hydroxybutyrate (GHB) in current users.

    PubMed

    Roll, John M; Newton, Thomas; Chudzynski, Joy; Cameron, Jennifer M; McPherson, Sterling; Fong, Timothy; Torrington, Matt

    2012-05-01

    Gamma-hydroxybutyrate (GHB) is a drug with significant abuse potential. The present study aimed to assess the relative value of escalating doses of GHB to current GHB users via the Multiple Choice Procedure (MCP), and to validate that the dose rated highest with the MCP would be self-administered at a greater rate than placebo. Participants were 5 current GHB users who were not currently trying to stop using GHB. To examine the value of escalating doses of GHB, the following doses of GHB were used: 0 (placebo), 12.5, 25, 37.5, and 50 mg/kg. Participants typically assigned higher doses of GHB had higher crossover points on the MCP. During choice sessions, participants made repeated choices between administering GHB, placebo or nothing. All participants selected GHB exclusively (5 out of 5 instances) except for one participant who selected GHB on 4 out of 5 instances, thus 96% (i.e., 24/25) of choices were for active GHB. Based on these data, GHB appears likely to function as a dose-dependent reinforcer for humans based on our sample.

  2. The clinical development of gamma-hydroxybutyrate (GHB).

    PubMed

    Wedin, Gregory P; Hornfeldt, Carl S; Ylitalo, Lisa M

    2006-01-01

    The discovery of gamma-hydroxybutyrate (GHB) over 40 years ago led to its immediate use as a general anesthetic agent. Subsequent research demonstrated that GHB is an endogenous compound in the mammalian brain and current research suggests that GHB is a probable neurotransmitter. In the United States, reports of anabolic effects lead to its misuse among body builders during the 1980's while the intoxicating properties of the drug lead to its popularization as a substance of abuse during the 1990's. GHB became associated with reports of drug-facilitated sexual assault and cases of physical dependence and withdrawal. Efforts to ban GHB caused increased use of GHB analogues and pro-drugs. Against this backdrop, GHB was being developed for the treatment of narcolepsy, leading to the approval of Xyrem (sodium oxybate) oral solution in 2002 for the treatment of cataplexy in patients with narcolepsy. A risk management program permits the safe handling and distribution of the approved product, minimizes the risk for diversion, provides professional and patient education about the risks and benefits of sodium oxybate, and includes physician and patient registries. Post-marketing surveillance indicates sodium oxybate has an acceptable safety profile and presents minimal risk for the development of physical dependence.

  3. Psychiatric comorbidity, psychological distress, and quality of life in gamma-hydroxybutyrate-dependent patients.

    PubMed

    Kamal, Rama M; Dijkstra, Boukje A G; de Weert-van Oene, Gerdien H; van Duren, Josja A M; de Jong, Cornelis A J

    2017-01-01

    Understanding the psychiatric state and psychological distress level of patients with gamma-hydroxybutyrate dependence is important to develop effective detoxification and relapse management methods. The aim of the current study was to assess the prevalence among gamma-hydroxybutyrate-dependent individuals of psychiatric comorbidity and psychological distress levels and their association with the individuals' pattern of misuse and quality of life. There were 98 patients tested with the Mini International Neuropsychiatric Interview-plus, the Brief Symptom Inventory, the Depression Anxiety Stress scale, and the EuroQoL-5D as a part of the Dutch gamma-hydroxybutyrate detoxification monitor in 7 addiction treatment centers. Participants were selected from those undergoing inpatient gamma-hydroxybutyrate detoxification treatment between March 2011 and September 2012. Males accounted for 68% of the participants and the average age was 28-years-old. A high rate of psychiatric comorbidity (79%) was detected, including anxiety (current 38%, lifetime 40%), mood (13%, 31%), and psychotic disorders (13%, 21%). The level of psychological distress was significantly higher than the standard outpatient reference group, especially in patients with current psychiatric comorbidity (Brief Symptom Inventory Global Severity Index mean 1.61 versus 1.09, p ≤ 0.01). Increased gamma-hydroxybutyrate misuse (higher dose and shorter interval between doses) was associated with the presence of lifetime psychosis, current mood disorders (rpb = 0.23, p = 0.025), and psychoticism as a symptom of psychological distress. Current anxiety, mood disorders and high psychological stress had a negative effect on participants' quality of life. Gamma-hydroxybutyrate dependence is characterized by serious psychiatric comorbidity and psychological distress, both of which are, in turn, associated with increased gamma-hydroxybutyrate use and a lower quality of life. This needs to be considered during

  4. Effects of Gamma Hydroxybutyric Acid on Inhibition and Excitation in Rat Neocortex

    PubMed Central

    Li, Qiang; Kuhn, Cynthia M.; Wilson, Wilkie A.; Lewis, Darrell V.

    2008-01-01

    The mechanism by which the sedative and amnestic recreational drug gamma hydroxybutyric acid (GHB) acts is controversial. Some studies indicate that it acts at its unique receptor, while others demonstrate effects mediated through the GABAB receptor. We examined the effect of GHB on evoked GABAA receptor mediated mono- and polysynaptic IPSCs as well as on NMDA and AMPA mediated EPSCs in layers II/III pyramidal cells of the frontal cortex of rat brain. One millimolar (mM) GHB suppressed monosynaptic IPSCs by 20%, whereas polysynaptic IPSCs were reduced by 56%. GHB (1mM) also produced a significant suppression of NMDA-mediated EPSCs by 53% compared to 27% suppression of AMPA-mediated EPSCs. All effects of GHB on IPSCs and EPSCs were reversed by the specific GABAB antagonist CGP62349, but not by the GHB receptor antagonist NCS 382. Consistent with a presynaptic site of action, GHB reduced the frequency but not the amplitude of AMPA receptor mediated mEPSCs and had no effect on postsynaptic currents evoked by direct application of NMDA. Finally, even though GHB appeared to be acting at presynaptic GABAB receptors, GHB and the GABAB agonist baclofen appeared to have opposite potencies for depression of NMDA vs AMPA mediated EPSCs. GHB showed a preference for depressing NMDA responses while baclofen more potently suppressed AMPA responses. The suppression of NMDA more than AMPA responses by GHB at intoxicating doses may make it attractive as a recreational drug and may explain why GHB is abused and baclofen is not. PMID:17904295

  5. Safety and tolerability of gamma-hydroxybutyric acid in the treatment of alcohol-dependent patients.

    PubMed

    Beghè, F; Carpanini, M T

    2000-04-01

    Gamma-hydroxybutyric acid (GHB) has been in clinical use in Italy since 1991 for treatment of alcohol dependence. Results of phase III and phase IV studies have shown that the drug is effective and well tolerated in the treatment of alcohol withdrawal syndrome and in reducing alcohol consumption and alcohol craving. Pharmacosurveillance indicates that abuse of gamma-hydroxybutyric acid is a limited phenomenon in clinical settings when the drug is dispensed under strict medical surveillance and entrusted to a referring familiar member of the patient.

  6. GC-MS Analysis of [gamma]-Hydroxybutyric Acid Analogs: A Forensic Chemistry Experiment

    ERIC Educational Resources Information Center

    Henck, Colin; Nally, Luke

    2007-01-01

    An upper-division forensic chemistry experiment is described. It involves using glycolic acid and sodium glycolate as analogs of [gamma]-hydroxybutyric acid and its sodium salt. The experiment shows the use of silylation in GC-MS analysis and gives students the opportunity to work with a commonly used silylating reagent,…

  7. "Grievous bodily harm:" gamma hydroxybutyrate abuse leading to a Wernicke-Korsakoff syndrome.

    PubMed

    Friedman, J; Westlake, R; Furman, M

    1996-02-01

    Gamma-hydroxybutyrate (GHB) is a naturally occurring GABA-like drug used illicitly by bodybuilders. Although there are reports of several cases of GHB abuse, with a variety of nervous system complications, we present the first case associated with a Wernicke-Korsakoff syndrome.

  8. GC-MS Analysis of [gamma]-Hydroxybutyric Acid Analogs: A Forensic Chemistry Experiment

    ERIC Educational Resources Information Center

    Henck, Colin; Nally, Luke

    2007-01-01

    An upper-division forensic chemistry experiment is described. It involves using glycolic acid and sodium glycolate as analogs of [gamma]-hydroxybutyric acid and its sodium salt. The experiment shows the use of silylation in GC-MS analysis and gives students the opportunity to work with a commonly used silylating reagent,…

  9. [EEG changes during sedation with gamma-hydroxybutyric acid].

    PubMed

    Entholzner, E; Mielke, L; Pichlmeier, R; Weber, F; Schneck, H

    1995-05-01

    Gamma-hydroxybutyric acid (GHB) is a naturally occurring transmitter in the mammalian brain, related to sleep regulation and possibly to energy balance in diving or hibernating animals. It has been used for almost 35 years as an intravenous agent for induction of anaesthesia and for long-term sedation. Its convincing pharmacological properties, without serious adverse effects on circulation or respiration, are compromised by its unpredictable duration of action. This is not a major problem with long-term sedation during ICU treatment. GHB has been used with good results for sedation of patients with severe brain injury, where it compares favourably with barbiturates. In animal studies, it seems to possess a protective action against hypoxia on a cellular and whole organ level. However, in some experimental animals GHB has been shown to produce seizure-like activities, and the compound is being used to produce absence-like seizures. GHB has been used in our ICU for years to provide adequate sedation for patients under controlled ventilation or for patients fighting the respirator during spontaneous respiration. No serious side effects were observed in these patients, while in some patients under haemodialysis hypernatraemia and metabolic alkalosis developed; both were reversible after discontinuation of GHB and restriction of additional sodium input (Somsanit, the commercially available GHB preparation in Germany, contains 9.2 mmol sodium/g; the daily dose averages 20-40 g GHB, i.e. 180-370 mmol sodium). PATIENTS AND METHODS. In 31 patients after major abdominal surgery, sedation was established with GHB 50 mg/kg BW injected via perfusion pump over a 20-min period. No centrally acting medication had been given for at least 2 h. A computer-based multichannel EEG system (CATEEM, MediSyst, Linden) was used, allowing for fast Fourier transformation, spectral analysis and topographical brain mapping. EEG during induction of sedation was followed after a baseline EEG (10

  10. gamma-Hydroxybutyrate/sodium oxybate: neurobiology, and impact on sleep and wakefulness.

    PubMed

    Pardi, Daniel; Black, Jed

    2006-01-01

    gamma-Hydroxybutyrate (GHB) is an endogenous short chain fatty acid and a, mostly oral, pharmacological compound that has been utilised in a variety of ways. Endogenously, GHB is synthesised locally within the CNS, mostly from its parent compound GABA. Sodium oxybate is the sodium salt of GHB and is used for the exogenous oral administration of GHB. It is likely that supraphysiological concentrations of GHB from exogenous administration produce qualitatively different neuronal actions than those produced by endogenous GHB concentrations. Evidence suggests a role for GHB as a neuromodulator/neurotransmitter. Under endogenous conditions and concentrations, and depending on the cell group affected, GHB may increase or decrease neuronal activity by inhibiting the release of neurotransmitters that are co-localised with GHB. After exogenous administration, most of the observed behavioural effects appear to be mediated via the activity of GHB at GABA(B) receptors, as long as the concentration is sufficient to elicit binding, which does not happen at endogenous concentrations. Endogenous and exogenous GHB is rapidly and completely converted into CO(2) and H(2)O through the tricarboxylic acid cycle (Krebs cycle). Sodium oxybate has been observed to modulate sleep in nonclinical study participants, and sleep and wakefulness in clinical populations, including groups with insomnia, fibromyalgia and narcolepsy. In narcolepsy, sodium oxybate has shown dose-related effects on various properties of sleep, including increases in slow-wave sleep duration and delta power, and a reduced number of night-time awakenings. Furthermore, multiple measures of daytime sleepiness and cataplexy demonstrated consistent short- and long-term improvement in response to night-time sodium oxybate therapy. The most common reported adverse events include dose-related headache, nausea, dizziness and somnolence.

  11. Recordkeeping and reporting requirements for drug products containing gamma-hydroxybutyric acid (GHB). Final rule.

    PubMed

    2005-01-04

    DEA is amending its regulations to require additional recordkeeping and reporting requirements for drug products containing gamma-hydroxybutyric acid (GHB) for which an application has been approved under the Federal Food, Drug, and Cosmetic Act. DEA makes these changes under section 4 of the "Hillory J. Farias and Samantha Reid Date-Rape Drug Prohibition Act of 2000." These additional requirements are necessary to protect against the diversion of GHB for illicit purposes.

  12. Low-carb diets, fasting and euphoria: Is there a link between ketosis and gamma-hydroxybutyrate (GHB)?

    PubMed

    Brown, Andrew J

    2007-01-01

    Anecdotal evidence links the initial phase of fasting or a low-carbohydrate diet with feelings of well-being and mild euphoria. These feelings have often been attributed to ketosis, the production of ketone bodies which can replace glucose as an energy source for the brain. One of these ketone bodies, beta-hydroxybutyrate (BHB), is an isomer of the notorious drug of abuse, GHB (gamma-hydroxybutyrate). GHB is also of interest in relation to its potential as a treatment for alcohol and opiate dependence and narcolepsy-associated cataplexy. Here I hypothesize that, the mild euphoria often noted with fasting or low-carbohydrate diets may be due to shared actions of BHB and GHB on the brain. Specifically, I propose that BHB, like GHB, induces mild euphoria by being a weak partial agonist for GABA(B) receptors. I outline several approaches that would test the hypothesis, including receptor binding studies in cultured cells, perception studies in trained rodents, and psychometric testing and functional magnetic resonance imaging in humans. These and other studies investigating whether BHB and GHB share common effects on brain chemistry and mood are timely and warranted, especially when considering their structural similarities and the popularity of ketogenic diets and GHB as a drug of abuse.

  13. Determination of gamma-hydroxybutyrate (GHB) and its precursors in blood and urine samples: a salting-out approach.

    PubMed

    Kankaanpää, Aino; Liukkonen, Raija; Ariniemi, Kari

    2007-08-06

    Gamma-hydroxybutyrate (GHB) is an increasingly popular drug of abuse that causes stimulation, euphoria, anxiolysis or hypnosis, depending on the dose used. Low doses of the drug are used recreationally, and also implicated in drug-facilitated sexual assaults. Because of the unusually steep dose-response curves, accidental GHB overdosing, leading to coma, seizures or death can occur. Being a controlled substance, GHB is often substituted with its non-scheduled precursors gamma-butyrolactone (GBL) and 1,4-butanediol (BD), which are rapidly metabolized into GHB in the body. Here we describe an assay for GHB, GBL and BD in blood and/or urine samples. GHB and BD were extracted from diluted 200 microL aliquots of samples with t-butylmethylether (plus internal standard benzyl alcohol) in test tubes preloaded with NaCl. After acidification and centrifugation the solvent phase was transferred to a test tube preloaded with Na(2)SO(4), incubated for 30 min, centrifuged again, and evaporated in vacuum. The residue was mixed with N-methyl-N-trimethylsilyl-trifluoroacetamide (MSTFA) in acetonitrile, and injected into a GC-MS. When analyzing GBL, the salting-out step was omitted, and analysis was performed with a GC-FID apparatus. As revealed by the validation data this procedure is suitable for quantitative determination of GHB and its precursors in blood and/or urine samples.

  14. Trans-gamma-hydroxycrotonic acid binding sites in brain: evidence for a subpopulation of gamma-hydroxybutyrate sites.

    PubMed

    Hechler, V; Schmitt, M; Bourguignon, J J; Maitre, M

    1990-03-02

    Trans-gamma-hydroxycrotonate (THCA), a compound naturally present in rat brain, possesses high-affinity binding sites with a heterogeneous distribution which are superimposable with those for gamma-hydroxybutyrate (GHB). Binding studies of THCA on rat brain membranes revealed two binding components, one of high affinity (Kd1, 7 nM, Bmax1 42 fmol/mg protein) and the other of low affinity (Kd2, 2 microM, Bmax2 13 pmol/mg protein). Displacement curves of [3H]THCA by THCA and GHB or of [3H]GHB by THCA are in favour of the existence of a specific high affinity site for THCA. Quantitative autoradiography with image analysis of [3H]THCA binding in rat brain slices indicated that [3H]THCA high affinity binding was displaced at a lower potency by GHB. THCA showed also some selectivity in displacing [3H]GHB from its high affinity binding site (Kd = 95 nM). This mutual overlap favours a subpopulation of GHB receptors, which have THCA as a natural ligand, showing partial agonistic properties compared to GHB. The functional significance of this result remains unknown.

  15. Illicit gamma-hydroxybutyrate (GHB) and pharmaceutical sodium oxybate (Xyrem): differences in characteristics and misuse.

    PubMed

    Carter, Lawrence P; Pardi, Daniel; Gorsline, Jane; Griffiths, Roland R

    2009-09-01

    There are distinct differences in the accessibility, purity, dosing, and misuse associated with illicit gamma-hydroxybutyrate (GHB) compared to pharmaceutical sodium oxybate. Gamma-hydroxybutyrate sodium and sodium oxybate are the chemical and drug names, respectively, for the pharmaceutical product Xyrem (sodium oxybate) oral solution. However, the acronym GHB is also used to refer to illicit formulations that are used for non-medical purposes. This review highlights important differences between illicit GHB and sodium oxybate with regard to their relative abuse liability, which includes the likelihood and consequences of abuse. Data are summarized from the scientific literature; from national surveillance systems in the U.S., Europe, and Australia (for illicit GHB); and from clinical trials and post-marketing surveillance with sodium oxybate (Xyrem). In the U.S., the prevalence of illicit GHB use, abuse, intoxication, and overdose has declined from 2000, the year that GHB was scheduled, to the present and is lower than that of most other licit and illicit drugs. Abuse and misuse of the pharmaceutical product, sodium oxybate, has been rare over the 5 years since its introduction to the market, which is likely due in part to the risk management program associated with this product. Differences in the accessibility, purity, dosing, and misuse of illicit GHB and sodium oxybate suggest that risks associated with illicit GHB are greater than those associated with the pharmaceutical product sodium oxybate.

  16. Effect of gamma-hydroxybutyrate on keratinocytes proliferation: A preliminary prospective controlled study in severe burn patients

    PubMed Central

    Rousseau, Anne-Françoise; Bargues, Laurent; Bever, Hervé Le; Vest, Philippe; Cavalier, Etienne; Ledoux, Didier; Piérard, Gérald E.; Damas, Pierre

    2014-01-01

    Background: Hypermetabolism and hyposomatotropism related to severe burns lead to impaired wound healing. Growth hormone (GH) boosts wound healing notably following stimulation of the production of insulin-like growth factor-1 (IGF1), a mitogen factor for keratinocytes. Gamma-hydroxybutyrate (GHB) stimulates endogenous GH secretion. Aim: To assess effects of GHB sedation on keratinocytes proliferation (based on immunohistochemical techniques). Design: Monocentric, prospective, controlled trial. Materials and Methods: Patients (aging 18-65 years, burn surface area >30%, expected to be sedated for at least one month) were alternately allocated, at the 5th day following injury, in three groups according to the intravenous GHB dose administered for 21 days: Evening bolus of 50 mg/kg (Group B), continuous infusion at the rate of 10 mg/kg/h (Group C), or absence of GHB (Group P). They all received local standard cares. Immunohistochemistry (Ki67/MIB-1, Ulex europaeus agglutinin-1 and Mac 387 antibodies) was performed at D21 on adjacent unburned skin sample for assessing any keratinocyte activation. Serum IGF1 levels were measured at initiation and completion of the protocol. Statistical Analysis: Categorical variables were compared with Chi-square test. Comparisons of medians were made using Kruskal-Wallis test. Post hoc analyses were performed using Mann-Whitney test with Bonferroni correction for multiple comparisons. A P < 0.05 was considered to be statistically significant. Results: A total of 14 patients completed the study (Group B: n = 5, Group C: n = 5, Group P: n = 4). Continuous administration of GHB was associated with a significant higher Ki67 immunolabeling at D21 (P = 0.049) and with a significant higher increase in the IGF1 concentrations at D21 (P = 0.024). No adverse effects were disclosed. Conclusions: Our preliminary data support a positive effect of GHB on keratinocyte proliferation and are encouraging enough to warrant large prospective studies. PMID

  17. Growth hormone response to hypoglycemia under gamma-hydroxybutyrate narco-analgesia in the rat.

    PubMed

    Bluet-Pajot, M T; Schaub, C; Nassiet, J

    1978-01-01

    Plasma immuno-reactive growth-hormone (RIA-GH) concentrations were investigated under in vivo continuous blood glucose (BG) monitoring after administration of gamma-hydroxybutyrate (GHB) as well as during spontaneous or insulin-induced hypoglycemia. During the narco-analgesia by GHB a marked secretory episode is consistently observed. This secretion peak is not accurately time related with GHB administration and seems to fade off in aging animals. Strictly controlled hypoglycemia elicits a consistent and specific GH release. In contrast deep hypoglycemic levels resulting in a state of metabolic stress inhibit GH secretion. Our results suggest that previous data on the GH regulation pattern during hypoglycemia may depend upon the anesthetic used and/or nonspecific stress responses following deep hypoglycemia. The above mentioned experimental conditions indicate that GH metabolic regulation is not fundamentally different in rodents and primates.

  18. [The knowledge about gamma-hydroxybutyric acid as by students of Physical Education Academy].

    PubMed

    Chwaluk, Paweł; Chwaluk, Agnieszka; Parnicki, Florian

    2009-01-01

    Gamma-hydroxybutyric acid is a substance stealthily used by criminals to facilitate sexual assaults. It is also known as doping agent in sports. Physical Education Academies should prepare their graduates to be educators for young people, their trainers, organizers of sports and recreational events. Second year students of two majors: physical education and tourism and recreation were surveyed by means of questionnaire on "date-rape drug". As much as 320 among 327 students surveyed had heard about "date-rape drug". However their knowledge on it was shallow and unsystematic. None of the surveyed knew that the substance of "date-rape drug" could also be used as a doping agent. Only 31% of respondents were aware of existence of the test to detect "date-rape drug" in drinks. Physical Education Academy students should be thoroughly and relevantly educated on the matter of pharmacologic doping agents and drugs endangerment.

  19. [Gamma-hydroxybutyric acid (GHB) dependence and the GHB withdrawal syndrome: diagnosis and treatment].

    PubMed

    van Noorden, Martijn S; Kamal, Rama; de Jong, Cor A J; Vergouwen, A C M Ton; Zitman, Frans G

    2010-01-01

    Gamma-hydroxybutyric acid (GHB) is a neurotransmitter that occurs naturally in the brain and is increasingly being used as a 'party drug' because of its relaxing and euphoria-inducing effects. GHB has a limited medical use in the treatment of narcolepsy. GHB-intoxications occur often in non-medical use, and generally result in a coma of short duration. GHB use several times a day can lead to tolerance and dependence. After sudden cessation or reduction of intensive GHB use, a severe withdrawal syndrome may occur with symptoms varying from tremor, anxiety and agitation to autonomic instability, hallucinations and delirium. Treatment of the GHB withdrawal syndrome consists of supportive care and benzodiazepines, often in high doses. The controlled detoxification of GHB using pharmaceutical GHB in an adjusted dose is currently being investigated in the Netherlands. There is no literature concerning the treatment of patients following GHB intoxication or after detoxification.

  20. A Web-Based Study of Gamma Hydroxybutyrate (GHB): Patterns, Experiences, and Functions of Use

    PubMed Central

    Stein, LAR; Lebeau, Rebecca; Clair, Mary; Martin, Rosemarie; Bryant, Monte; Storti, Susan; Monti, Peter

    2011-01-01

    GHB (gamma hydroxybutyrate) was developed as a general anesthetic. Due to dosing difficulty and side effects, regular use was discontinued. Medical uses include treating sleep and alcohol disorders. In the 1990s, it was promoted as a supplement and taken to improve mood and sex. GHB and its analogs (gamma butyrolactone and butanediol) were widely available until federal regulations were put into effect with mounting evidence of adverse events. This survey (N = 61) study was conducted to assess patterns, experiences, and functions of use. Much of what is understood regarding GHB treatment is based on hospital case studies for overdose and withdrawal. Not enough is known about prevention, reducing use and associated problems, or relapse. We know little about specific drug effect expectancies, triggers, coping skills, and consequences of use (positive/negative). While the drug treatment literature has a wealth of information to draw upon, GHB-specific information may greatly assist relapse prevention. PMID:21175918

  1. High-Risk Behaviors and Hospitalizations Among Gamma Hydroxybutyrate (GHB) Users

    PubMed Central

    Kim, Susan Y.; Anderson, Ilene B.; Dyer, Jo Ellen; Barker, Judith C.; Blanc, Paul D.

    2008-01-01

    Introduction Little is known about behaviors linked to gamma hydroxybutyrate (GHB) morbidity. Methods We surveyed 131 GHB users, using logistic regression to test the associations between the high risk behaviors and hospital treatment for GHB (26 [20%] of subjects). Results Increased risk of GHB hospital treatment was associated with: co-ingestion of ethanol (OR 5.2; 95% CI 1.7–16), driving under the influence of GHB (OR 3.2; 95%, CI 1.3–7.8), use of GHB to treat withdrawal symptoms (OR 2.9; 95% CI 1.1–7.9), and co-ingestion of ketamine (OR 2.7; 95% CI 1.1–6.7). Conclusion Targeted prevention activities could focus on selected high-risk behaviors. PMID:17613970

  2. Thyroid Stimulating Hormone Receptor

    PubMed Central

    Tuncel, Murat

    2017-01-01

    Thyroid stimulating hormone receptor (TSHR) plays a pivotal role in thyroid hormone metabolism. It is a major controller of thyroid cell function and growth. Mutations in TSHR may lead to several thyroid diseases, most commonly hyperthyroidism. Although its genetic and epigenetic alterations do not directly lead to carcinogenesis, it has a crucial role in tumor growth, which is initiated by several oncogenes. This article will provide a brief review of TSHR and related diseases. PMID:28117293

  3. Thyroid Stimulating Hormone Receptor.

    PubMed

    Tuncel, Murat

    2016-01-05

    Thyroid stimulating hormone receptor (TSHR) plays a pivotal role in thyroid hormone metabolism. It is a major controller of thyroid cell function and growth. Mutations in TSHR may lead to several thyroid diseases, most commonly hyperthyroidism. Although its genetic and epigenetic alterations do not directly lead to carcinogenesis, it has a crucial role in tumor growth, which is initiated by several oncogenes. This article will provide a brief review of TSHR and related diseases.

  4. Novel high-affinity and selective biaromatic 4-substituted gamma-hydroxybutyric acid (GHB) analogues as GHB ligands: design, synthesis, and binding studies.

    PubMed

    Høg, Signe; Wellendorph, Petrine; Nielsen, Birgitte; Frydenvang, Karla; Dahl, Ivar F; Bräuner-Osborne, Hans; Brehm, Lotte; Frølund, Bente; Clausen, Rasmus P

    2008-12-25

    Gamma-hydroxybutyrate (GHB) is a metabolite of gamma-aminobutyric acid (GABA) and has been proposed to function as a neurotransmitter or neuromodulator. GHB is used in the treatment of narcolepsy and is a drug of abuse. GHB binds to both GABA(B) receptors and specific high-affinity GHB sites in brain, of which the latter have not been linked unequivocally to function, but are speculated to be GHB receptors. In this study, a series of biaromatic 4-substituted GHB analogues, including 4'-phenethylphenyl, 4'-styrylphenyl, and 4'-benzyloxyphenyl GHB analogues, were synthesized and characterized pharmacologically in a [3H](E,RS)-(6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene)acetic acid ([3H]NCS-382) binding assay and in GABA(A) and GABA(B) receptor binding assays. The compounds were selective for the high-affinity GHB binding sites and several displayed Ki values below 100 nM. The affinity of the 4-[4'-(2-iodobenzyloxy)phenyl] GHB analogue 17b was shown to reside predominantly with the R-enantiomer (Ki = 22 nM), which has higher affinity than previously reported GHB ligands.

  5. Fatality due to gamma-hydroxybutyric acid (GHB) and heroin intoxication.

    PubMed

    Ferrara, S D; Tedeschi, L; Frison, G; Rossi, A

    1995-05-01

    The first case of fatal intoxication due to ingestion of gamma-hydroxybutyric acid (GHB) and intravenous use of heroin is reported. A 42-year-old man, known to have been a heroin addict and to have taken other psychoactive substances, who had been in treatment with GHB for several months, was found dead. Anatomohistopathologic examination showed generalized visceral congestion, edema and pulmonary anthracosis, chronic bronchitis and chronic active hepatitis. Toxicological findings included fluid and tissue distributions of GHB, morphine and 6-monoacetylmorphine. GHB and morphine concentrations were respectively 11.5 and 0.77 micrograms/mL (blood), 84.3 and 0.3 micrograms/mL (vitreous humor), 258.3 and 1.35 micrograms/mL (urine), 57.0 and 14.3 micrograms/mL (bile), 40.0 and 0.43 micrograms/g (brain), 43.0 and 0.60 micrograms/g (liver), 47.0 and 0.68 micrograms/g (kidney). Blood and urine levels of 6-monoacetylmorphine were 28.5 and 12.1 ng/mL respectively. The presumed mechanism of action and pharmacokinetics of GHB are briefly reviewed, with reference to its therapeutic use and to reports of non-fatal GHB intoxication.

  6. Identification of a new metabolite of GHB: gamma-hydroxybutyric acid glucuronide.

    PubMed

    Petersen, Ida Nymann; Tortzen, Christian; Kristensen, Jesper Langgaard; Pedersen, Daniel Sejer; Breindahl, Torben

    2013-06-01

    Gamma-hydroxybutyric acid (GHB) is an important analyte in clinical and forensic toxicology with a narrow detection window of 3-6 h. In the search of improved detection methods, the existence in vivo of a glucuronated GHB metabolite (GHB-GLUC) was hypothesized. Chemically pure standards of GHB-GLUC and a deuterated analogue for chromatography were synthesized. Liquid chromatography and tandem mass spectrometry were used for targeted analysis in anonymous clinical urine samples (n = 50). GHB-GLUC was found in concentrations ranging from 0.11 to 5.0 µg/mL (mean: 1.3 ± 1.2 µg/mL). Thus far, this is the first report of a GHB glucuronide detected in biological samples. Given that glucuronides generally have longer half-life values than their corresponding free drugs, GHB-GLUC should theoretically be a biomarker of GHB intoxication. It is also proposed that the hitherto unexplained reports of elevated GHB concentrations in some biological samples, which has caused the setting of a relatively high cutoff value (10 µg/mL), represent total GHB measurements (sum of free GHB and actively chemically hydrolyzed GHB-GLUC). To address these challenges, the present study must be followed by comprehensive pharmacokinetic and stability studies after the controlled administration of GHB.

  7. Baclofen as relapse prevention in the treatment of gamma-hydroxybutyrate dependence: a case series.

    PubMed

    Kamal, Rama M; Loonen, Anton J M; Dijkstra, Boukje A G; De Jong, Cornelis A J

    2015-06-01

    In the last decade, gamma-hydroxybutyrate (GHB) abuse and dependence have increased. It has been reported that GHB dependence has a high rate of relapse, serious complications of intoxication, and a potentially life-threatening withdrawal syndrome. Nevertheless, in clinical practice, there is no known medical treatment to support GHB relapse prevention. We describe a case series of patients who were supported through an off-label treatment with baclofen to avoid a relapse into GHB abuse, for a period of 12 weeks. Nine of 11 patients did not relapse while taking a dose ranging from 30 to 60 mg per day, one patient relapsed after 5 weeks, and one stopped after 7 weeks. Baclofen was well tolerated; patients reported mild side effects such as fatigue, nausea, dry mouth, excessive sweating, and depressive feelings. Although systematic evidence is still lacking, our practice-based experience suggests that treatment with baclofen to assist abstinence might be effective in patients with GHB dependence. Further systematic controlled studies are necessary to establish the exact efficacy and safety of baclofen as relapse prevention for GHB-dependent patients.

  8. Gamma-hydroxybutyrate (GHB): a scoping review of pharmacology, toxicology, motives for use, and user groups.

    PubMed

    Brennan, Rebekah; Van Hout, Marie Claire

    2014-01-01

    Gamma hydroxybutyrate (GHB) is a central nervous system depressant with euphoric and relaxant effects. Documentation of GHB prevalence and the underreporting of abuse remains problematic, given the availability of GHB and its precursors γ-butyrolactone (GBL) and 1,4-butanediol (1,4-BD) and the ease of synthesis from kits available on the Internet. The continued abuse of and dependence on GHB, and associated fatalities, present an on-going public health problem. As the drug GHB remains an underresearched topic, a scoping review was chosen as a technique to map the available literature into a descriptive summarized account. PRISMA was used to assist in data retrieval, with subsequent data charting into three key themes (pharmacology and toxicology, outcomes, and user groups). Administered orally, GHB is dose-dependent and popular for certain uses (therapeutic, body enhancement, sexual assault) and amongst user sub groups (recreational party drug users, homosexual men). Despite the low prevalence of use in comparison to other club drugs, rising abuse of the drug is associated with dependence, withdrawal, acute toxicity, and fatal overdose. Clinical diagnosis and treatment is complicated by the co-ingestion of alcohol and other drugs. Limitations of the scoping review and potential for further research and harm reduction initiatives are discussed.

  9. Intravenous self-administration of gamma-hydroxybutyrate (GHB) in baboons

    PubMed Central

    Goodwin, Amy K.; Kaminski, Barbara J.; Griffiths, Roland R.; Ator, Nancy A.; Weerts, Elise M.

    2010-01-01

    Background Abuse of gamma-hydroxybutyrate (GHB) poses a public health concern. In previous studies, intravenous (IV) self-administration of GHB doses up to 10 mg/kg was not maintained in non-human primates under limited-access conditions, which was inconsistent with the usual good correspondence between drugs abused by humans and those self-injected by laboratory animals. Methods Self-administration of GHB was studied in 10 baboons using procedures standard for our laboratory to assess drug abuse liability. Each self-injection depended on completion of 120 or 160 lever responses. Sessions ran continuously; a 3-h timeout limited the number of injections per 24 h to 8. Self-injection was established at 6–8 injections/day with cocaine (0.32 mg/kg/injection) prior to substitution of each GHB dose (3.2–178 mg/kg/injection) or vehicle for 15 days. Food pellets were available 24 h/day. Results GHB maintained significantly greater numbers of injections when compared to vehicle in 6 of the 9 baboons that completed GHB evaluations that included 32 mg/kg/injection or higher. The baboons that self-administered GHB at high rates were ones for which GHB was the first drug each had tested under the 24-hr/day cocaine baseline procedure. Self-injection of the highest doses of GHB decreased food-maintained responding. Conclusions High-dose GHB can function as a reinforcer in non-human primates under 24-h access, but self-administration history may be important. The findings are consistent with the demonstrated abuse liability of GHB in humans, and remove GHB as an exception to the typical good correspondence between those drugs abused by humans and those self-administered by nonhuman primates. PMID:21112162

  10. Fatal Combination with 3-Methylmethcathinone (3-MMC) and Gamma-Hydroxybutyric Acid (GHB).

    PubMed

    Jamey, Carole; Kintz, Pascal; Martrille, Laurent; Raul, Jean-Sébastien

    2016-09-01

    We reported the case of 69-year-old man who was discovered dead at a friend's home. 3-Methylmethcathinone (3-MMC) and poppers (alkyl nitrites) were found at the scene by the police. Autopsy specimens including peripheral and cardiac blood, urine, gastric content, bile and hair were sent to our laboratory to document a possible death involving abuse of drugs. Routine toxicological analysis was performed with gas chromatography with flame ionization detection (GC-FID), high performance liquid chromatography-diode array detection (HPLC-DAD), headspace gas chromatography-mass spectrometry (HS-GC-MS), gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS)-MS. After liquid-liquid extraction at alkaline pH, 3-MMC was identified with GC-MS (to allow the discrimination with 4-MMC) and quantified with ultra-performance liquid chromatography-mass spectrometry (UPLC-MS)-MS with the two following transitions: m/z 178.1 > 160 and 178.1 > 144.9. Gamma-hydroxybutyric acid (GHB) was analyzed by GC-MS for fluids and GC-MS-MS for hair. Toxicological analysis in peripheral blood revealed the presence of 3-MMC (0.33 mg/L), pseudoephedrine (0.03 mg/L) and GHB (576 mg/L). These molecules have also been found in other post-mortem fluids. Furthermore, testing for "poppers" by HS-GC-MS was negative. Hair analysis, without segmentation, demonstrated the presence of 3-MMC (206.7 ng/mg), pseudoephedrine (0.16 ng/mg) and GHB (96.3 ng/mg) and suggested a repeated consumption of these substances. However, one cannot exclude contamination by sweat during the agony period. Toxicological post-mortem results suggest a fatal combination of 3-MMC and GHB. Despite his age, the decedent was known to abuse drugs.

  11. Twenty-three deaths with gamma-hydroxybutyrate overdose in western Sweden between 2000 and 2007.

    PubMed

    Knudsen, K; Jonsson, U; Abrahamsson, J

    2010-09-01

    gamma-Hydroxybutyrate (GHB) is a drug of abuse with a status as being safe. In spite of a reputation of low toxicity, a huge number of deaths associated with this drug have been recorded during recent years in Sweden. It is unclear whether coingestion with other drugs or ethanol causes death in GHB overdoses or whether GHB itself is the main cause of death. The aim of this study was to analyze the cause of death in GHB-related fatalities seen in our region. All cases of deaths with GHB during the year 2000-2007 in the region of western Sweden were studied retrospectively. The cases were classified as either GHB poisonings without any, with a minor or a major influence of other drugs on the cause of death. Twenty-three cases were diagnosed as deaths due to GHB overdose. Ninety-one percent coingested other substances. Ninety-one percent of the decedents were male. Age varied between 16 and 46, with the median age at 25 years. Forty-three percent of the cases were classified as GHB poisonings without any or a minor influence of other drugs on the cause of death. Thirty percent also ingested ethanol. Two patients (9%) were only intoxicated with GHB. Intoxication with GHB carries some mortality. Combining GHB with ethanol does not explain the many deaths in our region, nor do extremely high plasma concentrations of GHB. The intake of opioids increases the toxicity of GHB. The drug itself has such biological activities that an overdose is dangerous and may lead to death.

  12. Baclofen and gamma-hydroxybutyrate differentially altered behavior, EEG activity and sleep in rats.

    PubMed

    Hodor, A; Palchykova, S; Gao, B; Bassetti, C L

    2015-01-22

    Animal and human studies have shown that sleep may have an impact on functional recovery after brain damage. Baclofen (Bac) and gamma-hydroxybutyrate (GHB) have been shown to induce physiological sleep in humans, however, their effects in rodents are unclear. The aim of this study is to characterize sleep and electroencelphalogram (EEG) after Bac and GHB administration in rats. We hypothesized that both drugs would induce physiological sleep. Adult male Sprague-Dawley rats were implanted with EEG/electromyogram (EMG) electrodes for sleep recordings. Bac (10 or 20 mg/kg), GHB (150 or 300 mg/kg) or saline were injected 1 h after light and dark onset to evaluate time of day effect of the drugs. Vigilance states and EEG spectra were quantified. Bac and GHB induced a non-physiological state characterized by atypical behavior and an abnormal EEG pattern. After termination of this state, Bac was found to increase the duration of non-rapid eye movement (NREM) and rapid eye movement (REM) sleep (∼90 and 10 min, respectively), reduce sleep fragmentation and affect NREM sleep episode frequency and duration (p<0.05). GHB had no major effect on vigilance states. Bac drastically increased EEG power density in NREM sleep in the frequencies 1.5-6.5 and 9.5-21.5 Hz compared to saline (p<0.05), while GHB enhanced power in the 1-5-Hz frequency band and reduced it in the 7-9-Hz band. Slow-wave activity in NREM sleep was enhanced 1.5-3-fold during the first 1-2 h following termination of the non-physiological state. The magnitude of drug effects was stronger during the dark phase. While both Bac and GHB induced a non-physiological resting state, only Bac facilitated and consolidated sleep, and promoted EEG delta oscillations thereafter. Hence, Bac can be considered a sleep-promoting drug and its effects on functional recovery after stroke can be evaluated both in humans and rats. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  13. Relationship between gamma-hydroxybutyrate plasma concentrations and its electroencephalographic effects in the rat.

    PubMed

    Van Sassenbroeck, D K; De Paepe, P; Belpaire, F M; Rosseel, M T; Martens, P; Boon, P A; Buylaert, W A

    2001-12-01

    In view of the potential interest in an objective parameter for the depth of coma in intoxications with the recreational drug gamma-hydroxybutyrate (GHB), we have studied the relationship between the plasma concentrations and the electroencephalographic (EEG) changes induced by GHB in the rat. Fifteen rats randomly received either 150 (n = 3), 200 (n = 6) or 300 mg kg(-1) (n = 6) GHB over 5 min, followed by a supramaximal dose of 450 mg kg(-1) over 5 min at the end of the experiment. Plasma concentrations were determined with HPLC. The EEG was continuously recorded and the amplitude in the 15.5-30 Hz frequency band was quantified using aperiodic analysis. The plasma concentration-time profiles were fitted to a two-compartment model with Michaelis-Menten elimination. The pharmacokinetic parameters Vmax, Km and the apparent volume of distribution (Vd) proved to be independent of the dose and the mean pooled values were Vmax 2068 +/- 140 microg min(-1) kg(-1), Km 58 +/- 16 microg mL(-1) and Vd 476 +/- 12 mL kg(-1). The EEG amplitude in the 15.5-30 Hz frequency band displayed a monophasic inhibition and the effect-plasma concentration curve showed hysteresis. This hysteresis between EEG effect and plasma concentrations was minimized by simultaneous calculation of hypothetical effect-site concentrations and fitting the effect vs effect-site concentration curve to a sigmoid inhibitory Emax model. The descriptors of this Emax model (Emax, EC50, k(e,0), gamma and E0) were independent of the dose with an equilibration half-life t1/2k(e,0) of 5.6 +/- 0.3 min (mean value of the pooled results of the 5-min treatment groups). To investigate the origin of this hysteresis, a dose of 600 mg kg(-1) GHB was infused over either 45 or 60 min each in three animals. The hysteresis was much less pronounced with 45 min than with 5 min and was absent with 60-min infusions. This indicated that the hysteresis was due to a distribution delay between the central compartment and the effect site

  14. Illicit gamma-hydroxybutyrate (GHB) and pharmaceutical sodium oxybate (Xyrem®): differences in characteristics and misuse

    PubMed Central

    Carter, Lawrence P.; Pardi, Daniel; Gorsline, Jane; Griffiths, Roland R.

    2009-01-01

    There are distinct differences in the accessibility, purity, dosing, and misuse associated with illicit gamma-hydroxybutyrate (GHB) compared to pharmaceutical sodium oxybate. Gamma-hydroxybutyrate sodium and sodium oxybate are the chemical and drug names, respectively, for the pharmaceutical product Xyrem® (sodium oxybate) oral solution. However, the acronym GHB is also used to refer to illicit formulations that are used for non-medical purposes. This review highlights important differences between illicit GHB and sodium oxybate with regard to their relative abuse liability, which includes the likelihood and consequences of abuse. Data are summarized from the scientific literature; from national surveillance systems in the U.S., Europe, and Australia (for illicit GHB); and from clinical trials and post-marketing surveillance with sodium oxybate (Xyrem). In the U.S., the prevalence of illicit GHB use, abuse, intoxication, and overdose has declined from 2000, the year that GHB was scheduled, to the present and is lower than that of most other licit and illicit drugs. Abuse and misuse of the pharmaceutical product, sodium oxybate, has been rare over the 5 years since its introduction to the market, which is likely due in part to the risk management program associated with this product. Differences in the accessibility, purity, dosing, and misuse of illicit GHB and sodium oxybate suggest that risks associated with illicit GHB are greater than those associated with the pharmaceutical product sodium oxybate. PMID:19493637

  15. Sensorimotor impairment and elevated levels of dopamine metabolites in the neostriatum occur rapidly after intranigral injection of 6-hydroxydopamine or gamma-hydroxybutyrate in awake rats.

    PubMed

    Altar, C A; O'Neil, S; Marshall, J F

    1984-03-01

    The unilateral injection of 6-hydroxydopamine (8 micrograms) into the ventral tegmental area of awake rats produced a rapidly developing and irreversible sensory neglect to contralateral tactile stimuli. This neglect developed in a caudal to rostral direction on the affected body surface and coincided with significant elevation in the concentrations of dopamine and two of its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the ipsilateral neostriatum. The unilateral injection of procaine or gamma-hydroxybutyric acid (GHB) into the substantia nigra of awake animals also produced a contralateral neglect that developed in a caudal to rostral direction, but the behavioral effect of these drugs diminished within 1 hr. Concentrations of dopamine, dihydroxyphenylacetic acid and homovanillic acid in the neostriatum were markedly elevated during continuous infusions of procaine or gamma-hydroxybutyric acid. The extent of sensory neglect and changes in dopamine metabolism in the neostriatum varied according to the amount of gamma-hydroxybutyric acid injected into the nigra and according to the proximity of injections of gamma-hydroxybutyric acid to the pars compacta. The rapid onset of sensory neglect following microinjections of 6-hydroxydopamine, procaine or gamma-hydroxybutyric acid is consistent with the ability of each of these drugs to block the conduction of impulses in mesostriatal neurons and suggests that concomitant increases in levels of dopamine, dihydroxyphenylacetic acid and homovanillic acid in the neostriatum resulted from decreases in the release of dopamine coupled with increased synthesis of dopamine. These findings also indicate that the catabolism of dopamine to dihydroxyphenylacetic acid or homovanillic acid may originate intraneuronally, without prior release of dopamine and its recapture by mesostriatal terminals, if the flow of impulses in this pathway has been blocked.

  16. Gamma-Hydroxybutyrate (GHB) Content in Hair Samples Correlates Negatively with Age in Succinic Semialdehyde Dehydrogenase Deficiency.

    PubMed

    Johansen, S S; Wang, X; Sejer Pedersen, D; Pearl, P L; Roullet, J-B; Ainslie, G R; Vogel, K R; Gibson, K M

    2017-02-18

    Gamma-hydroxybutyrate (GHB) is a drug of abuse, an approved therapeutic for narcolepsy, an agent employed for facilitation of sexual assault, as well as a biomarker of succinic semialdehyde dehydrogenase deficiency (SSADHD). Our laboratory seeks to identify surrogate biomarkers in SSADHD that can shed light on the developmental course of this neurometabolic disease. Since GHB may be quantified in hair as a potential surrogate to identify victims of drug-related assault, we have opted to examine its level in SSADHD. We quantified GHB in hair derived from ten patients with SSADHD, and documented a significant negative age correlation. These findings are consistent with recent results in patient biological fluids, including plasma and red blood cells. These findings may provide additional insight into the developmental course of SSADHD (Jansen et al., J Inherit Metab Dis 39:795-800, 2016).

  17. Sedative and hypothermic effects of gamma-hydroxybutyrate (GHB) in rats alone and in combination with other drugs: assessment using biotelemetry.

    PubMed

    van Nieuwenhuijzen, Petra S; McGregor, Iain S

    2009-08-01

    The recreational drug gamma-hydroxybutyrate (GHB) has euphoric effects and can induce sedation and body temperature changes. GHB is frequently combined with other recreational drugs although these interactions are not well characterised. The present study used biotelemetry to provide a fine-grained analysis of the effects of GHB on body temperature and locomotor activity in freely moving rats, and investigated interactions between GHB and 3,4-methylenedioxymethamphetamine (MDMA), methamphetamine (METH) and various antagonist drugs. GHB (1000mg/kg) caused profound sedation for more than 2h and a complex triphasic effect on body temperature: an initial hypothermia (5-40min), followed by hyperthermia (40-140min), followed again by hypothermia (140-360min). A lower GHB dose (500mg/kg) also caused sedation but only a hypothermic effect that lasted up to 6h. The dopamine D(1) receptor antagonist SCH 23390 (1mg/kg), the opioid antagonist naltrexone (1mg/kg), the benzodiazepine antagonist flumazenil (10mg/kg), and the 5-HT(2A/2C) receptor antagonist ritanserin (1mg/kg) did not prevent the overall sedative or body temperature effects of GHB (1000mg/kg). However the GABA(B) antagonist SCH 50911 (50mg/kg) prevented the hyperthermia induced by GHB (1000mg/kg). Repeated daily administration of GHB (1000mg/kg) produced tolerance to the sedative and hyperthermic effects of the drug and cross-tolerance to the sedative effects of the GABA(B) receptor agonist baclofen (10mg/kg). A high ambient temperature of 28 degrees C prevented the hypothermia obtained with GHB (500mg/kg) at 20 degrees C, while GHB (500mg/kg) reduced the hyperthermia and hyperactivity produced by co-administered doses of MDMA (5mg/kg) or METH (1mg/kg) at 28 degrees C. These results further confirm a role for GABA(B) receptors in the hypothermic and sedative effects of GHB and show an interaction between GHB and MDMA, and GHB and METH, that may be relevant to the experience of recreational users who mix these

  18. Gamma-hydroxybutyrate concentrations in the blood of impaired drivers, users of illicit drugs, and medical examiner cases.

    PubMed

    Jones, A Wayne; Holmgren, Anita; Kugelberg, Fredrik C

    2007-01-01

    Gamma-hydroxybutyrate (GHB) was determined in blood samples from impaired drivers, people arrested for petty drug offenses (non-traffic cases), and GHB-related deaths. The method of analysis involved conversion of GHB into gamma-butyrolactone and determination of the latter by gas chromatography with a flame ionization detector, and with gamma-valerolactone as the internal standard. The mean and median concentrations of GHB in blood from impaired drivers (N=473) were 90 and 84 mg/L, respectively, and offenders were predominantly men (96%) with an average age of 26 year (range 15-50 year). In 185 cases, GHB was the only drug present in blood at mean and median concentrations of 92 and 86 mg/L, respectively. People arrested for petty drug offenses (N=1061) had slightly higher GHB concentrations in their blood: median 118 mg/L for men and 111 mg/L for women. In GHB-related deaths (N=33), the mean and median concentrations were considerably higher: 307 mg/L and 190 mg/L, respectively, and the highest was 2200 mg/L. The typical signs of drug influence noted by the arresting police officers included sedation, agitation, slurred speech, irrational behaviour, jerky movements, and spitting. The short elimination half-life of GHB means that the concentrations in blood decrease rapidly and are probably a lot lower than at the time of driving, which was 30-90 min earlier.

  19. Estimation of gamma-hydroxybutyrate (GHB) co-consumption in serum samples of drivers positive for amphetamine or ecstasy.

    PubMed

    Lott, S; Musshoff, F; Madea, B

    2012-09-10

    There is no toxicological analysis of gamma-hydroxybutyrate (GHB) applied routinely in cases of driving under influence (DUI); therefore the extent of consumption of this drug might be underestimated. Its consumption is described as occurring often concurrently with amphetamine or ecstasy. This study examines 196 serum samples which were collected by police during road side testing for GHB. The samples subject to this study have already been found to be positive for amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA) and/or 3,4-methylenedioxyethamphetamine (MDEA). Analysis has been performed by LC/MS/MS in the multiple reaction monitoring (MRM) mode. Due to its polarity, chromatographic separation of GHB was achieved by a HILIC column. To differentiate endogenous and exogenous levels of GHB, a cut-off concentration of 4μg/ml was applied. Of the 196 samples, two have been found to be positive for GHB. Of these samples, one sample was also positive for amphetamine and one for MDMA. Whilst other amphetamine derivates were not detected in these samples, both samples were found to be positive for cannabinoids. These results suggest that co-consumption of GHB with amphetamine or ecstasy is relatively low (1%) for the collective of this study.

  20. The presence of gamma-hydroxybutyric acid (GHB) and gamma-butyrolactone (GBL) in alcoholic and non-alcoholic beverages.

    PubMed

    Elliott, Simon; Burgess, Victoria

    2005-07-16

    Gamma-hydroxybutyric acid (GHB) and its precursor gamma-butyrolactone (GBL) are regularly implicated in instances of surreptitious drug administration, particularly in beverages (so-called "spiked drinks"). In order to assist in the interpretation of cases where analysis of the actual beverage is required, over 50 beverages purchased in the UK were analysed for the presence of GHB and GBL. It was found that naturally occurring GHB and GBL were detected in those beverages involving the fermentation of white and particularly red grapes. No GHB or GBL was detected in other drinks such as beer, juice, spirits or liqueurs. GHB/GBL was detected in red wine vermouth (8.2 mg/L), sherry (9.7 mg/L), port (GBL), red wine (4.1-21.4 mg/L) and white wine (<3-9.6 mg/L). The presence of GHB/GBL did not appear to be influenced by the alcohol content or the pH of the beverage. In addition, the concentration in wines did not appear to be related to the geographical origin of the grape type. This is believed to be the first published data concerning the endogenous presence of GHB and GBL in the beverages described.

  1. Monitoring of the interconversion of gamma-butyrolactone (GBL) to gamma hydroxybutyric acid (GHB) by Raman spectroscopy.

    PubMed

    Munshi, Tasnim; Brewster, Victoria L; Edwards, Howell G M; Hargreaves, Michael D; Jilani, Shelina K; Scowen, Ian J

    2013-08-01

    Gamma-hydroxybutyric acid (GHB) is a drug-of-abuse that has recently become associated with drug-facilitated sexual assault, known as date rape. For this reason the drug is commonly found 'spiked' in alcoholic beverages. When GHB is in solution it may undergo conversion into the corresponding lactone, Gamma-butyrolactone (GBL). Studies have been carried out to determine the detection limits of GHB and GBL in various solutions by Raman spectroscopy and to monitor the interconversion of GHB and GBL in solution with different pH conditions and temperature. In this study, a portable Raman spectrometer was used to study the interconversion of GHB and GBL in water and ethanol solutions as a function of pH, time, and temperature. The aim of this was to determine the optimum pH range for conversion in order to relate this to the pH ranges that the drug is likely to be subjected to, first in spiked beverages and secondly after ingestion in the digestive system. The aim was also to identify a timescale for this conversion in relation to possible scenarios, for example if GHB takes a number of hours to convert to GBL, it is likely for the beverage to be ingested before esterification can take place. GHB and GBL were then spiked into a selection of beverages of known pH in order to study the stability of GHB and GBL in real systems. Copyright © 2012 John Wiley & Sons, Ltd.

  2. The Impact of Gamma Hydroxybutyrate (GHB) Legal Restrictions on Patterns of Use: Results from an International Survey

    PubMed Central

    Anderson, IB; Kim-Katz, SY; Dyer, JE; Blanc, PD

    2009-01-01

    Aims To conduct an Internet-based survey of GHB use, identifying differences by respondent residence. Methods We recruited GHB-knowledgeable persons via “social networking Internet sites.” Individuals (n=314) or groups (n=66) were approached based on GHB-use testimonials. Data collected location, use, reason for cessation (if applicable). Findings We recruited 155 GHB users. U.S. respondents (53 of 70; 76%) compared to non-U.S. respondents (38 of 85; 45%) were older and more highly educated (p<0.05) but manifest a 3-fold greater adjusted odds of GHB cessation (Odds Ratio [OR] 3.1; 95% CI 1.4–6.9; p < 0.05). Of the 80 respondents stating reason for cessation, 36 (45%) cited legal risk, price, or access; 44 (55%) cited health or related concerns. U.S. compared to non-U.S. respondents more frequently invoked legal and related concerns (OR 2.5; 95% CI 0.99–6.3; p=0.05). In a nested analysis, narrowly stated legal (n=4/5 U.S.) versus health (n=6/18 U.S.) reasons differed by location (p=0.048, one-tailed). Conclusions In the U.S., where GHB has stricter legal penalties, GHB cessation is more likely, with legal and related reasons more commonly invoked for cessation. These findings support a link between declining U.S. GHB abuse and more stringent restrictions; although other un-assessed factors may also explain this association. The Impact of Gamma Hydroxybutyrate (GHB) Legal Restrictions on Patterns of Use: Results from an International Survey PMID:20953310

  3. Catabolism of GABA, succinic semialdehyde or gamma-hydroxybutyrate through the GABA shunt impair mitochondrial substrate-level phosphorylation.

    PubMed

    Ravasz, Dora; Kacso, Gergely; Fodor, Viktoria; Horvath, Kata; Adam-Vizi, Vera; Chinopoulos, Christos

    2017-03-11

    GABA is catabolized in the mitochondrial matrix through the GABA shunt, encompassing transamination to succinic semialdehyde followed by oxidation to succinate by the concerted actions of GABA transaminase (GABA-T) and succinic semialdehyde dehydrogenase (SSADH), respectively. Gamma-hydroxybutyrate (GHB) is a neurotransmitter and a psychoactive drug that could enter the citric acid cycle through transhydrogenation with α-ketoglutarate to succinic semialdehyde and d-hydroxyglutarate, a reaction catalyzed by hydroxyacid-oxoacid transhydrogenase (HOT). Here, we tested the hypothesis that the elevation in matrix succinate concentration caused by exogenous addition of GABA, succinic semialdehyde or GHB shifts the equilibrium of the reversible reaction catalyzed by succinate-CoA ligase towards ATP (or GTP) hydrolysis, effectively negating substrate-level phosphorylation (SLP). Mitochondrial SLP was addressed by interrogating the directionality of the adenine nucleotide translocase during anoxia in isolated mouse brain and liver mitochondria. GABA eliminated SLP, and this was rescued by the GABA-T inhibitors vigabatrin and aminooxyacetic acid. Succinic semialdehyde was an extremely efficient substrate energizing mitochondria during normoxia but mimicked GABA in abolishing SLP in anoxia, in a manner refractory to vigabatrin and aminooxyacetic acid. GHB could moderately energize liver but not brain mitochondria consistent with the scarcity of HOT expression in the latter. In line with these results, GHB abolished SLP in liver but not brain mitochondria during anoxia and this was unaffected by either vigabatrin or aminooxyacetic acid. It is concluded that when mitochondria catabolize GABA or succinic semialdehyde or GHB through the GABA shunt, their ability to perform SLP is impaired.

  4. The Impact of Gamma Hydroxybutyrate (GHB) Legal Restrictions on Patterns of Use: Results from an International Survey.

    PubMed

    Anderson, Ib; Kim-Katz, Sy; Dyer, Je; Blanc, Pd

    2010-10-01

    AIMS: To conduct an Internet-based survey of GHB use, identifying differences by respondent residence. METHODS: We recruited GHB-knowledgeable persons via "social networking Internet sites." Individuals (n=314) or groups (n=66) were approached based on GHB-use testimonials. DATA COLLECTED: location, use, reason for cessation (if applicable). FINDINGS: We recruited 155 GHB users. U.S. respondents (53 of 70; 76%) compared to non-U.S. respondents (38 of 85; 45%) were older and more highly educated (p<0.05) but manifest a 3-fold greater adjusted odds of GHB cessation (Odds Ratio [OR] 3.1; 95% CI 1.4-6.9; p < 0.05). Of the 80 respondents stating reason for cessation, 36 (45%) cited legal risk, price, or access; 44 (55%) cited health or related concerns. U.S. compared to non-U.S. respondents more frequently invoked legal and related concerns (OR 2.5; 95% CI 0.99-6.3; p=0.05). In a nested analysis, narrowly stated legal (n=4/5 U.S.) versus health (n=6/18 U.S.) reasons differed by location (p=0.048, one-tailed). CONCLUSIONS: In the U.S., where GHB has stricter legal penalties, GHB cessation is more likely, with legal and related reasons more commonly invoked for cessation. These findings support a link between declining U.S. GHB abuse and more stringent restrictions; although other un-assessed factors may also explain this association. The Impact of Gamma Hydroxybutyrate (GHB) Legal Restrictions on Patterns of Use: Results from an International Survey.

  5. A critical evaluation of the gamma-hydroxybutyrate (GHB) model of absence seizures.

    PubMed

    Venzi, Marcello; Di Giovanni, Giuseppe; Crunelli, Vincenzo

    2015-02-01

    Typical absence seizures (ASs) are nonconvulsive epileptic events which are commonly observed in pediatric and juvenile epilepsies and may be present in adults suffering from other idiopathic generalized epilepsies. Our understanding of the pathophysiological mechanisms of ASs has been greatly advanced by the availability of genetic and pharmacological models, in particular the γ-hydroxybutyrate (GHB) model which, in recent years, has been extensively used in studies in transgenic mice. GHB is an endogenous brain molecule that upon administration to various species, including humans, induces not only ASs but also a state of sedation/hypnosis. Analysis of the available data clearly indicates that only in the rat does there exist a set of GHB-elicited behavioral and EEG events that can be confidently classified as ASs. Other GHB activities, particularly in mice, appear to be mostly of a sedative/hypnotic nature: thus, their relevance to ASs requires further investigation. At the molecular level, GHB acts as a weak GABA-B agonist, while the existence of a GHB receptor remains elusive. The pre- and postsynaptic actions underlying GHB-elicited ASs have been thoroughly elucidated in thalamus, but little is known about the cellular/network effects of GHB in neocortex, the other brain region involved in the generation of ASs.

  6. Modification of ethanol's reinforcing effectiveness in rhesus monkeys by cocaine, flunitrazepam, or gamma-hydroxybutyrate.

    PubMed

    Winger, Gail; Galuska, Chad M; Hursh, Steven R

    2007-09-01

    Although ethanol is frequently used in combination with other psychoactive drugs, the behavioral and pharmacological reasons for this form of polydrug abuse have not been well described. Rhesus monkeys with indwelling intravenous catheters produced intravenous injections of ethanol (50, 100, or 200 mg/kg/inj), flunitrazepam (0.001-0.03 mg/kg/inj), cocaine (0.01 or 0.03 mg/kg/inj), or combinations of ethanol and these drugs or gammahydroxybutyrate (GHB) (1.0 or 3.2 mg/kg/inj) by lever pressing according to a fixed-ratio schedule. The response requirement for each drug or drug combination was increased across sessions (10, 32, 100, 320, or 1,000). The dependent variables were rates of responding maintained by the drug or drug combination and the elasticity of drug demand when consumption was expressed as a function of price. Elasticity (P (max)) values for each drug varied among the monkeys but retained the same rank order for the monkeys, suggesting a fundamental difference in the animals' apparent sensitivities to the reinforcing effects of the drugs. Combining ethanol with the other drugs did not increase their reinforcing effectiveness. GHB (ineffective in previous studies) did not modify ethanol's reinforcing effects; demand functions for the combination of ethanol and flunitrazepam were slightly less elastic than for ethanol alone, but no different from that for flunitrazepam alone; adding ethanol to cocaine detracted from the reinforcing effectiveness of cocaine. The hypothesis that use of ethanol in combination with sedative and stimulant drugs is due to an ability of ethanol to enhance the reinforcing effects of these drugs is not supported.

  7. Postmortem concentrations of gamma-hydroxybutyrate (GHB) in peripheral blood and brain tissue - Differentiating between postmortem formation and antemortem intake.

    PubMed

    Thomsen, Ragnar; Rasmussen, Brian Schou; Johansen, Sys Stybe; Linnet, Kristian

    2017-03-01

    Gamma-hydroxybutyrate (GHB) is a recreational drug, a drug of abuse, as well as an endogenous molecule in mammals. The drug has become infamous as a tool for drug-facilitated sexual assault. GHB is found in low concentrations in living humans, while at postmortem the concentration of GHB rises due to fermentation processes. The endogenous nature of GHB leads to difficulty in interpretation of concentrations, as the source of GHB is not obvious. Postmortem brain and blood samples were collected from 221 individuals at autopsy. Of these, 218 were not suspected of having ingested GHB, while GHB intake was reported for the last three (cases A-C). Decomposition level was estimated and cases classified into no/minor and advanced decomposition. Brain samples were extracted from the frontal lobe; only gray matter from the cerebral cortex was used. Blood was drawn from the femoral vein. Brain samples were homogenized and diluted with water. Brain homogenates or femoral blood were then prepared using protein precipitation and GHB was quantified with UHPLC-MS/MS. For 189 cases where ingestion of GHB was not suspected and where no/minor decomposition had occurred the concentrations were in the range 4.8-45.4mg/kg (median 15.3mg/kg) in blood and not-detected to 9.8mg/kg (median 4.8mg/kg) in brain tissue. For case A, where intoxication with GHB was deemed to be the sole cause of death, the concentrations were 199 and 166mg/kg in blood and brain, respectively. For case B, where intoxication with GHB was a contributing factor of death, the respective concentrations were 142 and 78.4mg/kg. For case C, where GHB was ingested but the cause of death was opioid poisoning, the concentrations were 40.3 and 12.7mg/kg. The results demonstrate that postmortem-formed levels of GHB are much lower in brain than peripheral blood. Analysis of GHB in brain tissue thus provides for an improved capability to identify an exogenous source of GHB. By measuring GHB in brain tissue and employing a cut

  8. Endogenous gamma-hydroxybutyric acid (GHB) concentrations in post-mortem specimens and further recommendation for interpretative cut-offs.

    PubMed

    Andresen-Streichert, Hilke; Jensen, P; Kietzerow, J; Schrot, M; Wilke, N; Vettorazzi, E; Mueller, A; Iwersen-Bergmann, S

    2015-01-01

    When interpreting gamma-hydroxybutyric acid (GHB) concentrations in post-mortem specimens, a possible increase in GHB concentrations because of post-mortem generation must be considered. In this study, endogenous GHB concentrations in post-mortem biological fluids were investigated. Additionally, we review post-mortem GHB concentrations already published in the literature. Heart and peripheral blood samples, cerebrospinal fluid, urine, and vitreous humor were collected from 64 autopsies in subjects where the cause of death excluded GHB exposure. Sample analysis was carried out either on the day of autopsy or later after immediate freezing and storage at -20 °C. GHB concentrations in venous blood samples (n = 61) were <0.6-28.7 mg/L (mean 11.9 mg/L; median 10.6 mg/L), <0.6-65.3 mg/L (mean 15.2 mg/L; median 12.8 mg/L) in heart blood (n = 56), <0.6-25.1 mg/L (mean 6.0 mg/L; median 3.8 mg/L) in urine (n = 50), <0.6-39.0 mg/L (mean 9.6 mg/L; median 7.5 mg/L), in vitreous humor (n = 54), and <0.6-24.0 mg/L (mean 4.2 mg/L; median 3.2 mg/L) in cerebrospinal fluid (n = 52). There was no significant difference between GHB concentrations in cases where there were signs of beginning putrefaction at the time of autopsy (n = 9) and cases without obvious signs of putrefaction. In one case with advanced putrefaction, the GHB concentration in venous blood was 32.7 mg/L. In conclusion, for post-mortem venous blood, urine, and cerebrospinal fluid, an interpretative cut-off of 30 mg/L for GHB concentrations is suggested in cases where GHB analysis is conducted on the day of sample collection at autopsy or if samples have been stored at -20 °C immediately after collection.

  9. Pharmacokinetics of 1,4-butanediol in rats: bioactivation to gamma-hydroxybutyric acid, interaction with ethanol, and oral bioavailability.

    PubMed

    Fung, Ho-Leung; Tsou, Pei-Suen; Bulitta, Jurgen B; Tran, Doanh C; Page, Nathaniel A; Soda, David; Mi Fung, Sun

    2008-01-01

    1,4-Butanediol (BD), a substance of abuse, is bioactivated to gamma-hydroxybutyrate (GHB), but its fundamental pharmacokinetics (PK) have not been characterized. Because this bioactivation is partly mediated by alcohol dehydrogenase, we hypothesized that there may also be a metabolic interaction between ethanol (ETOH) and BD. We therefore studied, in rats, the plasma PK of GHB, BD and ETOH each at two intravenous (IV) doses, when each substance was given alone, and when GHB or BD was co-administered with ETOH. Results showed that bioconversion of intravenously administered BD to GHB was complete, and that both GHB and BD exhibited nonlinear PK. Various population PK models were analyzed using NONMEM VI, and the best disposition model was found to include two PK compartments each for BD, an (unmeasured) putative semialdehyde intermediate (ALD), GHB and ETOH, the presence of nonlinear (Michaelis-Menten) elimination for each compound, and several mutual inhibition processes. The most prominent mutual metabolic inhibition was found between ETOH and BD, while that between GHB and ETOH was not significant. In vitro studies using liver homogenates confirmed mutual metabolic inhibitions between GHB and BD. Oral absorption of BD was best described by a first-order process with lag-time and pre-systemic metabolism from BD to ALD. Oral absorption of BD (as BD plus ALD) was rapid and complete. The fraction of the absorbed dose entering the central compartment as BD was 30% for the 1.58 mmol/kg dose and 55% for the 6.34 mmol/kg dose. At 6.34 mmol/kg IV, the onset of loss of righting reflex (LRR) for BD was significantly delayed vs. that produced by GHB (72.0 +/- 9.1 min vs. 6.7 +/- 0.6 min, respectively, p < 0.001), and the total duration of LRR was prolonged for BD vs. GHB (192 +/- 28 min vs. 117 +/- 2 min, respectively, p < 0.05). Relative to IV dosing, oral BD produced similar but more variable LRR effects. These results may provide a quantitative PK framework for the

  10. Report on the analysis of common beverages spiked with gamma-hydroxybutyric acid (GHB) and gamma-butyrolactone (GBL) using NMR and the PURGE solvent-suppression technique.

    PubMed

    Lesar, Casey T; Decatur, John; Lukasiewicz, Elaan; Champeil, Elise

    2011-10-10

    In forensic evidence, the identification and quantitation of gamma-hydroxybutyric acid (GHB) in "spiked" beverages is challenging. In this report, we present the analysis of common alcoholic beverages found in clubs and bars spiked with gamma-hydroxybutyric acid (GHB) and gamma-butyrolactone (GBL). Our analysis of the spiked beverages consisted of using (1)H NMR with a water suppression method called Presaturation Utilizing Relaxation Gradients and Echoes (PURGE). The following beverages were analyzed: water, 10% ethanol in water, vodka-cranberry juice, rum and coke, gin and tonic, whisky and diet coke, white wine, red wine, and beer. The PURGE method allowed for the direct identification and quantitation of both compounds in all beverages except red and white wine where small interferences prevented accurate quantitation. The NMR method presented in this paper utilizes PURGE water suppression. Thanks to the use of a capillary internal standard, the method is fast, non-destructive, sensitive and requires no sample preparation which could disrupt the equilibrium between GHB and GBL.

  11. Quantitation of Gamma-Hydroxybutyric Acid in Dried Blood Spots: Feasibility Assessment for Newborn Screening of Succinic Semialdehyde Dehydrogenase (SSADH) Deficiency

    PubMed Central

    Forni, Sabrina; Pearl, Phillip L.; Gibson, K. Michael; Yu, Yuezhou; Sweetman, Lawrence

    2013-01-01

    Objective SSADH deficiency, the most prevalent autosomal recessive disorder of GABA degradation, is characterized by elevated gamma-hydroxybutyric acid (GHB). Neurological outcomes may be improved with early intervention and anticipatory guidance. Morbidity has been compounded by complications, e.g. hypotonia, in undiagnosed infants with otherwise routine childhood illnesses. We report pilot methodology on the feasibility of newborn screening for SSADH deficiency. Method Dried blood spot (DBS) cards from patients affected with SSADH deficiency were compared with 2831 archival DBS cards for gamma-hydroxybutyric acid content. Following extraction with methanol, GHB in DBS was separated and analyzed using ultra high-performance liquid chromatography tandem mass spectrometry. Results Methodology was validated to meet satisfactory accuracy and reproducibility criteria, including intra-day and inter-day validation. Archival refrigerated dried blood spots samples of babies, infants and children (N=2831) were screened for GHB, yielding a mean +/- S.D. of 8 ± 5 nM (99.9 %-tile 63 nM) (Min 0.0 Max 78 nM). The measured mean and median concentrations in blood spots derived from seven SSADH deficient patients were 1182 nM and 699 nM respectively (Min 124, Max 4851nM). Conclusions GHB concentration in all 2831 dried blood spot cards was well below the lowest concentration of affected children. These data provide proof-of-principle for screening methodology to detect SSADH deficiency with applicability to newborn screening and earlier diagnosis. PMID:23742746

  12. Gamma-Hydroxybutyrate (GHB)

    MedlinePlus

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  13. Post mortem concentrations of endogenous gamma hydroxybutyric acid (GHB) and in vitro formation in stored blood and urine samples.

    PubMed

    Busardò, Francesco Paolo; Bertol, Elisabetta; Vaiano, Fabio; Baglio, Giovanni; Montana, Angelo; Barbera, Nunziata; Zaami, Simona; Romano, Guido

    2014-10-01

    Gamma-hydroxybutyrate (GHB) is a central nervous system depressant, primarily used as a recreational drug of abuse with numerous names. It has also been involved in various instances of drug-facilitated sexual assault due to its potential incapacitating effects. The first aim of this paper is to measure the post-mortem concentration of endogenous GHB in whole blood and urine samples of 30 GHB free-users, who have been divided according to the post-mortem interval (PMI) in three groups (first group: 24-36h; second group: 37-72h; third group: 73-192h), trying to evaluate the role of PMI in affecting post mortem levels. Second, the Authors have evaluated the new formation of GHB in vitro in blood and urine samples of the three groups, which have been stored at -20°C, 4°C and 20°C over a period of one month. The concentrations were measured by GC-MS after liquid-liquid extraction according to the method validated and published by Elliot (For. Sci. Int., 2003). For urine samples, GHB concentrations were creatinine-normalized. In the first group the GHB mean concentration measured after autopsy was: 2.14mg/L (range 0.54-3.21mg/L) in blood and 3.90mg/g (range 0.60-4.81mg/g) in urine; in the second group it was: 5.13mg/L (range 1.11-9.60mg/L) in blood and 3.93mg/g (range 0.91-7.25mg/g) in urine; in the third group it was: 11.8mg/L (range 3.95-24.12mg/L) in blood and 9.83mg/g (range 3.67-21.90mg/g) in urine. The results obtained in blood and urine samples showed a statistically significant difference among groups (p<0.001) in the first analysis performed immediately after autopsy. Throughout the period of investigation up to 4 weeks, the comparison of storage temperatures within each group showed in blood and urine samples a mean difference at 20°C compared to -20°C not statistically significant at the 10% level. These findings allow us to affirm that the PMI strongly affects the post mortem production of GHB in blood and urine samples. Regarding the new formation of

  14. Analysis of gamma-hydroxybutyric acid, DL-lactic acid, glycolic acid, ethylene glycol and other glycols in body fluids by a direct injection gas chromatography-mass spectrometry assay for wide use.

    PubMed

    Van Hee, Paul; Neels, Hugo; De Doncker, Mireille; Vrydags, Nicolas; Schatteman, Katinka; Uyttenbroeck, Wim; Hamers, Nicole; Himpe, Dirk; Lambert, Willy

    2004-01-01

    Analysis of blood of severely intoxicated patients always requires prompt investigation. Diagnosis of intoxication with ethylene glycol, gamma-hydroxybutyric acid or D-lactic acid takes hours, since several different procedures are required. Rapid derivatization of the common hydroxyl function may resolve this analytical problem. Here we describe a fast method for the simultaneous measurement of ethylene glycol, glycolic acid, gamma-hydroxybutyric acid and racemic lactic acid. Only 20 microl of serum, plasma or urine are required for immediate derivatization at 70 degrees C with 750 microl of bis-N,O-trimethylsilyl trifluoroacetamide after adding 20 microl of internal standard solution (1,3-propylene glycol) and 20 microl of the catalyst dimethylformamide. After centrifugation an aliquot is transferred to a gas chromatographic system and analyzed with electron-impact mass spectrometry in selective ion monitoring mode. The derivatized acids and ethylene glycol are well separated and detected with a limit of detection ranging from 0.12 mg/l for ethylene glycol to 0.95 mg/l for gamma-hydroxybutyric acid, while the limit of quantification ranged from 0.4 mg/l for ethylene glycol to 3.15 mg/l for gamma-hydroxybutyric acid. The method is linear from 0.5 to 1800 mg/l blood for ethylene glycol, from 0.7 to 1200 mg/l for lactic acid, from 1.2 to 1800 mg/l for glycolic acid, and from 3.2 to 200 mg/l for gamma-hydroxybutyric acid, with analytical recoveries, accuracy, day-to-day and within-day precision well within the required limits. Total analysis time with one calibrator was 30 min, derivatization time included. This method is very suitable for emergency toxicology, since several toxic substances can be quantified simultaneously in a fast and sensitive manner.

  15. Chemical composition and structure of the microcrystals formed between silver(I) and gamma-hydroxybutyric acid and gamma-hydroxyvaleric acid.

    PubMed

    Bell, Suzanne C; Oldfield, Lucy S; Shakleya, Diaa M; Petersen, Jeffrey L; Mercer, Jennifer W

    2006-07-01

    This study examined microcrystals formed by silver with gamma-hydroxybutyric acid (GHB) and gamma-hydroxyvaleric acid (GHV), the five-carbon analog of GHB, in the presence of silver, copper, and lanthanide nitrates. Distinct microcrystals formed with silver (+1) and lanthanum (+3) ions but not with the copper (+2) ions. The crystals formed with GHB were distinctly different than those formed with GHV and in all cases, the drug microcrystals were easily distinguishable from reagent crystals. X-ray diffraction analysis provided definitive structure for the microcrystals. The morphological differences between the silver-GHB and silver-GHV crystals were characterized using simple measurements such as size and angles provided by image recognition software. The utility of the test for casework was demonstrated using spiked beverage samples.

  16. Interaction of beta-adrenergic agonists and antagonists with the stimulation of growth hormone release induced by clonidine or by morphine in the rat.

    PubMed

    Bluet-Pajot, M T; Durand, D; Mounier, F; Schaub, C; Kordon, C

    1982-09-01

    The beta-adrenergic agonist, isoprenaline, and antagonist, propranolol, had no effect on the delayed basal secretion of GH consistently observed in rats treated with the narco-analgesic gamma-hydroxybutyrate. Under the same experimental conditions, GH release was distinctly stimulated by infusion of the alpha-adrenergic agonist, clonidine, and by morphine; both responses were dose-dependent. The effects of beta-adrenergic agonists and antagonists on these GH responses were as follows: in rats pretreated with isoprenaline -the GH release induced by clonidine and morphine was abolished whereas it was enhanced in rats pretreated with propranolol. These data confirmed and extended previous reports from this laboratory on the inhibitory role of beta-adrenergic receptors on GH regulation.

  17. Carbon isotopic ratio analysis by gas chromatography/combustion/isotope ratio mass spectrometry for the detection of gamma-hydroxybutyric acid (GHB) administration to humans.

    PubMed

    Saudan, Christophe; Augsburger, Marc; Mangin, Patrice; Saugy, Martial

    2007-01-01

    Since GHB (gamma-hydroxybutyric acid) is naturally produced in the human body, clinical and forensic toxicologists must be able to discriminate between endogenous levels and a concentration resulting from exposure. To suggest an alternative to the use of interpretative concentration cut-offs, the detection of exogenous GHB in urine specimens was investigated by means of gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). GHB was isolated from urinary matrix by successive purification on Oasis MCX and Bond Elute SAX solid-phase extraction (SPE) cartridges prior to high-performance liquid chromatography (HPLC) fractioning using an Atlantis dC18 column eluted with a mixture of formic acid and methanol. Subsequent intramolecular esterification of GHB leading to the formation of gamma-butyrolactone (GBL) was carried out to avoid introduction of additional carbon atoms for carbon isotopic ratio analysis. A precision of 0.3 per thousand was determined using this IRMS method for samples at GHB concentrations of 10 mg/L. The (13)C/(12)C ratios of GHB in samples of subjects exposed to the drug ranged from -32.1 to -42.1 per thousand, whereas the results obtained for samples containing GHB of endogenous origin at concentration levels less than 10 mg/L were in the range -23.5 to -27.0 per thousand. Therefore, these preliminary results show that a possible discrimination between endogenous and exogenous GHB can be made using carbon isotopic ratio analyses.

  18. Gamma-hydroxybutyric acid endogenous production and post-mortem behaviour - the importance of different biological matrices, cut-off reference values, sample collection and storage conditions.

    PubMed

    Castro, André L; Dias, Mário; Reis, Flávio; Teixeira, Helena M

    2014-10-01

    Gamma-Hydroxybutyric Acid (GHB) is an endogenous compound with a story of clinical use, since the 1960's. However, due to its secondary effects, it has become a controlled substance, entering the illicit market for recreational and "dance club scene" use, muscle enhancement purposes and drug-facilitated sexual assaults. Its endogenous context can bring some difficulties when interpreting, in a forensic context, the analytical values achieved in biological samples. This manuscript reviewed several crucial aspects related to GHB forensic toxicology evaluation, such as its post-mortem behaviour in biological samples; endogenous production values, whether in in vivo and in post-mortem samples; sampling and storage conditions (including stability tests); and cut-off reference values evaluation for different biological samples, such as whole blood, plasma, serum, urine, saliva, bile, vitreous humour and hair. This revision highlights the need of specific sampling care, storage conditions, and cut-off reference values interpretation in different biological samples, essential for proper practical application in forensic toxicology. Copyright © 2014 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

  19. Incidence of craving for and abuse of gamma-hydroxybutyric acid (GHB) in different populations of treated alcoholics: an open comparative study.

    PubMed

    Caputo, F; Francini, S; Stoppo, M; Lorenzini, F; Vignoli, T; Del Re, A; Comaschi, C; Leggio, L; Addolorato, G; Zoli, G; Bernardi, M

    2009-11-01

    Gamma-hydroxybutyric acid (GHB) is a drug currently used for the treatment of alcohol dependence. The aim of our study was to investigate the incidence of craving for and abuse of GHB in 47 patients enrolled and divided into four groups: group A (pure alcoholics), group B (alcoholics with a sustained full remission from cocaine dependence), group C (alcoholics with a sustained full remission from heroin dependence) and group D (alcoholics in a methadone maintenance treatment [MMT] programme). All patients were treated with an oral dose of GHB (50 mg/kg of body weight t.i.d.) for three months. Craving for GHB was statistically significant higher in group B than in group A (P < 0.001), C (P = 0.01) and D (P < 0.001), and in group C than in group D (P < 0.05). Abuse of GHB proved to be statistically significant higher in group B than in group A (P < 0.001) and D (P < 0.01), and in group C than in group A (P = 0.01) and D (P < 0.05). Thus, the administration of GHB in alcoholics with a sustained full remission from heroin or cocaine dependence is not recommended; however, this should not discourage physicians from using GHB for the treatment of pure alcoholics or alcohol dependents following a MMT.

  20. Detection of gamma-hydroxybutyrate in hair: validation of GC-MS and LC-MS/MS methods and application to a real case.

    PubMed

    Bertol, Elisabetta; Argo, Antonina; Procaccianti, Paolo; Vaiano, Fabio; Di Milia, Maria Grazia; Furlanetto, Sandra; Mari, Francesco

    2012-11-01

    A gas chromatography-mass spectrometry (GC-MS) and a liquid chromatography tandem mass spectrometry (LC-MS/MS) method were validated for quantifying endogenous and exogenous hair concentrations of gamma-hydroxybutyrate (GHB). The GC-MS method is based on overnight extraction of 25 mg hair in NaOH at 56 °C, liquid/liquid extraction in ethylacetate and trimethylsylil derivatization; analysis is by electron ionization and single ion monitoring of three ions. The LC-MS/MS method entails a rapid digestion of 25 mg hair with NaOH at 75 °C for 40 min, liquid/liquid extraction in ethylacetate and reconstitution of the extract in the LC mobile phase; negative ion electrospray ionization and multiple reaction monitoring (MRM) analysis are employed for the LC-MS/MS detection. In both cases, GHB-d6 is used as an internal standard. The endogenous amount in "blank" hair are estimated by the standard addition method. Limits of detection are 0.4 and 0.5 ng/mg for GC-MS and LC-MS/MS respectively, while the limit of quantification (LOQ) is 0.6 ng/mg for both methods; the GC-MS method proved to be linear in the range 1-50 ng/mg whereas linearity was demonstrated from 0.6 to 50 ng/mg for the LC-MS/MS; imprecision and inaccuracy were always lower than 23% for quality controls samples. The two methods were applied to a real case of a man addicted to GHB; the drug concentration in segments from 17 cm hair strand well correlated with self-reported use of GHB in different periods of his life. Performances of the two methods were similar.

  1. Intoxication by gamma hydroxybutyrate and related analogues: Clinical characteristics and comparison between pure intoxication and that combined with other substances of abuse.

    PubMed

    Miró, Òscar; Galicia, Miguel; Dargan, Paul; Dines, Alison M; Giraudon, Isabelle; Heyerdahl, Fridtjof; Hovda, Knut E; Yates, Christopher; Wood, David M

    2017-08-05

    To study the profile of European gamma-hydroxybutyrate (GHB) and gammabutyrolactone (GBL) intoxication and analyse the differences in the clinical manifestations produced by intoxication by GHB/GBL alone and in combination with other substances of abuse. We prospectively collected data on all the patients attended in the Emergency Departments (ED) of the centres participating in the Euro-DEN network over 12 months (October 2013 to September 2014) with a primary presenting complaint of drug intoxication (excluding ethanol alone) and registered the epidemiological and clinical data and outcomes. We included 710 cases (83% males, mean age 31 years), representing 12.6% of the total cases attended for drug intoxication. Of these, 73.5% arrived at the ED by ambulance, predominantly during weekend, and 71.7% consumed GHB/GBL in combination with other substances of abuse, the most frequent additional agents being ethanol (50%), amphetamine derivatives (36%), cocaine (12%) and cannabis (8%). Among 15 clinical features pre-defined in the project database, the 3 most frequently identified were altered behaviour (39%), reduced consciousness (34%) and anxiety (14%). The severity ranged from mild cases requiring no treatment (308 cases, 43.4%) to severe cases requiring admission to intensive care (103 cases, 14.6%) and mechanical ventilation (49 cases, 6.9%). No deaths were reported. In comparison with only GHB/GBL consumption, patients consuming GHB/GBL with co-intoxicants presented more vomiting (15% vs. 3%, p<0.001) and cardiovascular symptoms (5.3% vs. 1.5%, p<0.05), a greater need for treatment (59.8% vs. 48.3%, p<0.01) and a longer ED stay (11.3% vs. 3.6% patients with ED stay >12h, p<0.01). The profile of the typical GHB/GBL-intoxicated European is a young male, requiring care for altered behaviour and reduced level of consciousness, mainly during the weekend. The clinical features are more severe when GHB is consumed in combination with other substances of abuse

  2. Effects of combining ethanol (EtOH) with gamma-hydroxybutyrate (GHB) on the discriminative stimulus, locomotor, and motor-impairing functions of GHB in mice.

    PubMed

    Cook, Charles D; Biddlestone, Laura; Coop, Andrew; Beardsley, Patrick M

    2006-03-01

    Gamma-hydroxybutyrate (GHB) is a drug of abuse that is often coabused with ethanol (EtOH) and has been implicated as a date rape agent in conjunction with EtOH. Much information is lacking regarding the manner in which GHB interacts with EtOH. This study was designed to further characterize the behavioral effects of GHB alone and in combination with EtOH in male Swiss-Webster mice. The effects of GHB (0.1-1.0 g/kg) and EtOH (2.0-5.0 g/kg) alone, as well as the effects of GHB in combination with EtOH, were examined using an automated locomotor activity procedure, a functional observational battery (FOB) and a GHB drug discrimination procedure. GHB decreased, whereas EtOH had little effect on locomotor activity. In the FOB, EtOH dose-dependently decreased activity in combination with 0.3 g/kg GHB. Alone, each drug had little effect on the righting reflex, but combining ineffective doses of GHB and EtOH significantly impaired righting. GHB and EtOH decreased forelimb grip strength. Combinations of ineffective doses of GHB and EtOH decreased forelimb grip strength when given together. GHB and EtOH impaired inverted screen performance, and EtOH increased the impairing effects of low, but not high, doses of GHB. GHB and EtOH increased hind limb splay, and EtOH increased the effects of 0.1 and 0.3 g/kg GHB on splay. GHB and EtOH decreased body temperature, and EtOH augmented the temperature-decreasing effects of GHB. EtOH produced less than 50% GHB-like discriminative stimulus effects, and GHB failed to alter the GHB-like discriminative stimulus effects of EtOH. Overall, EtOH increased the effects of GHB on several gross measures of behavior and only partially occasioned the discriminative stimulus properties of GHB.

  3. GHB - Gamma-Hydroxybutyric Acid

    MedlinePlus

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  4. P2X7 receptors stimulate AKT phosphorylation in astrocytes

    PubMed Central

    Jacques-Silva, Maria C; Rodnight, Richard; Lenz, Guido; Liao, Zhongji; Kong, Qiongman; Tran, Minh; Kang, Yuan; Gonzalez, Fernando A; Weisman, Gary A; Neary, Joseph T

    2004-01-01

    Emerging evidence indicates that nucleotide receptors are widely expressed in the nervous system. Here, we present evidence that P2Y and P2X receptors, particularly the P2X7 subtype, are coupled to the phosphoinositide 3-kinase (PI3K)/Akt pathway in astrocytes. P2Y and P2X receptor agonists ATP, uridine 5′-triphosphate (UTP) and 2′,3′-O-(4-benzoyl)-benzoyl ATP (BzATP) stimulated Akt phosphorylation in primary cultures of rat cortical astrocytes. BzATP induced Akt phosphorylation in a concentration- and time-dependent manner, similar to the effect of BzATP on Akt phosphorylation in 1321N1 astrocytoma cells stably transfected with the rat P2X7 receptor. Activation was maximal at 5 – 10 min and was sustained for 60 min; the EC50 for BzATP was approximately 50 μM. In rat cortical astrocytes, the positive effect of BzATP on Akt phosphorylation was independent of glutamate release. The effect of BzATP on Akt phosphorylation in rat cortical astrocytes was significantly reduced by the P2X7 receptor antagonist Brilliant Blue G and the P2X receptor antagonist iso-pyridoxal-5′-phosphate-6-azophenyl-2′,4′-disulfonic acid, but was unaffected by trinitrophenyl-ATP, oxidized ATP, suramin and reactive blue 2. Results with specific inhibitors of signal transduction pathways suggest that extracellular and intracellular calcium, PI3K and a Src family kinase are involved in the BzATP-induced Akt phosphorylation pathway. In conclusion, our data indicate that stimulation of astrocytic P2X7 receptors, as well as other P2 receptors, leads to Akt activation. Thus, signaling by nucleotide receptors in astrocytes may be important in several cellular downstream effects related to the Akt pathway, such as cell cycle and apoptosis regulation, protein synthesis, differentiation and glucose metabolism. PMID:15023862

  5. Reflex effects on the heart of stimulating left atrial receptors

    PubMed Central

    Furnival, C. M.; Linden, R. J.; Snow, H. M.

    1971-01-01

    1. Stimulation of left atrial receptors, by distension of the pulmonary vein/left atrial junctions, is known to cause a reflex increase in heart rate; the efferent pathway is known to be solely in the sympathetic nerves. 2. In expectation of a concomitant positive inotropic response the effect of stimulating the left atrial receptors on the inotropic state of the left ventricle was studied, using as a known sensitive index of inotropic changes the maximal rate of rise of pressure in the left ventricle (dP/dt max). 3. Stimulation of left atrial receptors resulted in an increase in heart rate but there were no significant concomitant changes in dP/dt max. 4. It is concluded that activity in this discrete efferent pathway does not include an inotropic effect on the left ventricle and therefore the reflex involves only those sympathetic nerves which innervate the sinu-atrial node. 5. The possible function of atrial receptors in the regulation of heart volumes is discussed. PMID:5124571

  6. Intrarenal dopamine D1-like receptor stimulation induces natriuresis via an angiotensin type-2 receptor mechanism.

    PubMed

    Salomone, Leslie J; Howell, Nancy L; McGrath, Helen E; Kemp, Brandon A; Keller, Susanna R; Gildea, John J; Felder, Robin A; Carey, Robert M

    2007-01-01

    We explored the effects of direct renal interstitial stimulation of dopamine D(1)-like receptors with fenoldopam, a selective D(1)-like receptor agonist, on renal sodium excretion and angiotensin type-2 (AT(2)) receptor expression and cellular distribution in rats on a high-sodium intake. In contrast to vehicle-infused rats, sodium excretion increased in fenoldopam-infused rats during each of three 1-hour experimental periods (<0.001). Blood pressure was unaffected by vehicle or fenoldopam. In plasma membranes of renal cortical cells, fenoldopam increased D(1) receptor expression by 38% (P<0.05) and AT(2) receptor expression by 69% (P<0.01). In plasma membranes of renal proximal tubule cells, fenoldopam increased AT(2) receptor expression by 108% (P<0.01). In outer apical membranes of proximal tubule cells, fenoldopam increased AT(2) receptor expression by 59% (P<0.01). No significant change in total AT(2) receptor protein expression was detectable in response to fenoldopam. Fenoldopam-induced natriuresis was abolished when either PD-123319, a specific AT(2) receptor antagonist, or SCH-23390, a potent D(1)-like receptor antagonist, was coinfused with F (P<0.001). In summary, direct renal D(1)-like receptor activation increased urinary sodium excretion and the plasma membrane expression of AT(2) receptors in renal cortical and proximal tubule cells. D(1)-like receptor-induced natriuresis was abolished by intrarenal AT(2) receptor inhibition. These findings suggest that dopaminergic regulation of sodium excretion involves recruitment of AT(2) receptors to the outer plasma membranes of renal proximal tubule cells and that dopamine-induced natriuresis requires AT(2) receptor activation.

  7. Purinergic receptor stimulation increases membrane trafficking in brown adipocytes

    PubMed Central

    1996-01-01

    Stimulation of brown adipocytes by their sympathetic innervation plays a major role in body energy homeostasis by regulating the energy- wasting activity of the tissue. The norepinephrine released by sympathetic activity acts on adrenergic receptors to activate a variety of metabolic and membrane responses. Since sympathetic stimulation may also release vesicular ATP, we tested brown fat cells for ATP responses. We find that micromolar concentrations of extracellular ATP initiates profound changes in the membrane trafficking of brown adipocytes. ATP elicited substantial increases in total cell membrane capacitance, averaging approximately 30% over basal levels and occurring on a time scale of seconds to minutes. The membrane capacitance increase showed an agonist sensitivity of 2-methylthio-ATP > or = ATP > ADP > > adenosine, consistent with mediation by a P2r type purinergic receptor. Membrane capacitance increases were not seen when cytosolic calcium was increased by adrenergic stimulation, and capacitance responses to ATP were similar in the presence and absence of extracellular calcium. These results indicate that increases in cytosolic calcium alone do not mediate the membrane response to ATP. Photometric assessment of surface-accessible membrane using the dye FM1- 43 showed that ATP caused an approximate doubling of the amount of membrane actively trafficking with the cell surface. The discrepancy in the magnitudes of the capacitance and fluorescence changes suggests that ATP both activates exocytosis and alters other aspects of membrane handling. These findings suggest that secretion, mobilization of membrane transporters, and/or surface membrane expression of receptors may be regulated in brown adipocytes by P2r purinergic receptor activity. PMID:8923265

  8. Analysis of gamma-hydroxybutyric acid (GHB) in spiked water and beverage samples using solid phase microextraction (SPME) on fiber derivatization/gas chromatography-mass spectrometry (GC/MS).

    PubMed

    Meyers, Jodi E; Almirall, José R

    2005-01-01

    Gamma-Hydroxybutyric acid (GHB) is a CNS depressant that has been abused recreationally for its purported euphoric and relaxation effects and for the purposes of drug facilitated sexual assault due to its sedative and amnesic effects at higher doses. The dramatic increase in the abuse of GHB and association in criminal investigations over the past decade has created the need for forensic laboratories to develop analytical methods to detect GHB in a variety of matrices. The method developed in this work used solid-phase microextraction (SPME) to extract GHB from aqueous samples followed by on-fiber derivatization and analysis by gas chromatography/mass spectrometry (GC/MS). This method detected GHB in aqueous matrices with good sensitivity, high precision, excellent linearity from 0.01 mg/mL to 0.25 mg/mL, and without the need for sample manipulation that could cause interconversion between GHB and its lactone, GBL. The method was successfully applied for detection of GHB in spiked water and beverage samples.

  9. Comparative study of equimolar doses of gamma-hydroxybutyrate (GHB), 1,4-butanediol (1,4-BD) and gamma-butyrolactone (GBL) on catalepsy after acute and chronic administration.

    PubMed

    Towiwat, Pasarapa; Phattanarudee, Siripan; Maher, Timothy J

    2013-01-01

    Gamma-hydroxybutyrate (GHB), and its precursors 1,4-butanediol (1,4-BD) and gamma-butyrolactone (GBL) are known drugs of abuse. The ability of acute and chronic administration of equimolar doses of GHB (200mg/kg), 1,4-BD (174mg/kg) and GBL (166mg/kg) to produce catalepsy in male Swiss Webster mice was examined. GHB, 1,4-BD, GBL produced catalepsy when injected acutely. Drug treatment was then continued for 14days. Tolerance development was determined on days 6, 14, and challenged with a higher dose on day 15 in those chronically pretreated mice, and compared with naïve mice. Chronic GHB produced tolerance to catalepsy, as evidenced from area under the curve (AUC) of catalepsy versus time (min-sec) on days 6 (678±254), 14 (272±247), which were less than those on day 1 (1923±269). However, less tolerance was seen from GBL or 1,4-BD, as AUCs on days 6 and 14 were not significantly lower than that of day 1. In conclusion, although equimolar doses were used, expecting similar levels of GHB in the body, 1,4-BD and GBL shared only some of the in vivo effects of GHB. The rate of metabolic conversion of 1,4-BD and GBL into GHB might be responsible for the differences in the tolerance development to these drugs.

  10. NMDA receptor blockade attenuates locomotion elicited by intrastriatal dopamine D1-receptor stimulation.

    PubMed

    Kreipke, Christian W; Walker, Paul D

    2004-07-01

    Previous behavioral studies suggest that the striatum mediates a hyperactive response to systemic NMDA receptor antagonism in combination with systemic D1 receptor stimulation. However, many experiments conducted at the cellular level suggest that inhibition of NMDA receptors should block D1 receptor-mediated locomotor activity. Therefore, we investigated the consequences of NMDA receptor blockade on the ability of striatal D1 receptors to elicit locomotor activity using systemic and intrastriatal injections of the NMDA antagonist MK-801 combined with intrastriatal injections of the D1 full agonist SKF 82958. Following drug treatment locomotor activity was measured via computerized activity monitors designed to quantify multiple parameters of rodent open-field behavior. Both systemic (0.1 mg/kg) and intrastriatal (1.0 microg) MK-801 pretreatments completely blocked locomotor and stereotypic activity elicited by 10 microg of SKF 82958 directly infused into the striatum. Further, increased activity triggered by intrastriatal SKF 82958 was attenuated by a posttreatment with intrastriatal infusion of 1 microg MK-801. These data suggest that D1-stimulated locomotor behaviors controlled by the striatum require functional NMDA channels.

  11. Cannabinoid receptor stimulation increases motivation for nicotine and nicotine seeking.

    PubMed

    Gamaleddin, Islam; Wertheim, Carrie; Zhu, Andy Z X; Coen, Kathleen M; Vemuri, Kiran; Makryannis, Alex; Goldberg, Steven R; Le Foll, Bernard

    2012-01-01

    The cannabinoid system appears to play a critical facilitative role in mediating the reinforcing effects of nicotine and relapse to nicotine-seeking behaviour in abstinent subjects based on the actions of cannabinoid (CB) receptor antagonists. However, the effects of CB receptor stimulation on nicotine self-administration and reinstatement have not been systematically studied. Here, we studied the effects of WIN 55,212-2, a CB1/2 agonist, on intravenous nicotine self-administration under fixed-ratio (FR) and progressive-ratio (PR) schedules of reinforcement in rats. The effects of WIN 55,212-2 on responding for food under similar schedules were also studied. In addition, the effects of WIN 55,212-2 on nicotine- and cue-induced reinstatement of nicotine seeking were also studied, as well as the effects of WIN 55,212-2 on nicotine discrimination. WIN 55,212-2 decreased nicotine self-administration under the FR schedule. However, co-administration of WIN 55,212-2 with nicotine decreased responding for food, which suggests that this effect was non-selective. In contrast, WIN 55,212-2 increased both nicotine self-administration and responding for food under the PR schedule, produced dose-dependent reinstatement of nicotine seeking, and enhanced the reinstatement effects of nicotine-associated cues. Some of these effects were reversed by the CB1 antagonist rimonabant, but not by the CB2 antagonist AM630. In the drug discrimination tests between saline and 0.4 mg/kg nicotine, WIN 55,212-2 produced no nicotine-like discriminative effects but significantly potentiated discriminative stimulus effects of nicotine at the low dose through a CB1-receptor-dependent mechanism. These findings indicate that cannabinoid CB1-receptor stimulation increases the reinforcing effects of nicotine and precipitates relapse to nicotine-seeking behaviour in abstinent subjects. Thus, modulating CB1-receptor signalling might have therapeutic value for treating nicotine dependence. © 2011 The

  12. Glycosides from edible sea cucumbers stimulate macrophages via purinergic receptors

    PubMed Central

    Aminin, Dmitry; Pislyagin, Evgeny; Astashev, Maxim; Es’kov, Andrey; Kozhemyako, Valery; Avilov, Sergei; Zelepuga, Elena; Yurchenko, Ekaterina; Kaluzhskiy, Leonid; Kozlovskaya, Emma; Ivanov, Alexis; Stonik, Valentin

    2016-01-01

    Since ancient times, edible sea cucumbers have been considered a jewel of the seabed and used in Asian folk medicine for stimulation of resistance against different diseases. However, the power of this sea food has not been established on a molecular level. A particular group of triterpene glycosides was found to be characteristic metabolites of the animals, responsible for this biological action. Using one of them, cucumarioside A2-2 (CA2-2) from the edible Cucumaria japonica species as an example as well as inhibitory analysis, patch-clamp on single macrophages, small interfering RNA technique, immunoblotting, SPR analysis, computer modeling and other methods, we demonstrate low doses of CA2-2 specifically to interact with P2X receptors (predominantly P2X4) on membranes of mature macrophages, enhancing the reversible ATP-dependent Ca2+ intake and recovering Ca2+ transport at inactivation of these receptors. As result, interaction of glycosides of this type with P2X receptors leads to activation of cellular immunity. PMID:28004778

  13. When a death apparently associated to sexual assault is instead a natural death due to idiopathic hypereosinophilic syndrome: The importance of gamma-hydroxybutyric acid analysis in vitreous humor.

    PubMed

    Busardò, Francesco Paolo; Portelli, Francesca; Montana, Angelo; Rotolo, Maria Concetta; Pichini, Simona; Maresi, Emiliano

    2017-05-01

    We here report a case involving a 21-year-old female, found dead in a central square of a city in the south of Italy. Initial evidences and circumstances were suggestive of a death associated with a sexual assault. Two peripheral blood and two vitreous humor samples were collected for the purpose of gamma-hydroxybutyric acid (GHB) testing from the dead body at two different post-mortem intervals (PMIs): approximately 2 (t0) and 36 (t1) hours. The obtained results showed that, between t0 and t1, there was an increase of GHB concentrations in peripheral blood and vitreous humor of 66.3% and 8.1%, respectively. This case was the first evidence of GHB post mortem production in a dead body and not in vitro, showing that vitreous humor is less affected than peripheral blood in GHB post-mortem production. The value detected at t1 in peripheral blood (53.4µg/mL) exceeded the proposed cut-off and if interpreted alone would have led to erroneous conclusions. This was not the case of vitreous humor GHB, whose post-mortem increase was minimal and it allowed to exclude a GHB exposure. Only after a broad forensic investigation including a complete autopsy, serological, histological, toxicological and haematology analyses, a diagnosis of idiopathic hypereosinophilic syndrome, a myeloproliferative disorder characterized by persistent eosinophilia associated with damage to multiple organs, was made and the cause of death was due to a pulmonary eosinophilic vasculitis responsible for an acute respiratory failure. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. THIP and isoguvacine are partial agonists of GABA-stimulated benzodiazepine receptor binding.

    PubMed

    Karobath, M; Lippitsch, M

    1979-10-15

    The effects of THIP and isoguvacine on 3H-flunitrazepam binding to washed membranes prepared from the cerebral cortex of adult rats have been examined. THIP, which has only minimal stimulatory effects on benzodiazepine (BZ) receptor binding, has been found to inhibit the stimulation induced by small concentrations (2 microM) of exogenous GABA. While isoguvacine stimulates BZ receptor binding, although to a smaller extent than GABA, it also antagonizes the stimulation of BZ receptor binding induced by GABA. Thus THIP and isoguvacine exhibit the properties of a partial agonist of GABA-stimulated BZ receptor binding.

  15. Transferrin receptor expression by stimulated cells in mixed lymphocyte culture.

    PubMed Central

    Salmon, M; Bacon, P A; Symmons, D P; Walton, K W

    1985-01-01

    Transferrin receptor (TRFr) expression by cells in mixed lymphocyte culture increases steadily for the first 5 days, but then reaches a plateau. By the sixth day in culture, about 20% of viable cells express TRFr in two-way mixed lymphocyte reactions. This subpopulation of TRFr-positive cells represents the proliferating population; it is heterogeneous, containing T-cell blasts and smaller cells which are a mixture of T and non-T cells. A small group of non-T cells have phenotypic similarity to natural killer (NK) cells. T cells appear to divide earlier in the course of the response than non-T cells. The biphasic nature of this response and the slower non-T reactivity may be due to a secondary stimulation of non-T cells by factors released from activated T cells (such as interleukin-2). PMID:2982734

  16. Chicken interferons, their receptors and interferon-stimulated genes.

    PubMed

    Goossens, Kate E; Ward, Alister C; Lowenthal, John W; Bean, Andrew G D

    2013-11-01

    The prevalence of pathogenic viruses is a serious issue as they pose a constant threat to both the poultry industry and to human health. To prevent these viral infections an understanding of the host-virus response is critical, especially for the development of novel therapeutics. One approach in the control of viral infections would be to boost the immune response through administration of cytokines, such as interferons. However, the innate immune response in chickens is poorly characterised, particularly concerning the interferon pathway. This review will provide an overview of our current understanding of the interferon system of chickens, including their cognate receptors and known interferon-stimulated gene products. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Thrombin stimulates insulin secretion via protease-activated receptor-3.

    PubMed

    Hänzelmann, Sonja; Wang, Jinling; Güney, Emre; Tang, Yunzhao; Zhang, Enming; Axelsson, Annika S; Nenonen, Hannah; Salehi, Albert S; Wollheim, Claes B; Zetterberg, Eva; Berntorp, Erik; Costa, Ivan G; Castelo, Robert; Rosengren, Anders H

    2015-01-01

    The disease mechanisms underlying type 2 diabetes (T2D) remain poorly defined. Here we aimed to explore the pathophysiology of T2D by analyzing gene co-expression networks in human islets. Using partial correlation networks we identified a group of co-expressed genes ('module') including F2RL2 that was associated with glycated hemoglobin. F2Rl2 is a G-protein-coupled receptor (GPCR) that encodes protease-activated receptor-3 (PAR3). PAR3 is cleaved by thrombin, which exposes a 6-amino acid sequence that acts as a 'tethered ligand' to regulate cellular signaling. We have characterized the effect of PAR3 activation on insulin secretion by static insulin secretion measurements, capacitance measurements, studies of diabetic animal models and patient samples. We demonstrate that thrombin stimulates insulin secretion, an effect that was prevented by an antibody that blocks the thrombin cleavage site of PAR3. Treatment with a peptide corresponding to the PAR3 tethered ligand stimulated islet insulin secretion and single β-cell exocytosis by a mechanism that involves activation of phospholipase C and Ca(2+) release from intracellular stores. Moreover, we observed that the expression of tissue factor, which regulates thrombin generation, was increased in human islets from T2D donors and associated with enhanced β-cell exocytosis. Finally, we demonstrate that thrombin generation potential in patients with T2D was associated with increased fasting insulin and insulinogenic index. The findings provide a previously unrecognized link between hypercoagulability and hyperinsulinemia and suggest that reducing thrombin activity or blocking PAR3 cleavage could potentially counteract the exaggerated insulin secretion that drives insulin resistance and β-cell exhaustion in T2D.

  18. In vivo stimulation of oestrogen receptor α increases insulin-stimulated skeletal muscle glucose uptake.

    PubMed

    Gorres, Brittany K; Bomhoff, Gregory L; Morris, Jill K; Geiger, Paige C

    2011-04-15

    Previous studies suggest oestrogen receptor α (ERα) is involved in oestrogen-mediated regulation of glucose metabolism and is critical for maintenance of whole body insulin action. Despite this, the effect of direct ERα modulation in insulin-responsive tissues is unknown. The purpose of the current study was to determine the impact of ERα activation, using the ER subtype-selective ligand propylpyrazoletriyl (PPT), on skeletal muscle glucose uptake. Two-month-old female Sprague-Dawley rats, ovariectomized for 1 week, were given subcutaneous injections of PPT (10 mg kg⁻¹), oestradiol benzoate (EB; 20 μg kg⁻¹), the ERβ agonist diarylpropionitrile (DPN, 10 mg kg⁻¹) or vehicle every 24 h for 3 days. On the fourth day, insulin-stimulated skeletal muscle glucose uptake was measured in vitro and insulin signalling intermediates were assessed via Western blotting.Activation of ERα with PPT resulted in increased insulin-stimulated glucose uptake into the slow-twitch soleus and fast-twitch extensor digitorum longus (EDL)muscles, activation of insulin signalling intermediates (as measured by phospho-Akt (pAkt) and pAkt substrate (PAS)) and phosphorylation of AMP-activated protein kinase (AMPK). GLUT4 protein was increased only in the EDL muscle. Rats treated with EB or DPN for 3 days did not show an increase in insulin-stimulated skeletal muscle glucose uptake compared to vehicle-treated animals. These new findings reveal that direct activation of ERα positively mediates glucose uptake and insulin action in skeletal muscle. Evidence that oestrogens and ERα stimulate glucose uptake has important implications for understanding mechanisms of glucose homeostasis, particularly in postmenopausal women.

  19. Odorant-stimulated phosphoinositide signaling in mammalian olfactory receptor neurons

    PubMed Central

    Klasen, K.; Corey, E.A.; Kuck, F.; Wetzel, C.H.; Hatt, H.; Ache, B.W.

    2009-01-01

    Recent evidence has revived interest in the idea that phosphoinositides (PIs) may play a role in signal transduction in mammalian olfactory receptor neurons (ORNs). To provide direct evidence that odorants indeed activate PI signaling in ORNs, we used adenoviral vectors carrying two different fluorescently tagged probes, the pleckstrin homology (PH) domains of phospholipase Cδ1 (PLCδ1) and the general receptor of phosphoinositides (GRP1), to monitor PI activity in the dendritic knobs of ORNs in vivo. Odorants mobilized PI(4,5)P2/IP3 and PI(3,4,5)P3, the substrates and products of PLC and PI3K. We then measured odorant activation of PLC and PI3K in olfactory ciliary-enriched membranes in vitro using a phospholipid overlay assay and ELISAs. Odorants activated both PLC and PI3K in the olfactory cilia within 2 sec of odorant stimulation. Odorant-dependent activation of PLC and PI3K in the olfactory epithelium could be blocked by enzyme-specific inhibitors. Odorants activated PLC and PI3K with partially overlapping specificity. These results provide direct evidence that odorants indeed activate PI signaling in mammalian ORNs in a manner that is consistent with the idea that PI signaling plays a role in olfactory transduction. PMID:19781634

  20. Behavioral meaningful opioidergic stimulation activates kappa receptor gene expression

    PubMed Central

    Teodorov, E.; Ferrari, M.F.R.; Fior-Chadi, D.R.; Camarini, R.; Felício, L.F.

    2012-01-01

    The periaqueductal gray (PAG) has been reported to be a location for opioid regulation of pain and a potential site for behavioral selection in females. Opioid-mediated behavioral and physiological responses differ according to the activity of opioid receptor subtypes. The present study investigated the effects of the peripheral injection of the kappa-opioid receptor agonist U69593 into the dorsal subcutaneous region of animals on maternal behavior and on Oprk1 gene activity in the PAG of female rats. Female Wistar rats weighing 200-250 g at the beginning of the study were randomly divided into 2 groups for maternal behavior and gene expression experiments. On day 5, pups were removed at 7:00 am and placed in another home cage that was distant from their mother. Thirty minutes after removing the pups, the dams were treated with U69593 (0.15 mg/kg, sc) or 0.9% saline (up to 1 mL/kg) and after 30 min were evaluated in the maternal behavior test. Latencies in seconds for pup retrieval, grouping, crouching, and full maternal behavior were scored. The results showed that U69593 administration inhibited maternal behavior (P < 0.05) because a lower percentage of U69593 group dams showed retrieval of first pup, retrieving all pups, grouping, crouching and displaying full maternal behavior compared to the saline group. Opioid gene expression was evaluated using real-time reverse-transcription polymerase chain reaction (RT-PCR). A single injection of U69593 increased Oprk1 PAG expression in both virgin (P < 0.05) and lactating female rats (P < 0.01), with no significant effect on Oprm1 or Oprd1 gene activity. Thus, the expression of kappa-opioid receptors in the PAG may be modulated by single opioid receptor stimulation and behavioral meaningful opioidergic transmission in the adult female might occur simultaneously to specific changes in gene expression of kappa-opioid receptor subtype. This is yet another alert for the complex role of the opioid system in female

  1. Dark chocolate receptors: epicatechin-induced cardiac protection is dependent on δ-opioid receptor stimulation

    PubMed Central

    Panneerselvam, Mathivadhani; Tsutsumi, Yasuo M.; Bonds, Jacqueline A.; Horikawa, Yousuke T.; Saldana, Michelle; Dalton, Nancy D.; Head, Brian P.; Patel, Piyush M.; Roth, David M.

    2010-01-01

    Epicatechin, a flavonoid, is a well-known antioxidant linked to a variety of protective effects in both humans and animals. In particular, its role in protection against cardiovascular disease has been demonstrated by epidemiologic studies. Low-dose epicatechin, which does not have significant antioxidant activity, is also protective; however, the mechanism by which low-dose epicatechin induces this effect is unknown. Our laboratory tested the hypothesis that low-dose epicatechin mediates cardiac protection via opioid receptor activation. C57BL/6 mice were randomly assigned to 1 of 10 groups: control, epicatechin, naloxone (nonselective opioid receptor antagonist), epicatechin + naloxone, naltrindole (δ-specific opioid receptor antagonist), epicatechin + naltrindole, norbinaltorphimine (nor-BNI, κ-specific opioid receptor antagonist), epicatechin + nor-BNI, 5-hydroxydecanoic acid [5-HD, ATP-sensitive potassium channel antagonist], and epicatechin + 5-HD. Epicatechin (1 mg/kg) or other inhibitors (5 mg/kg) were administered by oral gavage or intraperitoneal injection, respectively, daily for 10 days. Mice were subjected to 30 min coronary artery occlusion followed by 2 h of reperfusion, and infarct size was determined via planimetry. Whole heart homogenates were assayed for downstream opioid receptor signaling targets. Infarct size was significantly reduced in epicatechin- and epicatechin + nor-BNI-treated mice compared with control mice. This protection was blocked by naloxone, naltrindole, and 5-HD. Epicatechin and epicatechin + nor-BNI increased the phosphorylation of Src, Akt, and IκBα, while simultaneously decreasing the expression of c-Jun NH2-terminal kinase and caspase-activated DNase. All signaling effects are consistent with opioid receptor stimulation and subsequent cardiac protection. Naloxone, naltrindole, and 5-HD attenuated these effects. In conclusion, epicatechin acts via opioid receptors and more specifically through the δ-opioid receptor to

  2. Stimulation of Estrogen Receptor Signaling in Breast Cancer by a Novel Chaperone Synuclein Gamma

    DTIC Science & Technology

    2006-06-01

    AD_________________ Award Number: W81XWH- 04 -1-0569 TITLE: Stimulation of estrogen receptor...Stimulation of estrogen receptor signaling in breast cancer by a novel chaperone 5a. CONTRACT NUMBER synuclein gamma 5b. GRANT NUMBER W81XWH- 04 -1...UNIT NUMBER 7 . PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) 8. PERFORMING ORGANIZATION REPORT NUMBER North Shore University Hospital

  3. An Interactive Lesson in Acid/Base and Pro-Drug Chemistry Using Sodium Gamma-Hydroxybutyrate and Commercial Test Coasters

    PubMed Central

    Page, Nathaniel A.; Paganelli, Meaghan; Boje, Kathleen M.K.

    2007-01-01

    Objective To develop a classroom activity that applied pertinent pharmaceutical concepts to examine the use and limitations of a commercially available test drink coaster in detecting the presence of a date-rape drug, sodium γ-hydroxybutyrate (NaGHB), in beverages. Design An activity exercise involving a combination of self-study, hands on participation, and classroom discussion was developed. Topics incorporated into the activity were drug-assisted rape, the concepts of false positives and negatives, and prodrug and pH chemistry. Assessment Based on questionnaires completed by the students, the intended concepts were reinforced and students demonstrated an increased awareness of the potential shortcomings of the commercial test devices. The activity was well received by the majority of students. Conclusion The developed activity stimulated student awareness and interest in several principles relevant in pharamceutical education, including drug-assisted rape, consumer-based drug testing of NaGHB, and the chemical basis for its limitations. The activity requires no special equipment other than the drink coasters and can be easily completed in one 2-hour classroom session. PMID:17619654

  4. Thyroid-Stimulating Hormone Receptor Antibodies in Pregnancy: Clinical Relevance

    PubMed Central

    Bucci, Ines; Giuliani, Cesidio; Napolitano, Giorgio

    2017-01-01

    Graves’ disease is the most common cause of thyrotoxicosis in women of childbearing age. Approximately 1% of pregnant women been treated before, or are being treated during pregnancy for Graves’ hyperthyroidism. In pregnancy, as in not pregnant state, thyroid-stimulating hormone (TSH) receptor (TSHR) antibodies (TRAbs) are the pathogenetic hallmark of Graves’ disease. TRAbs are heterogeneous for molecular and functional properties and are subdivided into activating (TSAbs), blocking (TBAbs), or neutral (N-TRAbs) depending on their effect on TSHR. The typical clinical features of Graves’ disease (goiter, hyperthyroidism, ophthalmopathy, dermopathy) occur when TSAbs predominate. Graves’ disease shows some peculiarities in pregnancy. The TRAbs disturb the maternal as well as the fetal thyroid function given their ability to cross the placental barrier. The pregnancy-related immunosuppression reduces the levels of TRAbs in most cases although they persist in women with active disease as well as in women who received definitive therapy (radioiodine or surgery) before pregnancy. Changes of functional properties from stimulating to blocking the TSHR could occur during gestation. Drug therapy is the treatment of choice for hyperthyroidism during gestation. Antithyroid drugs also cross the placenta and therefore decrease both the maternal and the fetal thyroid hormone production. The management of Graves’ disease in pregnancy should be aimed at maintaining euthyroidism in the mother as well as in the fetus. Maternal and fetal thyroid dysfunction (hyperthyroidism as well as hypothyroidism) are in fact associated with several morbidities. Monitoring of the maternal thyroid function, TRAbs measurement, and fetal surveillance are the mainstay for the management of Graves’ disease in pregnancy. This review summarizes the biochemical, immunological, and therapeutic aspects of Graves’ disease in pregnancy focusing on the role of the TRAbs in maternal and fetal

  5. Monoclonal antibodies to the insulin receptor stimulate the intrinsic tyrosine kinase activity by cross-linking receptor molecules.

    PubMed

    O'Brien, R M; Soos, M A; Siddle, K

    1987-12-20

    The effect of monoclonal anti-insulin receptor antibodies on the intrinsic kinase activity of solubilized receptor was investigated. Antibodies for six distinct epitopes stimulated receptor autophosphorylation and kinase activity towards exogenous substrates. This effect of antibodies was seen only within a narrow concentration range and monovalent antibody fragments were ineffective. Evidence was obtained by sucrose density-gradient centrifugation for the formation of antibody-receptor complexes which involved both inter- and intra-molecular cross-linking, although stimulation of autophosphorylation appeared to be preferentially associated with the latter. There was partial additivity between the effects of insulin and antibodies in stimulating autophosphorylation, although the sites of phosphorylation appeared identical on two-dimensional peptide maps. Antibodies for two further epitopes failed to activate receptor kinase, but inhibited its stimulation by insulin. The effects of antibodies on kinase activity paralleled their metabolic effects on adipocytes, except for one antibody which was potently insulin-like in its metabolic effects, but which antagonized insulin stimulation of kinase activity. It is concluded that antibodies activate the receptor by cross-linking subunits rather than by reacting at specific epitopes. The ability of some antibodies to activate receptor may depend on receptor environment as well as the disposition of epitopes.

  6. Ventilatory stimulation by dopamine-receptor antagonists in the mouse.

    PubMed Central

    Olson, L. G.; Saunders, N. A.

    1985-01-01

    Ventilation was measured by a plethysmographic method in awake mice before and after intraperitoneal injection of neuroleptic drugs to test the hypothesis that dopaminergic mechanisms modulate control of breathing in this species. Dose-dependent augmentation of ventilation at rest and during hypoxia, and reduced ventilation during hypercapnia was demonstrated for haloperidol, droperidol, prochlorperazine and chlorpromazine (P less than 0.05 or less for each drug). Doses of drugs causing maximal increase of the ventilatory response to hypoxia were linearly related (r = 0.98, P less than 0.001) to in vitro affinity of the drugs for dopamine receptors. Despite presumed equal dopamine-receptor blockade, the drugs had unequal effects on the ventilatory response to hypoxia. Droperidol augmented hypoxic ventilation to 290% of the control value, chlorpromazine to 250% control, prochlorperazine to 190% control and haloperidol to 120% control. These differences in efficacy were in the same order as the affinities of the drugs for alpha-adrenoceptors. The effect of combined haloperidol (90 nmol kg-1) and varying doses of phentolamine (175-900 nmol kg-1) was assessed to test the hypothesis that alpha-antagonism was a factor in determining the increase in ventilation following dopamine blockade. Phentolamine caused dose-dependent augmentation of the ventilatory effects of haloperidol (P less than 0.01) but had no ventilatory effect when given alone. Carotid body resection in anaesthetized mice abolished the stimulation of hypoxic ventilation caused by droperidol. It is concluded that dopaminergic mechanisms in the carotid body modulate ventilatory control in the awake mouse. The drugs most effective in augmenting hypoxic ventilation are those that block both dopamine and alpha-adrenoceptors. PMID:2862937

  7. Effects of adenosine A2A receptor stimulation on cocaine-seeking behavior in rats.

    PubMed

    Bachtell, Ryan K; Self, David W

    2009-10-01

    Dopamine (DA) receptor stimulation in the nucleus accumbens (NAc) plays an important role in regulating cocaine-seeking behavior. Adenosine receptors antagonize the effects of DA receptor stimulation on intracellular signaling, neuronal output, and behavior. The goal of the present study is to determine the effects of adenosine A(2A) receptor stimulation on reinstatement of cocaine-seeking behavior in rats. Rats were trained to lever press for cocaine in daily self-administration sessions on a fixed-ratio 1 schedule for 3 weeks. After 1 week of abstinence, lever pressing was extinguished in six daily extinction sessions. We subsequently assessed the effects of the adenosine A(2A) receptor agonist, CGS 21680, on cocaine-, quinpirole (D(2) agonist)-, and cue-induced reinstatement to cocaine seeking. We also assessed the effects of CGS 21680 on sucrose seeking in rats extinguished from sucrose self-administration. Pretreatment of CGS 21680 dose-dependently blunted cocaine-induced reinstatement (15 mg/kg, i.p.). Pretreatment with CGS 21680 (0.03 mg/kg, i.p.) also attenuated quinpirole- and cue-induced reinstatement. A minimally effective dose of CGS 21680 failed to alter cocaine-induced locomotor activity or sucrose seeking. Stimulation of adenosine A(2A) receptors antagonizes reinstatement of cocaine seeking elicited by cocaine, DA D(2)-receptor stimulation, and cocaine-conditioned cues. These findings suggest that adenosine A(2A) receptor stimulation may oppose DA D(2) receptor signaling in the NAc that mediates cocaine relapse.

  8. Novel chimeric thyroid-stimulating hormone-receptor bioassay for thyroid-stimulating immunoglobulins

    PubMed Central

    Lytton, S D; Li, Y; Olivo, P D; Kohn, L D; Kahaly, G J

    2010-01-01

    Thyroid-stimulating immunoglobulins (TSI) are a functional biomarker of Graves' disease (GD). To develop a novel TSI bioassay, a cell line (MC4-CHO-Luc) was bio-engineered to constitutively express a chimeric TSH receptor (TSHR) and constructed with a cyclic adenosine monophosphate (cAMP)-dependent luciferase reporter gene that enables TSI quantification. Data presented as percentage of specimen-to-reference ratio (SRR%) were obtained from 271 patients with various autoimmune and thyroid diseases and 180 controls. Sensitivity of 96% and specificity of 99% for untreated GD were attained by receiver operating characteristic analysis, area under the curve 0·989, 95% confidence interval 0·969–0·999, P = 0·0001. Precision testing of manufactured reagents of high, medium, low and negative SRR% gave a percentage of coefficient-of-variation of 11·5%, 12·8%, 14·5% and 15·7%, respectively. There was no observed interference by haemoglobin, lipids and bilirubin and no non-specific stimulation by various hormones at and above physiological concentrations. TSI levels from GD patients without (SRR% 406 ± 134, mean ± standard deviation) or under anti-thyroid treatment (173 ± 147) were higher (P < 0·0001) compared with TSI levels of patients with Hashimoto's thyroiditis (51 ± 37), autoimmune diseases without GD (24 ± 10), thyroid nodules (30 ± 26) and controls (35 ± 18). The bioassay showed greater sensitivity when compared with anti-TSHR binding assays. In conclusion, the TSI-Mc4 bioassay measures the functional biomarker accurately in GD with a standardized protocol and could improve substantially the diagnosis of autoimmune diseases involving TSHR autoantibodies. PMID:21070207

  9. Epidermal growth factor receptor-dependent stimulation of amphiregulin expression in androgen-stimulated human prostate cancer cells.

    PubMed Central

    Sehgal, I; Bailey, J; Hitzemann, K; Pittelkow, M R; Maihle, N J

    1994-01-01

    Amphiregulin is a heparin-binding epidermal growth factor (EGF)-related peptide that binds to the EGF receptor (EGF-R) with high affinity. In this study, we report a role for amphiregulin in androgen-stimulated regulation of prostate cancer cell growth. Androgen is known to enhance EGF-R expression in the androgen-sensitive LNCaP human prostate carcinoma cell line, and it has been suggested that androgenic stimuli may regulate proliferation, in part, through autocrine mechanisms involving the EGF-R. In this study, we demonstrate that LNCaP cells express amphiregulin mRNA and peptide and that this expression is elevated by androgenic stimulation. We also show that ligand-dependent EGF-R stimulation induces amphiregulin expression and that androgenic effects on amphiregulin synthesis are mediated through this EGF-R pathway. Parallel studies using the estrogen-responsive breast carcinoma cell line, MCF-7, suggest that regulation of amphiregulin by estrogen may also be mediated via an EGF-R pathway. In addition, heparin treatment of LNCaP cells inhibits androgen-stimulated cell growth further suggesting that amphiregulin can mediate androgen-stimulated LNCaP proliferation. Together, these results implicate an androgen-regulated autocrine loop composed of amphiregulin and its receptor in prostate cancer cell growth and suggest that the mechanism of steroid hormone regulation of amphiregulin synthesis may occur through androgen upregulation of the EGF-R and subsequent receptor-dependent pathways. Images PMID:8049525

  10. Heparin stimulates epidermal growth factor receptor-mediated phosphorylation of tyrosine and threonine residues.

    PubMed

    Revis-Gupta, S; Abdel-Ghany, M; Koland, J; Racker, E

    1991-07-15

    We have described previously that in extracts of A431 cells epidermal growth factor (EGF) stimulates the phosphorylation of tyrosine as well as of threonine residues in the EGF receptor and in lipocortin 1. We now report that heparin at low concentrations also stimulates the autophosphorylation of the EGF receptor and of the recombinant 56-kDa domain of the EGF receptor that lacks the EGF binding site. To study the stimulations of phosphorylation of threonine residues, a fusion protein was prepared with glutathione S-transferase (GST) and an EGF receptor fragment, TK8 (residues 647-688), that contains the threonine phosphorylation site but no tyrosine. We show that the phosphorylation of threonine residues in GST-TK8 by extracts of A431 cells is stimulated by heparin but not by EGF. These and other results suggest that heparin acts as a chaperone, a substrate modulator, that enhances the susceptibility of the substrate to phosphorylation by protein kinases.

  11. The human thyrotropin receptor: a heptahelical receptor capable of stimulating members of all four G protein families.

    PubMed Central

    Laugwitz, K L; Allgeier, A; Offermanns, S; Spicher, K; Van Sande, J; Dumont, J E; Schultz, G

    1996-01-01

    Thyrotropin is the primary hormone that, via one heptahelical receptor, regulates thyroid cell functions such as secretion, specific gene expression, and growth. In human thyroid, thyrotropin receptor activation leads to stimulation of the adenylyl cyclase and phospholipase C cascades. However, the G proteins involved in thyrotropin receptor action have been only partially defined. In membranes of human thyroid gland, we immunologically identified alpha subunits of the G proteins Gs short, Gs long, Gi1, Gi2, Gi3, G(o) (Go2 and another form of Go, presumably Go1), Gq, G11, G12, and G13. Activation of the thyrotropin (TSH) receptor by bovine TSH led to increased incorporation of the photoreactive GTP analogue [alpha-32P]GTP azidoanilide into immunoprecipitated alpha subunits of all G proteins detected in thyroid membranes. This effect was receptor-dependent and not due to direct G protein stimulation because it was mimicked by TSH receptor-stimulating antibodies of patients suffering from Grave disease and was abolished by a receptor-blocking antiserum from a patient with autoimmune hypothyroidism. The TSH-induced activation of individual G proteins occurred with EC50 values of 5-50 milliunits/ml, indicating that the activated TSH receptor coupled with similar potency to different G proteins. When human thyroid slices were pretreated with pertussis toxin, the TSH receptor-mediated accumulation of cAMP increased by approximately 35% with TSH at 1 milliunits/ml, indicating that the TSH receptor coupled to Gs and G(i). Taken together, these findings show that, at least in human thyroid membranes, in which the protein is expressed at its physiological levels, the TSH receptor resembles a naturally occurring example of a general G protein-activating receptor. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8552586

  12. Histamine H1 and endothelin ETB receptors mediate phospholipase D stimulation in rat brain hippocampal slices.

    PubMed

    Sarri, E; Picatoste, F; Claro, E

    1995-08-01

    Different neurotransmitter receptor agonists [carbachol, serotonin, noradrenaline, histamine, endothelin-1, and trans-(1S,3R)-aminocyclopentyl-1,3-dicarboxylic acid (trans-ACPD)], known as stimuli of phospholipase C in brain tissue, were tested for phospholipase D stimulation in [32P]Pi-prelabeled rat brain cortical and hippocampal slices. The accumulation of [32P]phosphatidylethanol was measured as an index of phospholipase D-catalyzed transphosphatidylation in the presence of ethanol. Among the six neurotransmitter receptor agonists tested, only noradrenaline, histamine, endothelin-1, and trans-ACPD stimulated phospholipase D in hippocampus and cortex, an effect that was strictly dependent of the presence of millimolar extracellular calcium concentrations. The effect of histamine (EC50 18 microM) was inhibited by the H1 receptor antagonist mepyramine with a Ki constant of 0.7 nM and was resistant to H2 and H3 receptor antagonists (ranitidine and tioperamide, respectively). Endothelin-1-stimulated phospholipase D (EC50 44 nM) was not blocked by BQ-123, a specific antagonist of the ETA receptor. Endothelin-3 and the specific ETB receptor agonist safarotoxin 6c were also able to stimulate phospholipase D with efficacies similar to that of endothelin-1, and EC50 values of 16 and 3 nM, respectively. These results show that histamine and endothelin-1 stimulate phospholipase D in rat brain through H1 and ETB receptors, respectively.

  13. Mechanism of kinase activation in the receptor for colony-stimulating factor 1.

    PubMed Central

    Lee, A W; Nienhuis, A W

    1990-01-01

    Receptor tyrosine kinases remain dormant until activated by ligand binding to the extracellular domain. Two mechanisms have been proposed for kinase activation: (i) ligand binding to the external domain of a receptor monomer may induce a conformational change that is transmitted across the cell membrane (intramolecular model) or (ii) the ligand may facilitate oligomerization, thereby allowing interactions between the juxtaposed kinase domains (intermolecular model). The receptor for colony-stimulating factor 1 was used to test these models. Large insertions at the junction between the external and transmembrane domains of the receptor, introduced by site-directed mutagenesis of the cDNA, were positioned to isolate the external domain and prevent transmembrane conformational propagation while allowing for receptor oligomerization. Such mutant receptors were expressed on the cell surface, bound ligand with high affinity, exhibited ligand-stimulated autophosphorylation, and signaled mitogenesis and cellular proliferation in the presence of ligand. A second experimental strategy directly tested the intermolecular model of ligand activation. A hybrid receptor composed of the external domain of human glycophorin A and the transmembrane and cytoplasmic domains of the colony-stimulating factor 1 receptor exhibited anti-glycophorin antibody-induced kinase activity that supported mitogenesis. Our data strongly support a mechanism of receptor activation based on ligand-induced receptor oligomerization. Images PMID:2169623

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

    SciTech Connect

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

    1988-04-01

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

  15. Stimulation of the extracellular Ca²⁺-sensing receptor by denatonium.

    PubMed

    Rogachevskaja, Olga A; Churbanov, Gleb D; Bystrova, Marina F; Romanov, Roman A; Kolesnikov, Stanislav S

    2011-12-16

    The extracellular Ca(2+)-sensing receptor (CASR) is a promiscuous G-protein-coupled receptor closely related to the taste receptors T1R1-T1R3. Here we analyzed the possibility that apart from being stimulated by external Ca(2+) and amino acids, the substances effective as tastants, CASR might serve as a receptor for other sapid compounds. CASR was heterologously expressed in HEK-293 cells, and their responsivity to a variety of bitter and sweet substances was examined. Among them, solely denatonium was found to stimulate Ca(2+) signaling in CASR-positive HEK-293 cells. Apparently, these Ca(2+) responses were specific, as those were inhibited by the CASR antagonist NSP-4123. Altogether, our findings indicate that denatonium stimulates CASR by shifting a dose-response curve for the principal CASR agonist Ca(2+) to lower concentrations. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. IGF-II receptors and IGF-II-stimulated glucose transport in human fat cells

    SciTech Connect

    Sinha, M.K.; Buchanan, C.; Raineri-Maldonado, C.; Khazanie, P.; Atkinson, S.; DiMarchi, R.; Caro, J.F. )

    1990-03-01

    Insulin-like growth factor II (IGF-II) receptors have been described in rat but not in human adipocytes. In both species, IGF-II has been reported to stimulate glucose transport by interacting with the insulin receptor. In this study, we have unequivocally demonstrated the presence of IGF-II receptors in human adipocytes. 125I-labeled IGF-II specifically binds to intact adipocytes, membranes, and lectin-purified detergent solubilized extracts. Through the use of 0.5 mM disuccinimidyl suberate, 125I-IGF-II is cross-linked to a 260-kDa protein that is identified as the IGF-II receptor by displacement experiments with unlabeled IGF-II, IGF-I, and insulin and either by immunoprecipitation or by Western blot analysis with mannose 6-phosphate receptor antibodies. The concentrations of IGF-II required for half-maximal and maximal stimulation of glucose transport in human adipocytes are 35 and 100 times more than that of insulin. The possibility of IGF-II stimulating glucose transport by interacting predominantly with the insulin receptor is suggested by the following: (1) the concentration of IGF-II that inhibits half of insulin binding is only 20 times more than that of insulin; (2) the lack of an additive effect of IGF-II and insulin for maximal stimulation of glucose transport; (3) the ability of monoclonal insulin receptor antibodies to decrease glucose transport stimulated by submaximal concentrations of both IGF-II and insulin; and (4) the ability of IGF-II to stimulate insulin receptor autophosphorylation albeit at a reduced potency when compared with insulin.

  17. Metformin (Glucophage) inhibits tyrosine phosphatase activity to stimulate the insulin receptor tyrosine kinase.

    PubMed

    Holland, William; Morrison, Thomas; Chang, Ying; Wiernsperger, Nicholas; Stith, Bradley J

    2004-06-01

    Metformin is a commonly used anti-diabetic but whether its mechanism involves action on the insulin receptor or on downstream events is still controversial. With a time course that was slow compared with insulin action, metformin increased tyrosine phosphorylation of the regulatory domain of the insulin receptor (specifically, tyrosine residues 1150 and 1151). In a direct action, therapeutic levels of metformin stimulated the tyrosine kinase activity of the soluble intracellular portion of the beta subunit of the human insulin receptor toward a substrate derived from the insulin receptor regulatory domain. However, metformin did not alter the order of substrate phosphorylation by the insulin receptor kinase. Using a Xenopus oocyte preparation, we simultaneously recorded tyrosine kinase and phosphatase activities that regulate the insulin receptor by measuring the tyrosine phosphorylation and dephosphorylation of peptides derived from the regulatory domain of the human insulin receptor. In an indirect stimulation of the insulin receptor, metformin inhibited endogenous tyrosine phosphatases and purified human protein tyrosine phosphatase 1B that dephosphorylate and inhibit the insulin receptor kinase. Thus, there was evidence that metformin acted directly upon the insulin receptor and indirectly through inhibition of tyrosine phosphatases.

  18. Niacin stimulates adiponectin secretion through the GPR109A receptor.

    PubMed

    Plaisance, Eric P; Lukasova, Martina; Offermanns, Stefan; Zhang, Youyan; Cao, Guoqing; Judd, Robert L

    2009-03-01

    Niacin (nicotinic acid) has recently been shown to increase serum adiponectin concentrations in men with the metabolic syndrome. However, little is known about the mechanism(s) by which niacin regulates the intracellular trafficking and secretion of adiponectin. Since niacin appears to exert its effects on lipolysis through receptor (GPR109A)-dependent and -independent pathways, the purpose of this investigation was to examine the role of the recently identified GPR109A receptor in adiponectin secretion. Initial in vivo studies in rats demonstrated that niacin (30 mg/kg po) acutely increases serum adiponectin concentrations, whereas it decreases NEFAs. Further in vitro studies demonstrated an increase in adiponectin secretion and a decrease in lipolysis in primary adipocytes following treatment with niacin or beta-hydroxybutyrate (an endogenous ligand of the GPR109A receptor), but these effects were blocked when adipocytes were pretreated with pertussis toxin. Niacin had no effect on adiponectin secretion or lipolysis in 3T3-L1 adipocytes, which have limited cell surface expression of the GPR109A receptor. To further substantiate these in vitro findings, wild-type and GPR109A receptor knockout mice were administered a single dose of niacin or placebo, and serum was obtained for the determination of adiponectin and NEFA concentrations. Serum adiponectin concentrations increased and serum NEFAs decreased in the wild-type mice within 10 min following niacin administration. However, niacin administration had no effect on adiponectin and NEFA concentrations in the GPR109A receptor knockout mice. These results demonstrate that the GPR109A receptor plays an important role in the dual regulation of adiponectin secretion and lipolysis.

  19. Stimulation and inhibition of adenylyl cyclase by distinct 5-hydroxytryptamine receptors.

    PubMed

    De Vivo, M; Maayani, S

    1990-10-01

    5-Hydroxytryptamine (serotonin, 5-HT) stimulates basal adenylyl cyclase activity in membranes from guinea pig or rat hippocampi, but 5-HT inhibits forskolin-stimulated adenylyl cyclase activity in these same membranes. The opposing effects of 5-HT on adenylyl cyclase activity indicate that distinct 5-HT receptors, positively and negatively coupled to adenylyl cyclase, are present in these membranes. Stimulation of adenylyl cyclase activity is mediated by two distinct 5-HT receptors. The receptor with lower affinity for 5-HT, designated as RL, is apparently homologous with a 5-HT receptor present in rat collicular membranes, but it is not homologous with the stimulatory receptor characterized in neuroblastoma hybrid cell (NCB-20) membranes. The receptor with higher affinity for 5-HT is homologous with the 5-HT1A binding site. The magnitude of stimulation by 5-HT1A receptors is variable with respect to stimulation by RL and is sometimes completely absent. Inhibition of forskolin-stimulated adenylyl cyclase activity, in membranes from either rat or guinea pig hippocampus or rat cortex, is a functional correlate of the 5-HT1A binding site. This inhibitory response was used to determine the pharmacological characteristics of drugs that reportedly have high affinity for 5-HT1A binding sites, such as 1-[2-(4-aminophenyl)ethyl]-4-(3-trifluoromethylphenyl)piperazine (PAPP) and (-)pindolol. PAPP inhibited adenylyl cyclase activity in guinea pig hippocampal membranes with an EC50 value of 27 +/- 3 nM. (-)Pindolol was a partial agonist in inhibiting adenylyl cyclase activity in guinea pig and rat hippocampal membranes. Because of the low intrinsic activity of (-)pindolol, it was tested as an antagonist of the inhibition produced by 5-HT1A receptor agonists in rat hippocampal membranes. The Kb of (-)pindolol was 40 nM as measured by a Schild plot. (-)Propranolol was a simple competitive antagonist at the rat hippocampal receptor with a Kb value of 550 nM. In summary, guinea pig

  20. Dopamine D2High receptors stimulated by phencyclidines, lysergic acid diethylamide, salvinorin A, and modafinil.

    PubMed

    Seeman, Philip; Guan, Hong-Chang; Hirbec, Hélène

    2009-08-01

    Although it is commonly stated that phencyclidine is an antagonist at ionotropic glutamate receptors, there has been little measure of its potency on other receptors in brain tissue. Although we previously reported that phencyclidine stimulated cloned-dopamine D2Long and D2Short receptors, others reported that phencyclidine did not stimulate D2 receptors in homogenates of rat brain striatum. This study, therefore, examined whether phencyclidine and other hallucinogens and psychostimulants could stimulate the incorporation of [(35)S]GTP-gamma-S into D2 receptors in homogenates of rat brain striatum, using the same conditions as previously used to study the cloned D2 receptors. Using 10 microM dopamine to define 100% stimulation, phencyclidine elicited a maximum incorporation of 46% in rat striata, with a half-maximum concentration of 70 nM for phencyclidine, when compared with 80 nM for dopamine, 89 nM for salvinorin A (48 nM for D2Long), 105 nM for lysergic acid diethylamide (LSD), 120 nM for R-modafinil, 710 nM for dizocilpine, 1030 nM for ketamine, and >10,000 nM for S-modafinil. These compounds also inhibited the binding of the D2-selective ligand [(3)H]domperidone. The incorporation was inhibited by the presence of 200 microM guanylylimidodiphosphate and also by D2 blockade, using 10 microM S-sulpiride, but not by D1 blockade with 10 microM SCH23390. Hypertonic buffer containing 150 mM NaCl inhibited the stimulation by phencyclidine, which may explain negative results by others. It is concluded that phencyclidine and other psychostimulants and hallucinogens can stimulate dopamine D2 receptors at concentrations related to their behavioral actions.

  1. Dissociation and trafficking of rat GABAB receptor heterodimer upon chronic capsaicin stimulation.

    PubMed

    Laffray, Sophie; Tan, Kelly; Dulluc, Josette; Bouali-Benazzouz, Rabia; Calver, Andrew R; Nagy, Frédéric; Landry, Marc

    2007-03-01

    Gamma-aminobutyric acid type B receptors (GABAB) are G-protein-coupled receptors that mediate GABAergic inhibition in the brain. Their functional expression is dependent upon the formation of heterodimers between GABAB1 and GABAB2 subunits, a process that occurs within the endoplasmic reticulum. However, the mechanisms that regulate GABAB receptor oligomerization at the plasma membrane remain largely unknown. We first characterized the functional cytoarchitecture of an organotypic co-culture model of rat dorsal root ganglia and spinal cord. Subsequently, we studied the interactions between GABAB subunits after chronic stimulation of sensory fibres with capsaicin. Surface labelling of recombinant proteins showed a decrease in subunit co-localization and GABAB2 labelling, after capsaicin treatment. In these conditions, fluorescence lifetime imaging measurements further demonstrated a loss of interactions between green fluorescent protein-GABAB1b and t-dimer discosoma sp red fluorescent protein-GABAB2 subunits. Finally, we established that the GABAB receptor undergoes clathrin-dependent internalization and rapid recycling to the plasma membrane following activation with baclofen, a GABAB agonist. However, in cultures chronically stimulated with capsaicin, the agonist-induced endocytosis was decreased, reflecting changes in the dimeric state of the receptor. Taken together, our results indicate that the chronic stimulation of sensory fibres can dissociate the GABAB heterodimer and alters its responsiveness to the endogenous ligand. Chronic stimulation thus modulates receptor oligomerization, providing additional levels of control of signalling.

  2. Role of the Extracellular and Intracellular Loops of Follicle-Stimulating Hormone Receptor in Its Function

    PubMed Central

    Banerjee, Antara A.; Mahale, Smita D.

    2015-01-01

    Follicle-stimulating hormone receptor (FSHR) is a leucine-rich repeat containing class A G-protein coupled receptor belonging to the subfamily of glycoprotein hormone receptors (GPHRs), which includes luteinizing hormone/choriogonadotropin receptor (LH/CGR) and thyroid-stimulating hormone receptor. Its cognate ligand, follicle-stimulating hormone binds to, and activates FSHR expressed on the surface of granulosa cells of the ovary, in females, and Sertoli cells of the testis, in males, to bring about folliculogenesis and spermatogenesis, respectively. FSHR contains a large extracellular domain (ECD) consisting of leucine-rich repeats at the N-terminal end and a hinge region at the C-terminus that connects the ECD to the membrane spanning transmembrane domain (TMD). The TMD consists of seven α-helices that are connected to each other by means of three extracellular loops (ELs) and three intracellular loops (ILs) and ends in a short-cytoplasmic tail. It is well established that the ECD is the primary hormone binding domain, whereas the TMD is the signal transducing domain. However, several studies on the ELs and ILs employing site directed mutagenesis, generation of chimeric receptors and in vitro characterization of naturally occurring mutations have proven their indispensable role in FSHR function. Their role in every phase of the life cycle of the receptor like post translational modifications, cell surface trafficking, hormone binding, activation of downstream signaling, receptor phosphorylation, hormone–receptor internalization, and recycling of hormone–receptor complex have been documented. Mutations in the loops causing dysregulation of these processes lead to pathophysiological conditions. In other GPHRs as well, the loops have been convincingly shown to contribute to various aspects of receptor function. This review article attempts to summarize the extensive contributions of FSHR loops and C-terminal tail to its function. PMID:26236283

  3. Novel Insights on Thyroid-Stimulating Hormone Receptor Signal Transduction

    PubMed Central

    Neumann, Susanne; Grüters, Annette; Krude, Heiko

    2013-01-01

    The TSH receptor (TSHR) is a member of the glycoprotein hormone receptors, a subfamily of family A G protein-coupled receptors. The TSHR is of great importance for the growth and function of the thyroid gland. The TSHR and its endogenous ligand TSH are pivotal proteins with respect to a variety of physiological functions and malfunctions. The molecular events of TSHR regulation can be summarized as a process of signal transduction, including signal reception, conversion, and amplification. The steps during signal transduction from the extra- to the intracellular sites of the cell are not yet comprehensively understood. However, essential new insights have been achieved in recent years on the interrelated mechanisms at the extracellular region, the transmembrane domain, and intracellular components. This review contains a critical summary of available knowledge of the molecular mechanisms of signal transduction at the TSHR, for example, the key amino acids involved in hormone binding or in the structural conformational changes that lead to G protein activation or signaling regulation. Aspects of TSHR oligomerization, signaling promiscuity, signaling selectivity, phenotypes of genetic variations, and potential extrathyroidal receptor activity are also considered, because these are relevant to an understanding of the overall function of the TSHR, including physiological, pathophysiological, and pharmacological perspectives. Directions for future research are discussed. PMID:23645907

  4. Slowing of shortening velocity of rat cardiac myocytes by adenosine receptor stimulation regardless of beta-adrenergic stimulation.

    PubMed Central

    Strang, K T; Mentzer, R M; Moss, R L

    1995-01-01

    1. Single ventricular myocytes were enzymatically isolated, incubated with the A1-purinergic and beta-adrenergic receptor-specific agonists N6-cyclopentyladenosine (CPA) and isoprenaline (Iso), and then rapidly skinned. Ca2+ sensitivity of isometric tension and unloaded shortening velocity (Vo) were measured, and protein kinase A (PKA)-specific phosphorylations of troponin I (TnI) and C-protein were assessed by back-phosphorylation of cell suspensions with [gamma-32P]-ATP. 2. Isoprenaline treatment decreased the Ca2+ sensitivity of isometric tension relative to propranolol-treated controls, as did simultaneous stimulation with Iso and CPA (Iso + CPA). CPA alone had no effect on Ca2+ sensitivity. Vo was greater in Iso-treated cells than in paired controls, while Vo was significantly less than control in both Iso + CPA-treated and CPA-treated cells. 3. Phosphorylation of TnI and C-protein was increased by Iso treatment and also when Iso and CPA were simultaneously applied. CPA alone caused a significant decrease in the phosphorylation state of these two proteins. 4. From these results we conclude that A1-purinergic receptor stimulation does not inhibit beta-adrenergic receptor-mediated phosphorylation of myofilament proteins, nor does it alter the Ca2+ sensitivity of isometric tension at the level of the myofilaments. However, A1-receptor stimulation does decrease Vo at the level of the myofilaments by a mechanism that is independent of beta-adrenergically mediated phosphorylation of TnI and C-protein. Images Figure 1 Figure 4 PMID:7473228

  5. Discovery of substituted benzamides as follicle stimulating hormone receptor allosteric modulators.

    PubMed

    Yu, Henry N; Richardson, Thomas E; Nataraja, Selva; Fischer, David J; Sriraman, Venkataraman; Jiang, Xuliang; Bharathi, Pandi; Foglesong, Robert J; Haxell, Thomas F N; Heasley, Brian H; Jenks, Mathew; Li, Jane; Dugas, Melanie S; Collis, Regina; Tian, Hui; Palmer, Stephen; Goutopoulos, Andreas

    2014-05-01

    Follicle-stimulating hormone (FSH), acting on its receptor (FSHR), plays a pivotal role in the stimulation of follicular development and maturation. Multiple injections of protein formulations are used during clinical protocols for ovulation induction and for in vitro fertilization that are followed by a selection of assisted reproductive technologies. In order to increase patient convenience and compliance several research groups have searched for orally bioavailable FSH mimetics for innovative fertility medicines. We report here the discovery of a series of substituted benzamides as positive allosteric modulators (PAM) targeting FSHR. Optimization of this series has led to enhanced activity in primary rat granulosa cells, as well as remarkable selectivity against the closely related luteinizing hormone receptor (LHR) and thyroid stimulating hormone receptor (TSHR). Two modulators, 9j and 9k, showed promising in vitro and pharmacokinetic profiles.

  6. Prevention of Stimulant Induced Euphoria with an Opioid Receptor Antagonist

    DTIC Science & Technology

    2013-10-01

    ADHD symptoms over the course of the 6-week trial 15. SUBJECT TERMS- Clinical trial, methylphenidate, naltrexone, ADHD stimulant induced euphoria...In this double-blind study, subjects will receive methylphenidate and naltrexone or a placebo to treat their ADHD symptoms over the course of the 6...The Study Coordinator and Research Assistant have been trained to administer an IRB- approved phone screen asking about past and current ADHD symptoms

  7. GABAB receptor activation attenuates the stimulant but not mesolimbic dopamine response to ethanol in FAST mice

    PubMed Central

    Holstein, Sarah E.; Li, Na; Eshleman, Amy J.; Phillips, Tamara J.

    2012-01-01

    Neural processes influenced by γ-aminobutyric acid B (GABAB) receptors appear to contribute to acute ethanol sensitivity, including the difference between lines of mice bred for extreme sensitivity (FAST) or insensitivity (SLOW) to the locomotor stimulant effect of ethanol. One goal of the current study was to determine whether selection of the FAST and SLOW lines resulted in changes in GABAB receptor function, since the lines differ in sensitivity to the GABAB receptor agonist baclofen and baclofen attenuates the stimulant response to ethanol in FAST mice. A second goal was to determine whether the baclofen-induced reduction in ethanol stimulation in FAST mice is associated with an attenuation of the mesolimbic dopamine response to ethanol. In Experiment 1, the FAST and SLOW lines were found to not differ in GABAB receptor function (measured by baclofen-stimulated [35S]GTP!S binding) in whole brain or in several regional preparations, except in the striatum in one of the two replicate sets of selected lines. In Experiment 2, baclofen-induced attenuation of the locomotor stimulant response to ethanol in FAST mice was not accompanied by a reduction in dopamine levels in the nucleus accumbens, as measured by microdialysis. These data suggest that, overall, GABAB receptor function does not play an integral role in the genetic difference in ethanol sensitivity between the FAST and SLOW lines. Further, although GABAB receptors do modulate the locomotor stimulant response to ethanol in FAST mice, this effect does not appear to be due to a reduction in tonic dopamine signaling in the nucleus accumbens. PMID:22982185

  8. Palmitoylethanolamide enhances anandamide stimulation of human vanilloid VR1 receptors.

    PubMed

    De Petrocellis, L; Davis, J B; Di Marzo, V

    2001-10-12

    In human embryonic kidney cells over-expressing the human vanilloid receptor type 1 (VR1), palmitoylethanolamide (PEA, 0.5-10 microM) enhanced the effect of arachidonoylethanolamide (AEA, 50 nM) on the VR1-mediated increase of the intracellular Ca2+ concentration. PEA (5 microM) decreased the AEA half-maximal concentration for this effect from 0.44 to 0.22 microM. The PEA effect was not due to inhibition of AEA hydrolysis or adhesion to non-specific sites, since bovine serum albumin (0.01-0.25%) potently inhibited AEA activity, and PEA also enhanced the effect of low concentrations of the VR1 agonists resiniferatoxin and capsaicin. PEA (5 microM) enhanced the affinity of AEA for VR1 receptors as assessed in specific binding assays. These data suggest that PEA might be an endogenous enhancer of VR1-mediated AEA actions.

  9. Adenosine transiently modulates stimulated dopamine release in the caudate putamen via A1 receptors

    PubMed Central

    Ross, Ashley E.; Venton, B. Jill

    2014-01-01

    Adenosine modulates dopamine in the brain via A1 and A2A receptors, but that modulation has only been characterized on a slow time scale. Recent studies have characterized a rapid signaling mode of adenosine that suggests a possible rapid modulatory role. Here, fast-scan cyclic voltammetry was used to characterize the extent to which transient adenosine changes modulate stimulated dopamine release (5 pulses at 60 Hz) in rat caudate putamen brain slices. Exogenous adenosine was applied and dopamine concentration monitored. Adenosine only modulated dopamine when it was applied 2 or 5 s before stimulation. Longer time intervals and bath application of 5 µM adenosine did not decrease dopamine release. Mechanical stimulation of endogenous adenosine 2s before dopamine stimulation also decreased stimulated dopamine release by 41 ± 7 %, similar to the 54 ± 6 % decrease in dopamine after exogenous adenosine application. Dopamine inhibition by transient adenosine was recovered within 10 minutes. The A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) blocked the dopamine modulation, whereas dopamine modulation was unaffected by the A2A receptor antagonist SCH 442416. Thus, transient adenosine changes can transiently modulate phasic dopamine release via A1 receptors. These data demonstrate that adenosine has a rapid, but transient, modulatory role in the brain. PMID:25219576

  10. Mechanical Stimulation of Piezo1 Receptors Depends on Extracellular Matrix Proteins and Directionality of Force.

    PubMed

    Gaub, Benjamin M; Müller, Daniel J

    2017-02-08

    Piezo receptors convert mechanical forces into electrical signals. In mammals, they play important roles in basic physiological functions including proprioception, sensation of touch, and vascular development. However, basic receptor properties like the gating mechanism, the interaction with extracellular matrix (ECM) proteins, and the response to mechanical stimulation, remain poorly understood. Here, we establish an atomic force microscopy (AFM)-based assay to mechanically stimulate Piezo1 receptors in living animal cells, while monitoring receptor activation in real-time using functional calcium imaging. Our experiments show that in the absence of ECM proteins Piezo1 receptors are relatively insensitive to mechanical forces pushing the cellular membrane, whereas they can hardly be activated by mechanically pulling the membrane. Yet, if conjugated with Matrigel, a mix of ECM proteins, the receptors become sensitized. Thereby, forces pulling the cellular membrane activate the receptor much more efficiently compared to pushing forces. Finally, we found that collagen IV, a component of the basal lamina, which forms a cohesive network and mechanical connection between cells, sensitizes Piezo1 receptors to mechanical pulling.

  11. Dissociation of insulin receptor phosphorylation and stimulation of glucose transport in BC3H-1 myocytes

    SciTech Connect

    Mojsilovic, L.P.; Standaert, M.L.; Rosic, N.K.; Pollet, R.J.

    1986-05-01

    The authors have investigated insulin receptor phosphorylation in differentiated cultured BC3H-1 myocytes. As for other insulin-responsive cell systems in partially purified wheat germ agglutinin receptor preparations, insulin stimulates the phosphorylation of its own receptor (95K ..beta..-subunits) in a dose dependent manner (0-400 nM), as identified by immunoprecipitation with antiinsulin receptor antibodies and SDS-PAGE. In the same preparations they show that 12-0-tetradecanyl phorbol acetate (TPA), which in many respect ..beta..-subunits in the same dose dependent manner (0-5 ..mu..M). In addition, antiinsulin receptor antibodies (B-10) also induced phosphorylation of mimics insulin action, also induced phosphorylation of the insulin receptor and HPLC tryptic maps of the /sup 32/P-labeled ..beta..-subunit were identical to those for insulin-induced receptor phosphorylation. However, while insulin and TPA are potent stimulators of glucose transport in these muscle cells, the antireceptor antibodies alone failed to provoke glucose transport at any concentration. The specificity and activity of these antibodies were confirmed in their system by their ability to inhibit insulin binding and insulin-stimulated glucose transport in a concentration-dependent manner. Their results indicate that phosphorylation of insulin receptor is not a crucial event in mediating insulin action, at least with respect to glucose transport. While the effects of the B-10 antibody in the BC3H-1 myocyte differ from those in the adipocyte, their results provide independent confirmation of their essential conclusion that phosphorylation of the insulin receptor may not be necessary nor sufficient for its acute action in promoting glucose transport.

  12. Histamine receptors on adult rat cardiomyocytes: antagonism of alpha/sub 1/-receptor stimulation of cAMP degradation

    SciTech Connect

    Buxton, I.L.O.; Bowen, S.M.

    1986-03-01

    Incubation of intact cardiomyocytes with the histamine antagonist (/sup 3/H)mepyramine results in rapid reversible binding to a single class of high affinity sites (K/sub D/ = 1.2nM; 50,000 sites/myocyte). In membranes from purified myocytes histamine competition of (/sup 3/H)mepyramine binding (K/sub D/ = 300nM) is not altered by GTP (10..mu..M). Competition of (/sup 3/H)mepyramine binding by H-receptor subtype-selective antagonists suggests the presence of a single class of H/sub 1/-receptors. Incubation of intact myocytes with histamine (luM, H/sub 1/ receptor activation) plus norepinephrine (NE 1uM, alpha/sub 1/ + beta/sub 1/ receptor activation) for 3 min leads to significantly more cAMP accumulation (36.5 pmol/10/sup 6/ myocytes) than NE alone (30 pmol/10/sup 6/ myocytes). Histamine alone does not alter basal cAMP = 10.4 pmol/10/sup 6/ myocytes, or beta/sub 1/ stimulation (isoproternol, 1uM) = 39.6 pmol/10/sup 6/ myocytes. Cyclic AMP accumulation with NE plus prazosin 10nM, (alpha/sub 1/ + beta/sub 1/ + alpha/sub 1/ blockade) is indistinguishable from NE + histamine, (alpha/sub 1/ + beta/sub 1/ + H/sub 1/) stimulation. Histamine competition for (/sup 3/H)prazosin binding suggests that histamine does not block alpha/sub 1/ receptors on the myocyte. These data suggest that H/sub 1/ receptor activation leads to antagonism of the alpha/sub 1/ receptor mediated activation of cAMP phosphodiesterase the authors have recently described.

  13. Adenosine Receptor Stimulation Improves Glucocorticoid-Induced Osteoporosis in a Rat Model.

    PubMed

    Pizzino, Gabriele; Irrera, Natasha; Galfo, Federica; Oteri, Giacomo; Atteritano, Marco; Pallio, Giovanni; Mannino, Federica; D'Amore, Angelica; Pellegrino, Enrica; Aliquò, Federica; Anastasi, Giuseppe P; Cutroneo, Giuseppina; Squadrito, Francesco; Altavilla, Domenica; Bitto, Alessandra

    2017-01-01

    Glucocorticoid-induced osteoporosis (GIO) is a secondary cause of bone loss. Bisphosphonates approved for GIO, might induce jaw osteonecrosis; thus additional therapeutics are required. Adenosine receptor agonists are positive regulators of bone remodeling, thus the efficacy of adenosine receptor stimulation for treating GIO was tested. In a preventive study GIO was induced in Sprague-Dawley rats by methylprednisolone (MP) for 60 days. Animals were randomly assigned to receive polydeoxyribonucleotide (PDRN), an adenosine A2 receptor agonist, or PDRN and DMPX (3,7-dimethyl-1-propargylxanthine, an A2 antagonist), or vehicle (0.9% NaCl). Another set of animals was used for a treatment study, following the 60 days of MP-induction rats were randomized to receive (for additional 60 days) PDRN, or PDRN and DMPX (an adenosine A2 receptor antagonist), or zoledronate (as control for gold standard treatment), or vehicle. Control animals were administered with vehicle for either 60 or 120 days. Femurs were analyzed after treatments for histology, imaging, and breaking strength analysis. MP treatment induced severe bone loss, the concomitant use of PDRN prevented the developing of osteoporosis. In rats treated for 120 days, PDRN restored bone architecture and bone strength; increased b-ALP, osteocalcin, osteoprotegerin and stimulated the Wnt canonical and non-canonical pathway. Zoledronate reduced bone resorption and ameliorated the histological features, without significant effects on bone formation. Our results suggest that adenosine receptor stimulation might be useful for preventing and treating GIO.

  14. Tumor necrosis factor inhibits ligand-stimulated EGF receptor activation through a TNF receptor 1-dependent mechanism

    PubMed Central

    McElroy, Steven J.; Frey, Mark R.; Yan, Fang; Edelblum, Karen L.; Goettel, Jeremy A.; John, Sutha; Polk, D. Brent

    2008-01-01

    Tumor necrosis factor (TNF) and epidermal growth factor (EGF) are key regulators in the intricate balance maintaining intestinal homeostasis. Previous work from our laboratory shows that TNF attenuates ligand-driven EGF receptor (EGFR) phosphorylation in intestinal epithelial cells. To identify the mechanisms underlying this effect, we examined EGFR phosphorylation in cells lacking individual TNF receptors. TNF attenuated EGF-stimulated EGFR phosphorylation in wild-type and TNFR2−/−, but not TNFR1−/−, mouse colon epithelial (MCE) cells. Reexpression of wild-type TNFR1 in TNFR1−/− MCE cells rescued TNF-induced EGFR inhibition, but expression of TNFR1 deletion mutant constructs lacking the death domain (DD) of TNFR1 did not, implicating this domain in EGFR downregulation. Blockade of p38 MAPK, but not MEK, activation of ERK rescued EGF-stimulated phosphorylation in the presence of TNF, consistent with the ability of TNFR1 to stimulate p38 phosphorylation. TNF promoted p38-dependent EGFR internalization in MCE cells, suggesting that desensitization is achieved by reducing receptor accessible to ligand. Taken together, these data indicate that TNF activates TNFR1 by DD- and p38-dependent mechanisms to promote EGFR internalization, with potential impact on EGF-induced proliferation and migration key processes that promote healing in inflammatory intestinal diseases. PMID:18467504

  15. Modulation of sympathetic actions on the heart by opioid receptor stimulation.

    PubMed

    Wong, T M; Shan, J

    2001-01-01

    The sympathetic nervous system, the most important extrinsic regulatory mechanism of the heart, is inhibited postsynaptically and presynaptically by opioid peptides produced in the heart via their respective receptors. The cardiac actions of beta-adrenergic receptor (beta-AR) stimulation are attenuated by activation of the opioid receptor (OR) with OR agonist at ineffective concentrations, implying cross-talk between the OR and beta-AR. This cross-talk results from inhibition of the Gs protein and adenylyl cyclase of the beta-AR pathway by the pertussis toxin-sensitive G protein of the opioid pathway. Alterations in cross-talk between these two receptors occur in pathological situations to meet bodily needs. In myocardial ischemia, when the sympathetic activity is increased, the inhibition of beta-AR stimulation by kappa-opioid stimulation is also enhanced, thus reducing the workload, oxygen consumption and cardiac injury. Whereas cardiac responsiveness to sympathetic discharges is also reduced after chronic hypoxia, the cross-talk between kappa-OR and beta-AR is reduced to prevent undue suppression of the sympathetic influence on the heart. On the other hand, impairment of the cross-talk may result in abnormality. A lack or a significant reduction in the inhibition of beta-AR stimulation by kappa-OR stimulation may lead to an excessive increase in cardiac activities, which contribute to the maintenance of high arterial blood pressure in spontaneously hypertensive rats. Other than opioid peptides, female sex hormone and adenosine also inhibit the sympathetic actions on the heart. In addition, sympathetic action is also inhibited presynaptically by kappa-opioid peptides via their receptor. Copyright 2001 National Science Council, ROC and S. Karger AG, Basel

  16. Adenosine transiently modulates stimulated dopamine release in the caudate-putamen via A1 receptors.

    PubMed

    Ross, Ashley E; Venton, B Jill

    2015-01-01

    Adenosine modulates dopamine in the brain via A1 and A2A receptors, but that modulation has only been characterized on a slow time scale. Recent studies have characterized a rapid signaling mode of adenosine that suggests a possible rapid modulatory role. Here, fast-scan cyclic voltammetry was used to characterize the extent to which transient adenosine changes modulate stimulated dopamine release (5 pulses at 60 Hz) in rat caudate-putamen brain slices. Exogenous adenosine was applied and dopamine concentration monitored. Adenosine only modulated dopamine when it was applied 2 or 5 s before stimulation. Longer time intervals and bath application of 5 μM adenosine did not decrease dopamine release. Mechanical stimulation of endogenous adenosine 2 s before dopamine stimulation also decreased stimulated dopamine release by 41 ± 7%, similar to the 54 ± 6% decrease in dopamine after exogenous adenosine application. Dopamine inhibition by transient adenosine was recovered within 10 min. The A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine blocked the dopamine modulation, whereas dopamine modulation was unaffected by the A2A receptor antagonist SCH 442416. Thus, transient adenosine changes can transiently modulate phasic dopamine release via A1 receptors. These data demonstrate that adenosine has a rapid, but transient, modulatory role in the brain. Here, transient adenosine was shown to modulate phasic dopamine release on the order of seconds by acting at the A1 receptor. However, sustained increases in adenosine did not regulate phasic dopamine release. This study demonstrates for the first time a transient, neuromodulatory function of rapid adenosine to regulate rapid neurotransmitter release.

  17. Stochastic Resonance in crayfish hydrodynamic receptors stimulated with external noise

    NASA Astrophysics Data System (ADS)

    Douglass, J. K.; Wilkens, L. A.; Pantazelou, E.; Moss, F.

    1993-08-01

    Stochastic Resonance (SR) is a statistical process occurring only in nonlinear dynamical systems whereby a subthreshold coherent stimulus or signal can be enhanced by noise. The signal alone is too weak to cause a state change of the system. State changes are the carriers of information through the system. In the presence of random noise, however, the system can change state, more-or-less randomly, but with some degree of coherence with the signal. A measure of this coherence at the output shows a maximum at an optimal value of the noise intensity as the signature of SR. SR is the object of recent and continued experimental and theoretical research in statistical physics. While SR has been demonstrated in a variety of physical systems, it has not yet been discovered in any naturally occurring system. This paper was stimulated by the idea that the sensory nervous system might be an appropriate setting for a search for naturally occurring SR. The detection of weak stimuli, often in the presence of noise, is, after all, the first business of the sensory system. Moreover, the system is evolved, which admits the possibility that the process of natural selection might have resulted in an optimization with respect to the (inevitable) noise. This paper describes an experiment designed to observe SR in the mechanoreceptor cells of the crayfish Procambarus clarkii, shown on the left in Fig. 1, using external noise plus a weak coherent signal as the stimulus.

  18. D2 receptor block abolishes θ burst stimulation-induced neuroplasticity in the human motor cortex.

    PubMed

    Monte-Silva, Katia; Ruge, Diane; Teo, James T; Paulus, Walter; Rothwell, John C; Nitsche, Michael A

    2011-09-01

    Dopamine (DA) is a neurotransmitter with an important influence on learning and memory, which is thought to be due to its modulatory effect on plasticity at central synapses, which in turn depends on activation of D1 and D2 receptors. Methods of brain stimulation (transcranial direct current stimulation, tDCS; paired associative stimulation, PAS) lead to after-effects on cortical excitability that are thought to resemble long-term potentization (LTP)/long-term depression (LTD) in reduced preparations. In a previous study we found that block of D2 receptors abolished plasticity induced by tDCS but had no effect on the facilitatory plasticity induced by PAS. We postulated that the different effect of D2 receptor block on tDCS- and PAS-induced plasticity may be due to the different focality and associativity of the stimulation techniques. However, alternative explanations for this difference could not be ruled out. tDCS also differs from PAS in other aspects, as tDCS induces plasticity by subthreshold neuronal activation, modulating spontaneous activity, whereas PAS induces plasticity via phasic suprathreshold stimulation. The present study in 12 volunteers examined effects of D2 receptor blockade (sulpiride (SULP) 400 mg), on the LTP/LTD-like effects of theta burst transcranial magnetic stimulation (TBS), which has less restricted effects on cortical synapses than that of PAS, and does not induce associative plasticity, similar to tDCS, but on the other hand induces cortical excitability shifts by suprathreshold (rhythmic) activation of cortical neurons similarly to PAS. Administration of SULP blocked both the excitatory and inhibitory effects of intermittent (iTBS) and continuous TBS (cTBS), respectively. As the reduced response to TBS following SULP resembles its effect on tDCS, the results support an effect of DA on plasticity, which might be related to the focality and associativity of the plasticity induced.

  19. Phorbol ester stimulates secretory activity while inhibiting receptor-activated aminopyrine uptake by gastric glands

    SciTech Connect

    Brown, M.R.; Chew, C.S.

    1986-03-05

    Both cyclic AMP-dependent and -independent secretagogues stimulate pepsinogen release, respiration and H/sup +/ secretory activity (AP uptake) in rabbit gastric glands. 12-O-tetradecanoylphorbol-13-acetate (T), a diacyglycerol analog, activates protein kinase C (PKC) and stimulates secretion in many systems. T stimulated respiration and pepsinogen release by glands and increased AP uptake by both glands and purified parietal cells. However, T reduced AP uptake by glands stimulated with carbachol (C) or histamine (H) with an apparent IC/sub 50/ of 1 nM. Preincubation with T for 30 min produced maximum inhibition which was not reversed by removal of T. T accelerated the decline of the transient C peak while the late steady state response to H was most inhibited. H-stimulated AP uptake was also inhibited by 50 ..mu..g/ml 1-oleoyl-2-acetyl-glycerol, a reported PKC activator, but not by the inactive phorbol, 4..cap alpha..-phorbol-12,13-didecanoate. In contrast, T potentiated AP uptake by glands stimulated with submaximal doses of dibutyryl cyclic AMP. These results suggest inhibition by T is a specific effect of PKC activators. The differing effects of T on secretion indicators may result from a dual action of T on receptor and post-receptor intracellular events.

  20. Kappa opioid receptors stimulate phosphoinositide turnover in rat brain

    SciTech Connect

    Periyasamy, S.; Hoss, W. )

    1990-01-01

    The effects of various subtype-selective opioid agonists and antagonists on the phosphoinositide (PI) turnover response were investigated in the rat brain. The {kappa}-agonists U-50,488H and ketocyclazocine produced a concentration-dependent increase in the accumulation of IP's in hippocampal slices. The other {kappa}-agonists Dynorphin-A (1-13) amide, and its protected analog D(Ala){sup 2}-dynorphin-A (1-13) amide also produced a significant increase in the formation of ({sup 3}H)-IP's, whereas the {mu}-selective agonists (D-Ala{sup 2}-N-Me-Phe{sup 4}-Gly{sup 5}-ol)-enkephalin and morphine and the {delta}-selective agonist (D-Pen{sup 2,5})-enkephalin were ineffective. The increase in IP's formation elicited by U-50,488H was partially antagonized by naloxone and more completely antagonized by the {kappa}-selective antagonists nor-binaltorphimine and MR 2266. The formation of IP's induced by U-50,488H varies with the regions of the brain used, being highest in hippocampus and amygdala, and lowest in striatum and pons-medullar. The results indicate that brain {kappa}- but neither {mu}- nor {delta}- receptors are coupled to the PI turnover response.

  1. Pseudohypoparathyroidism, a novel mutation in the betagamma-contact region of Gsalpha impairs receptor stimulation.

    PubMed

    Farfel, Z; Iiri, T; Shapira, H; Roitman, A; Mouallem, M; Bourne, H R

    1996-08-16

    Pseudohypoparathyroidism, type Ia (PHP-Ia), is a dominantly inherited endocrine disorder characterized by resistance to hormones that act by stimulating adenylyl cyclase. It is caused by inheritance of an autosomal mutation that inactivates the alpha subunit (alphas) of Gs, the stimulatory regulator of adenylyl cyclase. In three members of a family, the PHP-Ia phenotype is associated with a mutation (R231H) that substitutes histidine for an arginine at position 231 in alphas. We assessed signaling function of alphas-WT versus alphas-R231H transiently transfected in HEK293 cells. Hormone receptor-dependent stimulation of cAMP accumulation in cells expressing alphas-R231H is reduced by approximately 75% in comparison to cAMP accumulation in cells expressing alphas-WT. A second mutation, alphas-R201C, inhibits the GTPase turnoff reaction of alphas, thus producing receptor-independent stimulation of cAMP accumulation. The double mutant, alphas-R231H/R201C, stimulates cAMP accumulation almost as well (approximately 80%) as does alphas-R201C itself, indicating that the R231H mutation selectively impairs receptor-dependent signaling. In three-dimensional structures of G protein heterotrimers, Arg-231 is located in a region, switch 2, that is thought to interact with the betagamma subunit rather than with the hormone receptor. Thus, the R231H phenotype suggests that switch 2 (perhaps in concert with betagamma) mediates G protein activation by receptors at a site distant from the receptor-G protein contact surface.

  2. Studies of the stimulation and desensitization of beta adrenergic receptors in the human lymphocyte

    SciTech Connect

    Borst, S.E.

    1985-01-01

    Lymphocytes, were employed in radioligand binding studies of beta-adrenergic receptor density and affinity for agonists and in studies of isoproterenol stimulated adenylate cyclase activity. Studies of isoproterenol stimulation of adenylate cyclase activity demonstrated a role for extracellular calcium ions but not for extracellular magnesium ions in this process. Responses were diminished by chelation or by removal of extracellular calcium, as well as by lanthanum ion which competes with calcium for membrane binding sites. Initial studies using /sup 3/H-dihydroalprenolol (/sup 3/H-DHA) indicated that 40% of cell surface beta receptors are lost during exposure of lymphocytes to isoproterenol and that the remaining receptors have a reduced affinity for beta agonists. Results from studies with /sup 125/iodocyanopindolol (/sup 125/ICYP) are consistent with the view that beta receptors lost from the cell surface during isoproterenol treatment are present in a internal compartment of the cell. In mild asthmatic patients receiving a three week regimen of oral terbutaline a 40% reduction in the total receptor population was observed, suggesting that degradation of internalized receptors had occurred.

  3. Blockade of NMDA receptors prevents analgesic tolerance to repeated transcutaneous electrical nerve stimulation (TENS) in rats

    PubMed Central

    Hingne, Priyanka M.; Sluka, Kathleen A.

    2008-01-01

    Repeated daily application transcutaneous electrical nerve stimulation (TENS) results in tolerance, at spinal opioid receptors, to the anti-hyperalgesia produced by TENS. Since N-Methyl-D-Aspartate (NMDA) receptor antagonists prevent analgesic tolerance to opioid agonists we hypothesized that blockade of NMDA receptors will prevent tolerance to TENS. In rats with knee joint inflammation, TENS was applied for 20 minute daily at high frequency (100 Hz), low frequency (4 Hz), or sham TENS. Rats were treated with the NMDA antagonist MK-801 (0.01 mg/kg-0.1 mg/kg) or vehicle daily before TENS. Paw withdrawal thresholds were tested before and after inflammation, and before and after TENS treatment for 4 days. On day 1 TENS reversed the decreased mechanical withdrawal threshold induced by joint inflammation. On day 4 TENS had no effect on the decreased withdrawal threshold in the group treated with vehicle demonstrating development of tolerance. However, in the group treated with 0.1 mg/kg MK-801, TENS significantly reversed the mechanical withdrawal thresholds on day 4 demonstrating that tolerance did not develop. Vehicle treated animals developed cross-tolerance at spinal opioid receptors. Treatment with MK-801 reversed this cross-tolerance at spinal opioid receptors. In summary, blockade of NMDA receptors prevents analgesic tolerance to daily TENS by preventing tolerance at spinal opioid receptors. Perspective Tolerance observed to the clinical treatment of TENS could be prevented by administration of pharmaceutical agents with NMDA receptors activity such as ketamine or dextromethorphan. PMID:18061543

  4. A role for sigma receptors in stimulant self-administration and addiction.

    PubMed

    Katz, Jonathan L; Hong, Weimin C; Hiranita, Takato; Su, Tsung-Ping

    2016-04-01

    Sigma-1 receptors (σ1Rs) are structurally unique intracellular proteins that function as chaperones. σ1Rs translocate from the mitochondria-associated membrane to other subcellular compartments, and can influence a host of targets, including ion channels, G-protein-coupled receptors, lipids, and other signaling proteins. Drugs binding to σRs can induce or block the actions of σRs. Studies indicate that stimulant self-administration induces the reinforcing effects of σR agonists, because of dopamine transporter actions. Once established, the reinforcing effects of σR agonists are independent of dopaminergic mechanisms traditionally thought to be critical to the reinforcing effects of stimulants. Self-administered doses of σR agonists do not increase dopamine concentrations in the nucleus accumbens shell, a transmitter and brain region considered important for the reinforcing effects of abused drugs. However, self-administration of σR agonists is blocked by σR antagonists. Several effects of stimulants have been blocked by σR antagonists, including the reinforcing effects, assessed by a place-conditioning procedure. However, the self-administration of stimulants is largely unaffected by σR antagonists, indicating fundamental differences in the mechanisms underlying these two procedures used to assess the reinforcing effects. When σR antagonists are administered in combination with dopamine uptake inhibitors, an effective and specific blockade of stimulant self-administration is obtained. Actions of stimulant drugs related to their abuse induce unique changes in σR activity and the changes induced potentially create redundant and, once established, independent reinforcement pathways. Concomitant targeting of both dopaminergic pathways and σR proteins produces a selective antagonism of stimulant self-administration, suggesting new avenues for combination chemotherapies to specifically combat stimulant abuse.

  5. Promotion of Cancer Cell Invasiveness and Metastasis Emergence Caused by Olfactory Receptor Stimulation

    PubMed Central

    Sanz, Guenhaël; Leray, Isabelle; Dewaele, Aurélie; Sobilo, Julien; Lerondel, Stéphanie; Bouet, Stéphan; Grébert, Denise; Monnerie, Régine; Pajot-Augy, Edith; Mir, Lluis M.

    2014-01-01

    Olfactory receptors (ORs) are expressed in the olfactory epithelium, where they detect odorants, but also in other tissues with additional functions. Some ORs are even overexpressed in tumor cells. In this study, we identified ORs expressed in enterochromaffin tumor cells by RT-PCR, showing that single cells can co-express several ORs. Some of the receptors identified were already reported in other tumors, but they are orphan (without known ligand), as it is the case for most of the hundreds of human ORs. Thus, genes coding for human ORs with known ligands were transfected into these cells, expressing functional heterologous ORs. The in vitro stimulation of these cells by the corresponding OR odorant agonists promoted cell invasion of collagen gels. Using LNCaP prostate cancer cells, the stimulation of the PSGR (Prostate Specific G protein-coupled Receptor), an endogenously overexpressed OR, by β-ionone, its odorant agonist, resulted in the same phenotypic change. We also showed the involvement of a PI3 kinase γ dependent signaling pathway in this promotion of tumor cell invasiveness triggered by OR stimulation. Finally, after subcutaneous inoculation of LNCaP cells into NSG immunodeficient mice, the in vivo stimulation of these cells by the PSGR agonist β-ionone significantly enhanced metastasis emergence and spreading. PMID:24416348

  6. Peripheral NMDA Receptors Mediate Antidromic Nerve Stimulation-Induced Tactile Hypersensitivity in the Rat

    PubMed Central

    Jang, Jun Ho; Nam, Taick Sang; Jun, Jaebeom; Jung, Se Jung; Kim, Dong-Wook; Leem, Joong Woo

    2015-01-01

    We investigated the role of peripheral NMDA receptors (NMDARs) in antidromic nerve stimulation-induced tactile hypersensitivity outside the skin area innervated by stimulated nerve. Tetanic electrical stimulation (ES) of the decentralized L5 spinal nerve, which induced enlargement of plasma extravasation, resulted in tactile hypersensitivity in the L4 plantar dermatome of the hind-paw. When intraplantar (i.pl.) injection was administered into the L4 dermatome before ES, NMDAR and group-I metabotropic Glu receptor (mGluR) antagonists and group-II mGluR agonist but not AMPA/kainate receptor antagonist prevented ES-induced hypersensitivity. I.pl. injection of PKA or PKC inhibitors also prevented ES-induced hypersensitivity. When the same injections were administered after establishment of ES-induced hypersensitivity, hypersensitivity was partially reduced by NMDAR antagonist only. In naïve animals, i.pl. Glu injection into the L4 dermatome induced tactile hypersensitivity, which was blocked by NMDAR antagonist and PKA and PKC inhibitors. These results suggest that the peripheral release of Glu, induced by antidromic nerve stimulation, leads to the expansion of tactile hypersensitive skin probably via nociceptor sensitization spread due to the diffusion of Glu into the skin near the release site. In addition, intracellular PKA- and PKC-dependent mechanisms mediated mainly by NMDAR activation are involved in Glu-induced nociceptor sensitization and subsequent hypersensitivity. PMID:26770021

  7. Peripheral NMDA Receptors Mediate Antidromic Nerve Stimulation-Induced Tactile Hypersensitivity in the Rat.

    PubMed

    Jang, Jun Ho; Nam, Taick Sang; Jun, Jaebeom; Jung, Se Jung; Kim, Dong-Wook; Leem, Joong Woo

    2015-01-01

    We investigated the role of peripheral NMDA receptors (NMDARs) in antidromic nerve stimulation-induced tactile hypersensitivity outside the skin area innervated by stimulated nerve. Tetanic electrical stimulation (ES) of the decentralized L5 spinal nerve, which induced enlargement of plasma extravasation, resulted in tactile hypersensitivity in the L4 plantar dermatome of the hind-paw. When intraplantar (i.pl.) injection was administered into the L4 dermatome before ES, NMDAR and group-I metabotropic Glu receptor (mGluR) antagonists and group-II mGluR agonist but not AMPA/kainate receptor antagonist prevented ES-induced hypersensitivity. I.pl. injection of PKA or PKC inhibitors also prevented ES-induced hypersensitivity. When the same injections were administered after establishment of ES-induced hypersensitivity, hypersensitivity was partially reduced by NMDAR antagonist only. In naïve animals, i.pl. Glu injection into the L4 dermatome induced tactile hypersensitivity, which was blocked by NMDAR antagonist and PKA and PKC inhibitors. These results suggest that the peripheral release of Glu, induced by antidromic nerve stimulation, leads to the expansion of tactile hypersensitive skin probably via nociceptor sensitization spread due to the diffusion of Glu into the skin near the release site. In addition, intracellular PKA- and PKC-dependent mechanisms mediated mainly by NMDAR activation are involved in Glu-induced nociceptor sensitization and subsequent hypersensitivity.

  8. Stimulation of sky receptor tyrosine kinase by the product of growth arrest-specific gene 6.

    PubMed

    Ohashi, K; Nagata, K; Toshima, J; Nakano, T; Arita, H; Tsuda, H; Suzuki, K; Mizuno, K

    1995-09-29

    Sky (also called Rse, Brt, and Tyro3) is a member of a subfamily of related receptor tyrosine kinases, including Axl/Ufo/Ark and c-Eyk/Mer. We obtained evidence that Gas6 (the product of growth arrest-specific gene 6) is a ligand of the Sky receptor tyrosine kinase. Gas6, but not protein S (an anticoagulant protein structurally similar to Gas6), specifically bound to the soluble form of Sky (Sky-Fc), composed of the extracellular domain of Sky fused to the Fc domain of human immunoglobulin G1. The native and recombinant Gas6, but not protein S, stimulated tyrosine phosphorylation of Sky ectopically expressed in Chinese hamster ovary cells. Stimulation of Sky in response to Gas6 was inhibited by Sky-Fc. The half-maximal concentration of Gas6 that stimulated Sky was about 1 nM. Thus, Gas6 as a ligand for Sky specifically binds to and stimulates Sky receptor tyrosine kinase.

  9. Promotion of cancer cell invasiveness and metastasis emergence caused by olfactory receptor stimulation.

    PubMed

    Sanz, Guenhaël; Leray, Isabelle; Dewaele, Aurélie; Sobilo, Julien; Lerondel, Stéphanie; Bouet, Stéphan; Grébert, Denise; Monnerie, Régine; Pajot-Augy, Edith; Mir, Lluis M

    2014-01-01

    Olfactory receptors (ORs) are expressed in the olfactory epithelium, where they detect odorants, but also in other tissues with additional functions. Some ORs are even overexpressed in tumor cells. In this study, we identified ORs expressed in enterochromaffin tumor cells by RT-PCR, showing that single cells can co-express several ORs. Some of the receptors identified were already reported in other tumors, but they are orphan (without known ligand), as it is the case for most of the hundreds of human ORs. Thus, genes coding for human ORs with known ligands were transfected into these cells, expressing functional heterologous ORs. The in vitro stimulation of these cells by the corresponding OR odorant agonists promoted cell invasion of collagen gels. Using LNCaP prostate cancer cells, the stimulation of the PSGR (Prostate Specific G protein-coupled Receptor), an endogenously overexpressed OR, by β-ionone, its odorant agonist, resulted in the same phenotypic change. We also showed the involvement of a PI3 kinase γ dependent signaling pathway in this promotion of tumor cell invasiveness triggered by OR stimulation. Finally, after subcutaneous inoculation of LNCaP cells into NSG immunodeficient mice, the in vivo stimulation of these cells by the PSGR agonist β-ionone significantly enhanced metastasis emergence and spreading.

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

    SciTech Connect

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

    2011-11-18

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

  11. Extracellular polysaccharides produced by Ganoderma formosanum stimulate macrophage activation via multiple pattern-recognition receptors

    PubMed Central

    2012-01-01

    Background The fungus of Ganoderma is a traditional medicine in Asia with a variety of pharmacological functions including anti-cancer activities. We have purified an extracellular heteropolysaccharide fraction, PS-F2, from the submerged mycelia culture of G. formosanum and shown that PS-F2 exhibits immunostimulatory activities. In this study, we investigated the molecular mechanisms of immunostimulation by PS-F2. Results PS-F2-stimulated TNF-α production in macrophages was significantly reduced in the presence of blocking antibodies for Dectin-1 and complement receptor 3 (CR3), laminarin, or piceatannol (a spleen tyrosine kinase inhibitor), suggesting that PS-F2 recognition by macrophages is mediated by Dectin-1 and CR3 receptors. In addition, the stimulatory effect of PS-F2 was attenuated in the bone marrow-derived macrophages from C3H/HeJ mice which lack functional Toll-like receptor 4 (TLR4). PS-F2 stimulation triggered the phosphorylation of mitogen-activated protein kinases JNK, p38, and ERK, as well as the nuclear translocation of NF-κB, which all played essential roles in activating TNF-α expression. Conclusions Our results indicate that the extracellular polysaccharides produced by G. formosanum stimulate macrophages via the engagement of multiple pattern-recognition receptors including Dectin-1, CR3 and TLR4, resulting in the activation of Syk, JNK, p38, ERK, and NK-κB and the production of TNF-α. PMID:22883599

  12. Effects of combined carotid chemoreceptor and atrial receptor stimulation on renal blood flow in anaesthetized dogs.

    PubMed Central

    Kappagoda, C T; Karim, F; Mackay, D

    1983-01-01

    In dogs anaesthetized with chloralose and artificially ventilated, carotid chemoreceptors were stimulated by changing the perfusate of the vascularly isolated carotid bifurcations from arterial to venous blood. Left atrial receptors were stimulated by distending balloons in two pulmonary vein-left atrial junctions and in this left atrial appendage. The left renal blood flow was measured by an electromagnetic flow meter at a constant systemic (renal) arterial pressure in preparations in which heart rate changes were prevented by administration of propranolol hydrochloride (0.5 mg kg-1) and atropine sulphate (0.4 mg kg-1). Muscular movement was prevented by gallamine triethiodide (0.2 mg kg-1). Stimulation of left atrial receptors resulted in a significant increase (P less than 0.001) in renal blood flow of 5.6 +/- 0.88 ml min-1 100 g-1 renal mass from a control of 223 ml min-1 100 g-1 renal mass. The responses were abolished by cooling the cervical vagus nerves to 6-8 degrees C. Stimulation of carotid chemoreceptors, by perfusion of the carotid bifurcations by venous blood, caused a decrease in renal blood flow of 20 +/- 6.9 ml min-1 100 g-1 renal mass from 224 ml min-1 100 g-1 renal mass. Stimulation of left atrial receptors during venous perfusion of carotid chemoreceptors resulted in an increase in renal blood flow of 10.9 +/- 1.82 ml min-1 100 g-1 renal mass from 208 ml min-1 100 g-1 renal mass. These results show that atrial receptors and chemoreceptors can interact in their effects on renal blood flow. PMID:6410054

  13. Norgestrel and gestodene stimulate breast cancer cell growth through an oestrogen receptor mediated mechanism.

    PubMed Central

    Catherino, W. H.; Jeng, M. H.; Jordan, V. C.

    1993-01-01

    There is great concern over the long-term influence of oral contraceptives on the development of breast cancer in women. Oestrogens are known to stimulate the growth of human breast cancer cells, and this laboratory has previously reported (Jeng & Jordan, 1991) that the 19-norprogestin norethindrone could stimulate the proliferation of MCF-7 human breast cancer cells. We studied the influence of the 19-norprogestins norgestrel and gestodene compared to a 'non' 19-norprogestin medroxyprogesterone acetate (MPA) on MCF-7 cell proliferation. The 19-norprogestins stimulated proliferation at a concentration of 10(-8) M, while MPA could not stimulate proliferation at concentrations as great as 3 x 10(-6) M. The stimulatory activity of the 19-norprogestins could be blocked by the antioestrogen ICI 164,384, but not by the antiprogestin RU486. Transfection studies with the reporter plasmids containing an oestrogen response element or progesterone response element (vitERE-CAT, pS2ERE-CAT, and PRE15-CAT) were performed to determine the intracellular action of norgestrel and gestodene. The 19-norprogestins stimulated the vitERE-CAT activity maximally at 10(-6) M, and this stimulation was inhibited by the addition of ICI 164,384. MPA did not stimulate vitERE-CAT activity. A single base pair alteration in the palindromic sequence of vitERE (resulting in the pS2ERE) led to a dramatic decrease in CAT expression by the 19-norprogestins, suggesting that the progestin activity required specific response element base sequencing. PRE15-CAT activity was stimulated by norgestrel, gestodene and MPA at concentrations well below growth stimulatory activity. This stimulation could be blocked by RU486. These studies suggest that the 19-norprogestins norgestrel and gestodene stimulate MCF-7 breast cancer cell growth by activating the oestrogen receptor. PMID:8494728

  14. Effects of kappa-opioid receptor ligands on intracranial self-stimulation in rats.

    PubMed

    Todtenkopf, Mark S; Marcus, Jacqueline F; Portoghese, Philip S; Carlezon, William A

    2004-04-01

    Elevations in cAMP response element binding protein (CREB) function within the mesolimbic system of rats reduce cocaine reward in place conditioning studies and increase immobility in the forced swim test. Each of these behavioral adaptations can be interpreted as a depressive-like effect (i.e., anhedonia, despair) that may reflect reduced activity of brain reward systems. Furthermore, each effect appears due to increases in CREB-mediated expression of dynorphin, since each is attenuated by intracranial injections of the kappa-opioid receptor antagonist norBNI. Intracranial self-stimulation (ICSS) studies were conducted in rats to determine whether administration of a kappa-agonist would have depressive-like effects on brain stimulation reward, and whether pretreatment with a kappa-antagonist would attenuate any such effects. Conditions that have depressive effects in people (e.g., drug withdrawal) increase the threshold amounts of stimulation required to sustain ICSS in rats. Sprague-Dawley rats with lateral hypothalamic stimulating electrodes were tested in a "curve-shift" variant of the ICSS procedure after systemic administration of the kappa-agonist U-69593 alone, the novel kappa-antagonist 5'-acetamidinoethylnaltrindole (ANTI) alone, or co-administration of both drugs. U-69593 dose dependently increased ICSS thresholds, suggesting that activation of kappa-receptors reduced the rewarding impact of the brain stimulation. ANTI had no effects on its own, but it attenuated increases in ICSS thresholds caused by the agonist. These data provide further evidence that stimulation of brain kappa-receptors may trigger certain depressive-like signs, and that kappa antagonists may have efficacy as antidepressants without having reward-related actions of their own.

  15. Peroxisome proliferator-activated receptor gamma activation inhibits progesterone-stimulated human MUC1 expression.

    PubMed

    Wang, Peng; Dharmaraj, Neeraja; Brayman, Melissa J; Carson, Daniel D

    2010-07-01

    Mucin 1 (MUC1) is a type I transmembrane glycoprotein abundantly expressed on nearly all epithelial tissues and overexpressed by many cancer cells. Previous studies from our lab showed that progesterone receptor (PR)B is a strong stimulator of MUC1 gene expression. It is reported that liganded peroxisome proliferator-activated receptor gamma (PPARgamma) stimulates Muc1 expression in murine trophoblast. Here, we demonstrate that although the PPARgamma ligand, rosiglitazone, stimulates the murine Muc1 promoter in HEC1A, a human uterine epithelial cell line, rosiglitazone alone, has no significant effect on basal human MUC1 promoter activity. In fact, rosiglitazone treatment antagonizes progesterone-stimulated human MUC1 promoter activity and protein expression in two human uterine epithelial cell lines and T47D human breast cancer cells. This response is antagonized by the PPARgamma antagonist, GW9662, as well as a dominant-negative form of PPARgamma, demonstrating the response is mediated by PPARgamma. Additional studies indicate that PPARgamma activation does not change PR binding to the MUC1 promoter but generally antagonizes progesterone activity by stimulating PRB degradation and inhibiting progesterone-induced PRB phosphorylation. Collectively, these studies indicate that PPARgamma activation inhibits PRB activity through both acute (phosphorylation) and long-term (PRB degradation) pathways.

  16. Artificial sweeteners stimulate adipogenesis and suppress lipolysis independently of sweet taste receptors.

    PubMed

    Simon, Becky R; Parlee, Sebastian D; Learman, Brian S; Mori, Hiroyuki; Scheller, Erica L; Cawthorn, William P; Ning, Xiaomin; Gallagher, Katherine; Tyrberg, Björn; Assadi-Porter, Fariba M; Evans, Charles R; MacDougald, Ormond A

    2013-11-08

    G protein-coupled receptors mediate responses to a myriad of ligands, some of which regulate adipocyte differentiation and metabolism. The sweet taste receptors T1R2 and T1R3 are G protein-coupled receptors that function as carbohydrate sensors in taste buds, gut, and pancreas. Here we report that sweet taste receptors T1R2 and T1R3 are expressed throughout adipogenesis and in adipose tissues. Treatment of mouse and human precursor cells with artificial sweeteners, saccharin and acesulfame potassium, enhanced adipogenesis. Saccharin treatment of 3T3-L1 cells and primary mesenchymal stem cells rapidly stimulated phosphorylation of Akt and downstream targets with functions in adipogenesis such as cAMP-response element-binding protein and FOXO1; however, increased expression of peroxisome proliferator-activated receptor γ and CCAAT/enhancer-binding protein α was not observed until relatively late in differentiation. Saccharin-stimulated Akt phosphorylation at Thr-308 occurred within 5 min, was phosphatidylinositol 3-kinase-dependent, and occurred in the presence of high concentrations of insulin and dexamethasone; phosphorylation of Ser-473 occurred more gradually. Surprisingly, neither saccharin-stimulated adipogenesis nor Thr-308 phosphorylation was dependent on expression of T1R2 and/or T1R3, although Ser-473 phosphorylation was impaired in T1R2/T1R3 double knock-out precursors. In mature adipocytes, artificial sweetener treatment suppressed lipolysis even in the presence of forskolin, and lipolytic responses were correlated with phosphorylation of hormone-sensitive lipase. Suppression of lipolysis by saccharin in adipocytes was also independent of T1R2 and T1R3. These results suggest that some artificial sweeteners have previously uncharacterized metabolic effects on adipocyte differentiation and metabolism and that effects of artificial sweeteners on adipose tissue biology may be largely independent of the classical sweet taste receptors, T1R2 and T1R3.

  17. Artificial Sweeteners Stimulate Adipogenesis and Suppress Lipolysis Independently of Sweet Taste Receptors*

    PubMed Central

    Simon, Becky R.; Parlee, Sebastian D.; Learman, Brian S.; Mori, Hiroyuki; Scheller, Erica L.; Cawthorn, William P.; Ning, Xiaomin; Gallagher, Katherine; Tyrberg, Björn; Assadi-Porter, Fariba M.; Evans, Charles R.; MacDougald, Ormond A.

    2013-01-01

    G protein-coupled receptors mediate responses to a myriad of ligands, some of which regulate adipocyte differentiation and metabolism. The sweet taste receptors T1R2 and T1R3 are G protein-coupled receptors that function as carbohydrate sensors in taste buds, gut, and pancreas. Here we report that sweet taste receptors T1R2 and T1R3 are expressed throughout adipogenesis and in adipose tissues. Treatment of mouse and human precursor cells with artificial sweeteners, saccharin and acesulfame potassium, enhanced adipogenesis. Saccharin treatment of 3T3-L1 cells and primary mesenchymal stem cells rapidly stimulated phosphorylation of Akt and downstream targets with functions in adipogenesis such as cAMP-response element-binding protein and FOXO1; however, increased expression of peroxisome proliferator-activated receptor γ and CCAAT/enhancer-binding protein α was not observed until relatively late in differentiation. Saccharin-stimulated Akt phosphorylation at Thr-308 occurred within 5 min, was phosphatidylinositol 3-kinase-dependent, and occurred in the presence of high concentrations of insulin and dexamethasone; phosphorylation of Ser-473 occurred more gradually. Surprisingly, neither saccharin-stimulated adipogenesis nor Thr-308 phosphorylation was dependent on expression of T1R2 and/or T1R3, although Ser-473 phosphorylation was impaired in T1R2/T1R3 double knock-out precursors. In mature adipocytes, artificial sweetener treatment suppressed lipolysis even in the presence of forskolin, and lipolytic responses were correlated with phosphorylation of hormone-sensitive lipase. Suppression of lipolysis by saccharin in adipocytes was also independent of T1R2 and T1R3. These results suggest that some artificial sweeteners have previously uncharacterized metabolic effects on adipocyte differentiation and metabolism and that effects of artificial sweeteners on adipose tissue biology may be largely independent of the classical sweet taste receptors, T1R2 and T1R3. PMID

  18. Inhibition of forskolin-stimulated adenylate cyclase activity by 5-HT receptor agonists.

    PubMed

    Devivo, M; Maayani, S

    1985-12-17

    We measured the inhibition of forskolin-stimulated adenylate cyclase activity in guinea pig hippocampal membranes by 5-HT, 5-carboxamidotryptamine (CAT) and 8-hydroxy-2-(di-n-propylamino) tetralin (PAT). Low concentrations of these agonists inhibited forskolin-stimulated adenylate cyclase activity in a concentration-dependent and saturable manner. The antagonist spiperone shifted the concentration-response curve to CAT to the right in a parallel manner. The EC50 values of CAT, PAT and 5-HT and the KB of spiperone suggest that this receptor may correspond to the 5-HT1A binding site.

  19. Alpha/sub 1/ receptor stimulated phosphatidylinositol hydrolysis in rat cerebral cortex

    SciTech Connect

    Raulli, R.; Crews, F.T.

    1986-03-05

    The potency of various alpha adrenergic compounds on stimulation of phosphatidylinositol (PI) hydrolysis was determined using (/sup 3/H)-inositol labelled cerebral cortical slices. Norepinephrine-induced PI hydrolysis was inhibited by the alpha/sub 1/ selective antagonist prazosin (1 ..mu..M) but not the beta receptor antagonist propranolol (1 ..mu..M). Tramazoline, (-)-ephedrine, and (+/-)-phenylpropanolamine were all found to be partial agonists at 1 mM concentrations. Clonidine, naphazoline, trazodone, and the novel antidepressant mianserin at concentrations of 100 ..mu..M to 1 mM produced no significant increase in PI hydrolysis above control levels. The relationship between responses and receptor binding will be discussed.

  20. Combinatorial diffusion assay used to identify topically active melanocyte-stimulating hormone receptor antagonists.

    PubMed Central

    Quillan, J M; Jayawickreme, C K; Lerner, M R

    1995-01-01

    alpha-Melanocyte-stimulating hormone (alpha-MSH) is implicated in pigmentation, central nervous system and immune system functions, growth, mitogenesis, and melanoma. Evaluation of these roles has been hindered by the lack of alpha-MSH antagonists. A combinatorial chemistry-based diffusion assay is used to find random tripeptides that antagonize normal frog and human melanoma MSH receptors and to identify pharmacological groups responsible for receptor interaction. The alpha-MSH antagonist D-Trp-Arg-Leu-NH2 is used to demonstrate directly the contribution of MSH to normal skin tone in frogs following injection or topical application. Images Fig. 1 Fig. 3 Fig. 4 Fig. 6 PMID:7708744

  1. Evidence for a central 5-hydroxytryptamine receptor stimulation by lysergic acid diethylamide

    PubMed Central

    Andén, N.-E.; Corrodi, H.; Fuxe, K.; Hökfelt, T.

    1968-01-01

    1. Lysergic acid diethylamide (LSD) and the 5-hydroxytryptamine (5-HT) precursor, 5-hydroxytryptophan produced similar functional effects in rat spinal cord and brain to the 5-hydroxytryptamine precursor 5-hydroxytryptophan, which indicates that LSD stimulates central 5-HT receptors. 2. By means of combined histochemical and biochemical techniques it was found that LSD reduced the turnover rate of brain and spinal cord 5-HT, studied after inhibition of the tryptophan hydroxylase by α-propyldopacetamide. The turnover of brain noradrenaline but not dopamine was somewhat accelerated. 3. The functional and chemical effects by LSD were related to dose and to time. They were not observed after the LSD analogues 2-bromo-LSD and methylsergide. 4. The retardation of the 5-HT turnover by LSD may be due to negative feed-back mechanisms evoked by direct stimulation of the central 5-HT receptors. ImagesFIG. 1FIG. 2 PMID:5302837

  2. Extracellular matrix hyaluronan signals via its CD44 receptor in the increased responsiveness to mechanical stimulation.

    PubMed

    Ferrari, L F; Araldi, D; Bogen, O; Levine, J D

    2016-06-02

    We propose that the extracellular matrix (ECM) signals CD44, a hyaluronan receptor, to increase the responsiveness to mechanical stimulation in the rat hind paw. We report that intradermal injection of hyaluronidase induces mechanical hyperalgesia, that is inhibited by co-administration of a CD44 receptor antagonist, A5G27. The intradermal injection of low (LMWH) but not high (HMWH) molecular weight hyaluronan also induces mechanical hyperalgesia, an effect that was attenuated by pretreatment with HMWH or A5G27. Pretreatment with HMWH also attenuated the hyperalgesia induced by hyaluronidase. Similarly, intradermal injection of A6, a CD44 receptor agonist, produced hyperalgesia that was inhibited by HMWH and A5G27. Inhibitors of protein kinase A (PKA) and Src, but not protein kinase C (PKC), significantly attenuated the hyperalgesia induced by both A6 and LMWH. Finally, to determine if CD44 receptor signaling is involved in a preclinical model of inflammatory pain, we evaluated the effect of A5G27 and HMWH on the mechanical hyperalgesia associated with the inflammation induced by carrageenan. Both A5G27 and HMWH attenuated carrageenan-induced mechanical hyperalgesia. Thus, while LMWH acts at its cognate receptor, CD44, to induce mechanical hyperalgesia, HMWH acts at the same receptor as an antagonist. That the local administration of HMWH or A5G27 inhibits carrageenan-induced hyperalgesia supports the suggestion that carrageenan produces changes in the ECM that contributes to inflammatory pain. These studies define a clinically relevant role for signaling by the hyaluronan receptor, CD44, in increased responsiveness to mechanical stimulation.

  3. Skeletal muscle beta-receptors and isoproterenol-stimulated vasodilation in canine heart failure

    SciTech Connect

    Frey, M.J.; Lanoce, V.; Molinoff, P.B.; Wilson, J.R. )

    1989-11-01

    To investigate whether heart failure alters beta-adrenergic receptors on skeletal muscle and its associated vasculature, the density of beta-adrenergic receptors, isoproterenol-stimulated adenylate cyclase activity, and coupling of the guanine nucleotide-binding regulatory protein were compared in 18 control dogs and 16 dogs with heart failure induced by 5-8 wk of ventricular pacing at 260 beats/min. Hindlimb vascular responses to isoproterenol were compared in eight controls and eight of the dogs with heart failure. In dogs with heart failure, the density of beta-receptors on skeletal muscle was reduced in both gastrocnemius (control: 50 +/- 5; heart failure: 33 +/- 8 fmol/mg of protein) and semitendinosus muscle (control: 43 +/- 9; heart failure: 27 +/- 9 fmol/mg of protein, both P less than 0.05). Receptor coupling to the ternary complex, as determined by isoproterenol competition curves with and without guanosine 5'-triphosphate (GTP), was unchanged. Isoproterenol-stimulated adenylate cyclase activity was significantly decreased in semitendinosus muscle (control: 52.4 +/- 4.6; heart failure: 36.5 +/- 9.5 pmol.mg-1.min-1; P less than 0.05) and tended to be decreased in gastrocnemius muscle (control: 40.1 +/- 8.5; heart failure: 33.5 +/- 4.5 pmol.mg-1.min-1; P = NS). Isoproterenol-induced hindlimb vasodilation was not significantly different in controls and in dogs with heart failure. These findings suggest that heart failure causes downregulation of skeletal muscle beta-adrenergic receptors, probably due to receptor exposure to elevated catecholamine levels, but does not reduce beta-receptor-mediated vasodilation in muscle.

  4. Stimulation of Estrogen Receptor Signaling in Breast Cancer by a Novel Chaperone Synuclein Gamma

    DTIC Science & Technology

    2007-06-01

    dependent cancers of breast and ovary promoted us to investigate the role of SNCG in regulation of ERα. SNCG strongly stimulated the ligand-dependent...promoted us to investigate the role of SNCG in regulation of estrogen receptor ER-α. BODY A notable finding relevant to this study is that SNCG...and antiestrogens. Cell growth was measured using a cell proliferation kit (XTT). Data are the mean ± SD of quadruplicate cultures. Open bar

  5. Stimulation of Estrogen Receptor Signaling in Breast Cancer by a Novel Chaperone Gamma Synuclein

    DTIC Science & Technology

    2009-06-01

    AD_________________ AWARD NUMBER: W81XWH- 04 -1-0569 TITLE: Stimulation of Estrogen Receptor...Synuclein 5b. GRANT NUMBER W81XWH- 04 -1-0569 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT NUMBER Yuenian Shi 5e. TASK NUMBER E-Mail...EShi@LIJ.EDU 5f. WORK UNIT NUMBER 7 . PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) North Shore-Long Island Jewish Research Institute

  6. Abluminal Stimulation of Sphingosine 1-Phosphate Receptors 1 and 3 Promotes and Stabilizes Endothelial Sprout Formation

    PubMed Central

    Lenz, Steven M.; Awojoodu, Anthony O.

    2015-01-01

    Local delivery of lipid mediators has become a promising new approach for therapeutic angiogenesis and regenerative medicine. In this study, we investigated how gradient stimulation (either abluminal/distal or luminal/proximal) of engineered microvessels with sphingosine 1-phosphate (S1P) receptor-subtype-targeted molecules affects endothelial sprout growth using a microfluidic device. Our studies show that distal stimulation of microvessels with FTY720, an S1P1/3 selective agonist, promotes both arterial and venular sprout growth, whereas proximal stimulation does not. Using novel pharmacological antagonists of S1P receptor subtypes, we further show that S1P3 functionality is necessary for VEGF-induced sprouting, and confirmed these findings ex vivo using a murine aortic ring assay from S1P3-deficient mice. S1P3 agonist stimulation enhanced vascular stability in both cell types via upregulation of the interendothelial junction protein VE-cadherin. Lastly, S1P3 activation under flow promoted endothelial sprouting and branching while decreasing migratory cell fate in the microfluidic device. We used an in vivo murine dorsal skinfold window chamber model to confirm S1P3's role in neovascular branching. Together, these data suggest that a distal transendothelial gradient of S1P1/3-targeted drugs is an effective technique for both enhancing and stabilizing capillary morphogenesis in angiogenic applications. PMID:25315888

  7. Synaptic NMDA receptor stimulation activates PP1 by inhibiting its phosphorylation by Cdk5

    PubMed Central

    Hou, Hailong; Sun, Lu; Siddoway, Benjamin A.; Petralia, Ronald S.; Yang, Hongtian; Gu, Hua; Nairn, Angus C.

    2013-01-01

    The serine/threonine protein phosphatase protein phosphatase 1 (PP1) is known to play an important role in learning and memory by mediating local and downstream aspects of synaptic signaling, but how PP1 activity is controlled in different forms of synaptic plasticity remains unknown. We find that synaptic N-methyl-d-aspartate (NMDA) receptor stimulation in neurons leads to activation of PP1 through a mechanism involving inhibitory phosphorylation at Thr320 by Cdk5. Synaptic stimulation led to proteasome-dependent degradation of the Cdk5 regulator p35, inactivation of Cdk5, and increased auto-dephosphorylation of Thr320 of PP1. We also found that neither inhibitor-1 nor calcineurin were involved in the control of PP1 activity in response to synaptic NMDA receptor stimulation. Rather, the PP1 regulatory protein, inhibitor-2, formed a complex with PP1 that was controlled by synaptic stimulation. Finally, we found that inhibitor-2 was critical for the induction of long-term depression in primary neurons. Our work fills a major gap regarding the regulation of PP1 in synaptic plasticity. PMID:24189275

  8. Abluminal stimulation of sphingosine 1-phosphate receptors 1 and 3 promotes and stabilizes endothelial sprout formation.

    PubMed

    Das, Anusuya; Lenz, Steven M; Awojoodu, Anthony O; Botchwey, Edward A

    2015-01-01

    Local delivery of lipid mediators has become a promising new approach for therapeutic angiogenesis and regenerative medicine. In this study, we investigated how gradient stimulation (either abluminal/distal or luminal/proximal) of engineered microvessels with sphingosine 1-phosphate (S1P) receptor-subtype-targeted molecules affects endothelial sprout growth using a microfluidic device. Our studies show that distal stimulation of microvessels with FTY720, an S1P1/3 selective agonist, promotes both arterial and venular sprout growth, whereas proximal stimulation does not. Using novel pharmacological antagonists of S1P receptor subtypes, we further show that S1P3 functionality is necessary for VEGF-induced sprouting, and confirmed these findings ex vivo using a murine aortic ring assay from S1P3-deficient mice. S1P3 agonist stimulation enhanced vascular stability in both cell types via upregulation of the interendothelial junction protein VE-cadherin. Lastly, S1P3 activation under flow promoted endothelial sprouting and branching while decreasing migratory cell fate in the microfluidic device. We used an in vivo murine dorsal skinfold window chamber model to confirm S1P3's role in neovascular branching. Together, these data suggest that a distal transendothelial gradient of S1P1/3-targeted drugs is an effective technique for both enhancing and stabilizing capillary morphogenesis in angiogenic applications.

  9. Gastrointestinal hormones stimulate growth of Foregut Neuroendocrine Tumors by transactivating the EGF receptor.

    PubMed

    Di Florio, Alessia; Sancho, Veronica; Moreno, Paola; Delle Fave, Gianfranco; Jensen, Robert T

    2013-03-01

    Foregut neuroendocrine tumors [NETs] usually pursuit a benign course, but some show aggressive behavior. The treatment of patients with advanced NETs is marginally effective and new approaches are needed. In other tumors, transactivation of the EGF receptor (EGFR) by growth factors, gastrointestinal (GI) hormones and lipids can stimulate growth, which has led to new treatments. Recent studies show a direct correlation between NET malignancy and EGFR expression, EGFR inhibition decreases basal NET growth and an autocrine growth effect exerted by GI hormones, for some NETs. To determine if GI hormones can stimulate NET growth by inducing transactivation of EGFR, we examined the ability of EGF, TGFα and various GI hormones to stimulate growth of the human foregut carcinoid,BON, the somatostatinoma QGP-1 and the rat islet tumor,Rin-14B-cell lines. The EGFR tyrosine-kinase inhibitor, AG1478 strongly inhibited EGF and the GI hormones stimulated cell growth, both in BON and QGP-1 cells. In all the three neuroendocrine cell lines studied, we found EGF, TGFα and the other growth-stimulating GI hormones increased Tyr(1068) EGFR phosphorylation. In BON cells, both the GI hormones neurotensin and a bombesin analogue caused a time- and dose-dependent increase in EGFR phosphorylation, which was strongly inhibited by AG1478. Moreover, we found this stimulated phosphorylation was dependent on Src kinases, PKCs, matrix metalloproteinase activation and the generation of reactive oxygen species. These results raise the possibility that disruption of this signaling cascade by either EGFR inhibition alone or combined with receptor antagonists may be a novel therapeutic approach for treatment of foregut NETs/PETs.

  10. Gastrointestinal hormones stimulate growth of Foregut Neuroendocrine Tumors by transactivating the EGF receptor

    PubMed Central

    Di Florio, Alessia; Sancho, Veronica; Moreno, Paola; Fave, Gianfranco Delle; Jensen, Robert T.

    2012-01-01

    Foregut Neuroendocrine Tumors[NETs] usually pursuit a benign course, but some show aggressive behavior. The treatment of patients with advanced NETs is marginally effective and new approaches are needed. In other tumors, transactivation of the EGF receptor(EGFR) by growth factors, gastrointestinal(GI) hormones and lipids can stimulate growth, which has led to new treatments. Recent studies show a direct correlation between NET malignancy and EGFR expression, EGFR inhibition decreases basal NET growth and an autocrine growth effect exerted by GI hormones, for some NETs. To determine if GI hormones can stimulate NET growth by inducing transactivation of EGFR, we examined the ability of EGF, TGFα and various GI hormones to stimulate growth of the human foregut carcinoid, BON, the somatostatinoma QGP-1 and the rat islet tumor, Rin-14B-cell lines. The EGFR tyrosine-kinase inhibitor, AG1478 strongly inhibited EGF and the GI hormones stimulated cell growth, both in BON and QGP-1 cells. In all the three neuroendocrine cell lines studied, we found EGF, TGFα and the other growth-stimulating GI hormones increased Tyr1068 EGFR phosphorylation. In BON cells, both the GI hormones neurotensin and a bombesin analogue caused a time- and dose-dependent increase in EGFR phosphorylation, which was strongly inhibited by AG1478. Moreover, we found this stimulated phosphorylation was dependent on Src kinases, PKCs, matrix metalloproteinase activation and the generation of reactive oxygen species. These results raise the possibility that disruption of this signaling cascade by either EGFR inhibition alone or combined with receptor antagonists may be a novel therapeutic approach for treatment of foregut NETs/PETs. PMID:23220008

  11. Activation of G-proteins by receptor-stimulated nucleoside diphosphate kinase in Dictyostelium.

    PubMed Central

    Bominaar, A A; Molijn, A C; Pestel, M; Veron, M; Van Haastert, P J

    1993-01-01

    Recently, interest in the enzyme nucleoside diphosphate kinase (EC2.7.4.6) has increased as a result of its possible involvement in cell proliferation and development. Since NDP kinase is one of the major sources of GTP in cells, it has been suggested that the effects of an altered NDP kinase activity on cellular processes might be the result of altered transmembrane signal transduction via guanine nucleotide-binding proteins (G-proteins). In the cellular slime mould Dictyostelium discoideum, extracellular cAMP induces an increase of phospholipase C activity via a surface cAMP receptor and G-proteins. In this paper it is demonstrated that part of the cellular NDP kinase is associated with the membrane and stimulated by cell surface cAMP receptors. The GTP produced by the action of NDP kinase is capable of activating G-proteins as monitored by altered G-protein-receptor interaction and the activation of the effector enzyme phospholipase C. Furthermore, specific monoclonal antibodies inhibit the effect of NDP kinase on G-protein activation. These results suggest that receptor-stimulated NDP kinase contributes to the mediation of hormone action by producing GTP for the activation of GTP-binding proteins. Images PMID:8389692

  12. Grb10 mediates insulin-stimulated degradation of the insulin receptor: a mechanism of negative regulation.

    PubMed

    Ramos, Fresnida J; Langlais, Paul R; Hu, Derong; Dong, Lily Q; Liu, Feng

    2006-06-01

    Growth factor receptor-bound protein 10 (Grb10) is an adapter protein that interacts with a number of tyrosine-phosphorylated growth factor receptors, including the insulin receptor (IR). To investigate the role of Grb10 in insulin signaling, we generated cell lines in which the expression levels of Grb10 are either overexpressed by stable transfection or suppressed by RNA interference. We found that suppressing endogenous Grb10 expression led to increased IR protein levels, whereas overexpression of Grb10 led to reduced IR protein levels. Altering Grb10 expression levels had no effect on the mRNA levels of IR, suggesting that the modulation occurs at the protein level. Reduced IR levels were also observed in cells with prolonged insulin treatment, and this reduction was inhibited in Grb10-deficient cells. The insulin-induced IR reduction was greatly reversed by MG-132, a proteasomal inhibitor, but not by chloroquine, a lysosomal inhibitor. IR underwent insulin-stimulated ubiquitination in cells, and this ubiquitination was inhibited in the Grb10-suppressed cell line. Together, our results suggest that, in addition to inhibiting IR kinase activity by directly binding to the IR, Grb10 also negatively regulates insulin signaling by mediating insulin-stimulated degradation of the receptor.

  13. Direct Demonstration of Separate Receptors for Growth and Metabolic Activities of Insulin and Multiplication-stimulating Activity (an Insulinlike Growth Factor) Using Antibodies to the Insulin Receptor

    PubMed Central

    King, George L.; Kahn, C. Ronald; Rechler, Matthew M.; Nissley, S. Peter

    1980-01-01

    Insulin and such insulinlike growth factors as multiplication stimulating activity (MSA) are related polypeptides that have common biological activities. Both insulin and MSA produce acute metabolic responses (stimulation of glucose oxidation in isolated fat cells) as well as growth effects (stimulation of [3H]thymidine incorporation into DNA in cultured fibroblasts). In addition, most cells have separate receptors for insulin and insulinlike growth factors, and both peptides have weaker affinity for each other's specific receptors than for their own. To determine, therefore, whether these effects are mediated by receptors for insulin, insulinlike growth factors, or both, we have selectively blocked insulin receptors with a specific antagonist, namely Fab fragments derived from naturally occurring antibodies to the insulin receptor. In rat adipocytes, 10 μg/ml of antireceptor Fab inhibited insulin binding by 90%, whereas it inhibited MSA binding <5%. The anti-insulin receptor Fab is without intrinsic biological activity, but acts as a competitive inhibitor of insulin receptors. Blockade of insulin receptors with Fab fragments produced a 30-fold rightward shift in the dose response for stimulation of glucose oxidation by both insulin and MSA. The dose-response curves for stimulation of oxidation by vitamin K5 and spermine, agents that stimulate glucose oxidation through noninsulin receptor pathways, were not affected by the blockade of insulin receptors with Fab antibody fragments. These data suggest that this acute metabolic effect of both insulin and MSA is mediated via the insulin receptor. In cultured human fibroblasts, 10 μg/ml of Fab inhibited insulin binding by 90% and MSA binding by 15%. In fibroblasts, however, blockade of the insulin receptor did not alter the dose response for stimulation of thymidine incorporation into DNA by either insulin or MSA. Furthermore, intact antireceptor antibody immunoglobulin (Ig)G, which produces multiple other insulinlike

  14. Reconstitution of hormone-responsive detergent-solubilized follicle stimulating hormone receptors into liposomes

    SciTech Connect

    Grasso, P.; Dattatreyamurty, B.; Reichert, L.E. Jr.

    1988-05-01

    An FSH receptor-enriched fraction that responds to exogenous FSH by activation of adenylate cyclase was prepared by ultrafiltration of sucrose density gradient-purified light membranes derived from bovine calf testes homogenates and solubilized with Triton X-100. To further confirm the functional nature of the detergent-solubilized FSH receptor, the extract was incorporated by lipid hydration into large multilamellar vesicles composed of dioleoyl phosphatidylcholine and cholesterol, 2:1 molar ratio. Receptor incorporation was determined by measurement of specific binding of (125I) human FSH ((125I) hFSH). Substitution of dioleoyl phosphatidylcholine with dipalmitoyl phosphatidylcholine or increasing the cholesterol concentration of the vesicles reduced specific binding of (125I)hFSH. Under conditions favoring optimal incorporation of the receptor, specific binding of (125I)hFSH was time and temperature dependent and saturable when increasing concentrations of radioligand were added to a constant amount of proteoliposomes. Reconstituted proteoliposomes bound 1600 fmol FSH/mg protein with an affinity of 3.54 x 10(9) M-1. Inhibition of (125I) hFSH binding by hFSH was comparable to that seen with the membrane-bound receptor (ED50 = 10 ng). Equilibrium binding studies with (3H)Gpp(NH)p indicated that a single class of high affinity GTP binding sites with an association constant (Ka) of 3.33 x 10(7) m-1 which bound 2.19 fmol (3H)Gpp(NH)p/mg protein had also been incorporated into the proteoliposomes. Addition of FSH induced a 2-fold stimulation of (3H)Gpp(NH)p binding, supporting our earlier studies suggesting that the detergent-solubilized FSH receptor is complexed to the G protein. Of particular significance in the present study was the observation that both NaF and FSH stimulated cAMP production in the reconstituted system.

  15. Effects of stimulation of glutamate receptors in medial septum on some immune responses in rats.

    PubMed

    Dutta, Goutam; Raj Goswami, Ananda; Ghosh, Tusharkanti

    2013-11-13

    The immunomodulatory role of medial septum (MS) has been explored so far only in MS lesioned rats. But in MS lesioned rats, all the nerve cells and fibres of the lesioned area are damaged and the specific role of the neural circuits of MS on immunomodulation cannot be assessed from the lesion of MS. Considering the presence of a large number of glutamate receptors in MS, the specific role of glutamate receptors stimulation on some immune responses has been investigated in the present study. Hyperreactive behaviour, TC and DC of WBC, phagocytic activity of peripheral leukocytes, adhesibility and cytotoxicity of splenic mononuclear cells (MNC), delayed type of hypersensitivity (DTH) responses and the serum corticosterone (CORT) were measured after microinfusion of glutamate into MS of rats. To ascertain the specific role of those glutamate receptors, the parameters were also measured after microinfusion of glutamate receptor blocker 6, 7-dinitroquinoxaline-2, 3-dione (DNQX). The hyperreactive behaviour, TC and DC of WBC remained unaltered after stimulation or blocking of glutamate receptors. The phagocytic activity, adhesibility and cytotoxicity of splenic MNC, and DTH responses were increased after infusion of 0.25 and 0.5µM glutamate. But after infusion of higher dose of glutamate (1µM), the phagocytic activity and the adhesibility of splenic MNC were decreased and other parameters remained unaltered in that condition. After infusion of 4 and 8mM DNQX all the observed immunological parameters were decreased. The CORT concentration was decreased after infusion of 0.25 and 0.5µM of glutamate but it was increased after infusion of 1µM glutamate or 4 and 8mM DNQX. Results indicate that the medial septal glutamate receptors play an important role in the modulation of some immune responses.

  16. Similar clinical performance of a novel chimeric thyroid-stimulating hormone receptor bioassay and an automated thyroid-stimulating hormone receptor binding assay in Graves' disease.

    PubMed

    Kamijo, Keiichi; Murayama, Hiroshi; Uzu, Takahiro; Togashi, Kazuyoshi; Olivo, Paul D; Kahaly, George J

    2011-12-01

    Graves' disease (GD) is caused by the continuous stimulation of the thyroid gland by autoantibodies directed against the thyroid-stimulating hormone receptor (TSHR). Two frequent assays for the measurement of TSHR autoantibodies (TSHRAb) were compared, one measuring stimulation of cyclic adenosine monophosphate (cAMP) production and one measuring inhibition of TSH binding, with regard to diagnostic accuracy for GD as well as whether there was an existence of their discordant results in patients with GD and painless thyroiditis (PT). Using 106 sera from untreated GD and 80 sera from autoimmune PT, we compared the diagnostic performance of two TSHRAb assays that have been recently developed. The first one is a bioreporter assay using chimera TSHR (Mc-4), which detects a stimulation signal of cAMP level in cultured CHO cells (Mc4-TSAb assay). The second is a binding inhibition assay using the extracelluar domain of porcine TSHR and a monoclonal antibody (M22) closely mimicking the binding to TSH (M22-TRAb assay). In addition, we compared both assays by using eight sera from eight GD subjects becoming spontaneously hypothyroid due to appearance of thyroid blocking autoantibodies (TBAb) that were measured with inhibition rates of TSH-stimulated cAMP in porcine cells. The Mc4-TSAb assay and the M22-TRAb assay were positive in 94.3% and 92.5% of the GD patients, respectively, whereas they were negative in 95.0% and 98.8% of the PT subjects. However, 10 of 106 GD sera (9.4%) showed discordant results. Six of 106 cases with untreated GD (5.7%) were Mc4-TSAb positive and M22-TRAb negative. In contrast, 4 of 106 sera (3.8%) were Mc4-TSAb negative but M22-TRAb positive. Two cases of untreated GD were negative for both Mc4-TSAb and M22-TRAb. In eight GD subjects with TBAb and hypothyroidism, the binding assay was highly positive, although Mc4-TSAb was negative. Similar and excellent performance was noted for the Mc4-TSAb and M22-TRAb assays in a large group of patients with GD

  17. TGF-beta 1 stimulation of cell locomotion utilizes the hyaluronan receptor RHAMM and hyaluronan

    PubMed Central

    1993-01-01

    TGF-beta is a potent stimulator of motility in a variety of cell types. It has recently been shown that hyaluronan (HA) can directly promote locomotion of cells through interaction with the HA receptor RHAMM. We have investigated the role of RHAMM and HA in TGF-beta-stimulated locomotion and show that TGF-beta triggers the transcription, synthesis and membrane expression of the RHAMM receptor and the secretion of HA coincident with the induction of the locomotory response. This was demonstrated by both incubating cells with exogenous TGF-beta 1 and by stimulating the production of bioactive TGF-beta 1 in tumor cells transfected with TGF-beta 1 under the control of the metallothionein promoter. TGF-beta 1-induced locomotion was suppressed by antibodies that prevented HA/RHAMM interaction, using polyclonal antibodies to either RHAMM fusion protein or RHAMM peptides, or mAbs to purified RHAMM. Peptides corresponding to the HA-binding motif of RHAMM also suppressed TGF-beta 1-induced increases in motility rate. Spontaneous locomotion of fibrosarcoma cells was blocked by neutralizing secreted TGF-beta with panspecific TGF-beta antibodies and by inhibition of TGF- beta 1 secretion with antisense oligonucleotides. Polyclonal anti-RHAMM fusion protein antibodies and peptide from the RHAMM HA-binding motif also suppressed the spontaneous motility rate of fibrosarcoma cells. These data suggest that fibrosarcoma cell locomotion requires TGF-beta, and the pathway by which TGF-beta stimulates locomotion uses the HA receptor RHAMM and HA. PMID:7693717

  18. TGF-beta 1 stimulation of cell locomotion utilizes the hyaluronan receptor RHAMM and hyaluronan.

    PubMed

    Samuel, S K; Hurta, R A; Spearman, M A; Wright, J A; Turley, E A; Greenberg, A H

    1993-11-01

    TGF-beta is a potent stimulator of motility in a variety of cell types. It has recently been shown that hyaluronan (HA) can directly promote locomotion of cells through interaction with the HA receptor RHAMM. We have investigated the role of RHAMM and HA in TGF-beta-stimulated locomotion and show that TGF-beta triggers the transcription, synthesis and membrane expression of the RHAMM receptor and the secretion of HA coincident with the induction of the locomotory response. This was demonstrated by both incubating cells with exogenous TGF-beta 1 and by stimulating the production of bioactive TGF-beta 1 in tumor cells transfected with TGF-beta 1 under the control of the metallothionein promoter. TGF-beta 1-induced locomotion was suppressed by antibodies that prevented HA/RHAMM interaction, using polyclonal antibodies to either RHAMM fusion protein or RHAMM peptides, or mAbs to purified RHAMM. Peptides corresponding to the HA-binding motif of RHAMM also suppressed TGF-beta 1-induced increases in motility rate. Spontaneous locomotion of fibrosarcoma cells was blocked by neutralizing secreted TGF-beta with panspecific TGF-beta antibodies and by inhibition of TGF-beta 1 secretion with antisense oligonucleotides. Polyclonal anti-RHAMM fusion protein antibodies and peptide from the RHAMM HA-binding motif also suppressed the spontaneous motility rate of fibrosarcoma cells. These data suggest that fibrosarcoma cell locomotion requires TGF-beta, and the pathway by which TGF-beta stimulates locomotion uses the HA receptor RHAMM and HA.

  19. Growth hormone receptor polymorphisms and growth hormone response to stimulation test: a pilot study.

    PubMed

    Pagani, Sara; DE Filippo, Gianpaolo; Genoni, Giulia; Rendina, Domenico; Meazza, Cristina; Bozzola, Elena; Bona, Gianni; Bozzola, Mauro

    2016-06-29

    No gold standard pharmacological stimulation test exists for the diagnosis of growth hormone deficiency (GHD). In addition, the genetic factors that influence growth hormone (GH) responses remain unclear. This study aimed to determine whether polymorphisms in exon 6 of the GH receptor gene influence responses to the L-arginine GH stimulation test. This study included 27 prepubertal patients with confirmed GHD. GHD was defined as a peak GH level <8 ng/ml in response to pharmacological stimulation. The mean GH peak after L-arginine stimulation was 2.9 ± 2.9 ng/ml. The included patients had the following genotypes at the third position of codon 168: AA (n=1), AG (n=15) and GG (n=11). Patients carrying the AA and AG genotypes exhibited stronger responses to arginine than patients with the GG genotype (3.1 ± 2.7 vs. 1.5 ± 1.3 ng/ml, p = 0.01). The approach employed in this study could elucidate GH profiles under physiological and pathological conditions, facilitating improved interpretation of pharmacological stimulation tests.

  20. Effects of erythropoietin receptors and erythropoiesis-stimulating agents on disease progression in cancer

    PubMed Central

    Aapro, M; Jelkmann, W; Constantinescu, S N; Leyland-Jones, B

    2012-01-01

    Erythropoiesis-stimulating agents (ESAs) increase red blood cell (RBC) production in bone marrow by activating the erythropoietin receptor (EpoR) on erythrocytic-progenitor cells. Erythropoiesis-stimulating agents are approved in the United States and Europe for treating anaemia in cancer patients receiving chemotherapy based on randomised, placebo-controlled trials showing that ESAs reduce RBC transfusions. Erythropoiesis-stimulating agent-safety issues include thromboembolic events and concerns regarding whether ESAs increase disease progression and/or mortality in cancer patients. Several trials have reported an association between ESA use and increased disease progression and/or mortality, whereas other trials in the same tumour types have not provided similar findings. This review thoroughly examines available evidence regarding whether ESAs affect disease progression. Both clinical-trial data on ESAs and disease progression, and preclinical data on how ESAs could affect tumour growth are summarised. Preclinical topics include (i) whether tumour cells express EpoR and could be directly stimulated to grow by ESA exposure and (ii) whether endothelial cells express EpoR and could be stimulated by ESA exposure to undergo angiogenesis and indirectly promote tumour growth. Although assessment and definition of disease progression vary across studies, the current clinical data suggest that ESAs may have little effect on disease progression in chemotherapy patients, and preclinical data indicate a direct or indirect effect of ESAs on tumour growth is not strongly supported. PMID:22395661

  1. Direct stimulation of angiotensin II type 2 receptor initiated after stroke ameliorates ischemic brain damage.

    PubMed

    Min, Li-Juan; Mogi, Masaki; Tsukuda, Kana; Jing, Fei; Ohshima, Kousei; Nakaoka, Hirotomo; Kan-No, Harumi; Wang, Xiao-Li; Chisaka, Toshiyuki; Bai, Hui-Yu; Iwanami, Jun; Horiuchi, Masatsugu

    2014-08-01

    Stroke is a leading cause of death and disability; however, meta-analysis of randomized controlled trials of blood pressure-lowering drugs in acute stroke has shown no definite evidence of a beneficial effect on functional outcome. Accumulating evidence suggests that angiotensin II type 1 receptor blockade with angiotensin II type 2 (AT2) receptor stimulation could contribute to protection against ischemic brain damage. We examined the possibility that direct AT2 receptor stimulation by compound 21 (C21) initiated even after stroke can prevent ischemic brain damage. Stroke was induced by middle cerebral artery (MCA) occlusion, and the area of cerebral infarction was measured by magnetic resonant imaging. C21 (10 µg/kg/day) treatment was initiated immediately after MCA occlusion by intraperitoneal injection followed by treatment with C21 once daily. We observed that ischemic area was enlarged in a time dependent fashion and decreased on day 5 after MCA occlusion. Treatment with C21 initiated after MCA occlusion significantly reduced the ischemic area, with improvement of neurological deficit in a time-dependent manner without affecting blood pressure. The decrease of cerebral blood flow after MCA occlusion was also ameliorated by C21 treatment. Moreover, treatment with C21 significantly attenuated superoxide anion production and expression of proinflammatory cytokines, monocyte chemoattractant protein 1, and tumor necrosis factor α. Interestingly, C21 administration significantly decreased blood-brain barrier permeability and cerebral edema on the ischemic side. These results provide new evidence that direct AT2 receptor stimulation with C21 is a novel therapeutic approach to prevent ischemic brain damage after acute stroke. © American Journal of Hypertension, Ltd 2014. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  2. Dissociation between neural and vascular responses to sympathetic stimulation : contribution of local adrenergic receptor function

    NASA Technical Reports Server (NTRS)

    Jacob, G.; Costa, F.; Shannon, J.; Robertson, D.; Biaggioni, I.

    2000-01-01

    Sympathetic activation produced by various stimuli, eg, mental stress or handgrip, evokes regional vascular responses that are often nonhomogeneous. This phenomenon is believed to be the consequence of the recruitment of differential central neural pathways or of a sympathetically mediated vasodilation. The purpose of this study was to determine whether a similar heterogeneous response occurs with cold pressor stimulation and to test the hypothesis that local differences in adrenergic receptor function could be in part responsible for this diversity. In 8 healthy subjects, local norepinephrine spillover and blood flow were measured in arms and legs at baseline and during sympathetic stimulation induced by baroreflex mechanisms (nitroprusside infusion) or cold pressor stimulation. At baseline, legs had higher vascular resistance (27+/-5 versus 17+/-2 U, P=0.05) despite lower norepinephrine spillover (0.28+/-0.04 versus 0.4+/-0.05 mg. min(-1). dL(-1), P=0.03). Norepinephrine spillover increased similarly in both arms and legs during nitroprusside infusion and cold pressor stimulation. On the other hand, during cold stimulation, vascular resistance increased in arms but not in legs (20+/-9% versus -7+/-4%, P=0.03). Increasing doses of isoproterenol and phenylephrine were infused intra-arterially in arms and legs to estimate beta-mediated vasodilation and alpha-induced vasoconstriction, respectively. beta-Mediated vasodilation was significantly lower in legs compared with arms. Thus, we report a dissociation between norepinephrine spillover and vascular responses to cold stress in lower limbs characterized by a paradoxical decrease in local resistance despite increases in sympathetic activity. The differences observed in adrenergic receptor responses cannot explain this phenomenon.

  3. Dissociation between neural and vascular responses to sympathetic stimulation : contribution of local adrenergic receptor function

    NASA Technical Reports Server (NTRS)

    Jacob, G.; Costa, F.; Shannon, J.; Robertson, D.; Biaggioni, I.

    2000-01-01

    Sympathetic activation produced by various stimuli, eg, mental stress or handgrip, evokes regional vascular responses that are often nonhomogeneous. This phenomenon is believed to be the consequence of the recruitment of differential central neural pathways or of a sympathetically mediated vasodilation. The purpose of this study was to determine whether a similar heterogeneous response occurs with cold pressor stimulation and to test the hypothesis that local differences in adrenergic receptor function could be in part responsible for this diversity. In 8 healthy subjects, local norepinephrine spillover and blood flow were measured in arms and legs at baseline and during sympathetic stimulation induced by baroreflex mechanisms (nitroprusside infusion) or cold pressor stimulation. At baseline, legs had higher vascular resistance (27+/-5 versus 17+/-2 U, P=0.05) despite lower norepinephrine spillover (0.28+/-0.04 versus 0.4+/-0.05 mg. min(-1). dL(-1), P=0.03). Norepinephrine spillover increased similarly in both arms and legs during nitroprusside infusion and cold pressor stimulation. On the other hand, during cold stimulation, vascular resistance increased in arms but not in legs (20+/-9% versus -7+/-4%, P=0.03). Increasing doses of isoproterenol and phenylephrine were infused intra-arterially in arms and legs to estimate beta-mediated vasodilation and alpha-induced vasoconstriction, respectively. beta-Mediated vasodilation was significantly lower in legs compared with arms. Thus, we report a dissociation between norepinephrine spillover and vascular responses to cold stress in lower limbs characterized by a paradoxical decrease in local resistance despite increases in sympathetic activity. The differences observed in adrenergic receptor responses cannot explain this phenomenon.

  4. Stimulation of pulmonary rapidly adapting receptors by inhaled wood smoke in rats

    PubMed Central

    Lai, C J; Kou, Y R

    1998-01-01

    The stimulation of pulmonary rapidly adapting receptors (RARs) by wood smoke was investigated. Impulses from seventy RARs were recorded in fifty-nine anaesthetized, open-chest and artificially ventilated rats; responses to delivery of 6 ml of wood smoke into the lungs were studied in sixty-one receptors whereas responses to histamine (10 or 100 μg kg−1, i.v.) were studied in the other nine. Delivery of wood smoke stimulated fifty-two of the sixty-one RARs studied. When stimulated, an intense burst of discharge was evoked within 1 or 2 s of smoke delivery. This increased activity quickly peaked in 1-3 s (Δ= 15.8 ± 1.6 impulses s−1; n = 61; mean ± s.e.m.), then declined and yet remained at a level higher than the baseline activity. The mean duration of the stimulation was 25.1 ± 2.7 s. In contrast, smoke delivery did not affect tracheal pressure. Peak responses of RARs to wood smoke were partially reduced by removal of smoke particulates and were largely attenuated by pretreatment with dimethylthiourea (DMTU, a hydroxyl radical scavenger), indomethacin (Indo, a cyclo-oxygenase inhibitor), or both DMTU and Indo (DMTU + Indo). Conversely, the peak responses of RARs were not significantly affected by pretreatment with isoprenaline (a bronchodilator) or vehicle for these chemicals. Additionally, pretreatment with DMTU, Indo, or DMTU + Indo did not significantly alter the RAR sensitivity to mechanical stimulation (constant-pressure lung inflation; 20 cmH2O). Of the nine RARs tested, six were stimulated by histamine and their sensitivity to this chemical irritant was not altered by pretreatment with DMTU + Indo. The results suggest that both the particulates and gas phases are responsible for, and both the hydroxyl radical and cyclo-oxygenase products are involved in, the stimulation of RARs by wood smoke. Furthermore, changes in lung mechanics following smoke delivery are not the cause of this afferent stimulation. PMID:9508820

  5. Chemical inhibitors of c-Met receptor tyrosine kinase stimulate osteoblast differentiation and bone regeneration.

    PubMed

    Kim, Jung-Woo; Nam Lee, Mi; Jeong, Byung-Chul; Oh, Sin-Hye; Kook, Min-Suk; Koh, Jeong-Tae

    2017-03-16

    The c-Met receptor tyrosine kinase and its ligand, hepatocyte growth factor (HGF), have been recently introduced to negatively regulate bone morphogenetic protein (BMP)-induced osteogenesis. However, the effect of chemical inhibitors of c-Met receptor on osteoblast differentiation process has not been examined, especially the applicability of c-Met chemical inhibitors on in vivo bone regeneration. In this study, we demonstrated that chemical inhibitors of c-Met receptor tyrosine kinase, SYN1143 and SGX523, could potentiate the differentiation of precursor cells to osteoblasts and stimulate regeneration in calvarial bone defects of mice. Treatment with SYN1143 or SGX523 inhibited HGF-induced c-Met phosphorylation in MC3T3-E1 and C3H10T1/2 cells. Cell proliferation of MC3T3-E1 or C3H10T1/2 was not significantly affected by the concentrations of these inhibitors. Co-treatment with chemical inhibitor of c-Met and osteogenic inducing media enhanced osteoblast-specific genes expression and calcium nodule formation accompanied by increased Runx2 expression via c-Met receptor-dependent but Erk-Smad signaling independent pathway. Notably, the administration of these c-Met inhibitors significantly repaired critical-sized calvarial bone defects. Collectively, our results suggest that chemical inhibitors of c-Met receptor tyrosine kinase might be used as novel therapeutics to induce bone regeneration.

  6. Interaction of urokinase with specific receptors stimulates mobilization of bovine adrenal capillary endothelial cells

    SciTech Connect

    Fibbi, G.; Ziche, M.; Morbidelli, L. ); Magnelli, L.; Del Rosso, M. )

    1988-12-01

    On the basis of {sup 125}I-labeled plasminogen activator binding analysis the authors have found that bovine adrenal capillary endothelial cells have specific receptors for human urinary-type plasminogen activator on the cell membrane. Each cell exposes about 37,000 free receptors with a K{sub d} of 0.8958{times}10{sup {minus}12} M. A monoclonal antibody against the 17,500 proteolytic fragment of the A chain of the plasminogen activator, not containing the catalytic site of the enzyme, impaired the specific binding, thus suggesting the involvement of a sequence present on the A chain in the interaction with the receptor, as previously shown in other cell model systems. Both the native molecule and the A chain are able to stimulate endothelial cell motility in the Boyden chamber, when used at nanomolar concentrations. The use of the same monoclonal antibody that can inhibit ligand-receptor interaction can impair the plasminogen activator and A-chain-induced endothelial cell motility, suggesting that under the conditions used in this in vitro model system, the motility of bovine adrenal capillary endothelial cells depends on the specific interaction of the ligand with free receptors on the surface of endothelial cells.

  7. 5-HT₄ receptor stimulation leads to soluble AβPPα production through MMP-9 upregulation.

    PubMed

    Hashimoto, Gakuji; Sakurai, Mikako; Teich, Andrew F; Saeed, Faisal; Aziz, Fahad; Arancio, Ottavio

    2012-01-01

    Serotonin 4 (5-HT4) receptor signaling does not only have the physiological function of improving cognition, but might also be helpful in the therapy of Alzheimer's disease (AD) through regulation of the production of soluble amyloid-β protein precursor alpha (sAβPPα). To analyze the relationship between 5-HT4 receptor signaling and sAβPPα production, we stably transfected H4 cells with AβPP and 5-HT4 receptor (H4/AβPP/5-HT4 cells). We found that 24-h incubation with the 5-HT4 receptor agonist RS-67333 upregulates matrix metalloproteinase-9 (MMP-9). Furthermore, MMP-9 overexpression enhanced sAβPPα levels, whereas knockdown with MMP-9 siRNA decreased sAβPPα levels. When RS-67333 was injected for 10 days in Tg2576 mice, a model of amyloid-β peptide (Aβ) deposition, there was an increase in hippocampal levels of sAβPPα, C-terminal fragment α, and MMP-9, as well as a decrease in hippocampal senile plaque number and levels of the 40 amino acid peptide, Aβ40. Taken all together, these experiments demonstrate that 5-HT4 receptor stimulation induces expression of MMP-9 which cleaves AβPP through α-secretase-like activity, leading to an increase of sAβPPα levels and a reduction of Aβ load.

  8. Excitatory amino acid receptor-stimulated phosphoinositide turnover in primary cerebrocortical cultures.

    PubMed Central

    Birrell, G. J.; Marcoux, F. W.

    1993-01-01

    1. Characterization of excitatory amino acid-induced accumulation of [3H]-phosphoinositides was carried out in primary cerebrocortical cultures isolated from foetal rats. 2. All of the excitatory amino acid receptor agonists examined caused concentration-dependent enhancement of phosphoinositide (PI) formation. The most potent excitatory amino acid receptor agonists were quisqualate, (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid ((1S,3R)-ACPD), ibotenate and glutamate with mean EC50 values of 0.9 +/- 0.4 microM, 15 +/- 5 microM, 15 +/- 3 microM and 41 +/- 8 microM respectively. 3. The selective ionotropic receptor antagonists kynurenic acid (1 mM), 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline (NBQX, 10 microM) and (+/-)-4-(3-phosphonopropyl)-2 piperazinecarboxylic acid (CPP, 100 microM), failed to block responses to quisqualate, (1S,3R)-ACPD or glutamate. D,L-2-Amino-3-phosphonopropionate (D,L-AP3) did not block 1S,3R-ACPD or quisqualate-induced PI turnover, but had an additive effect with quisqualate or (1S,3R)-ACPD. 4. Exposure of cultures to agonists in the absence of added extracellular calcium reduced the maximal quisqualate response by approximately 45%, revealing a two-component concentration-response curve. Concentration-response curves to ibotenate and glutamate became flattened by omission of extracellular calcium, whereas (1S,3R)-ACPD-stimulated PI turnover was unaffected. 5. Pretreatment of cultures with pertussis toxin markedly inhibited PI responses evoked by (1S,3R)-ACPD. 6. These results suggest that excitatory amino acid-stimulated PI turnover in cerebrocortical cultures is independent of ionotropic receptor activation and is mediated via specific G-protein-linked metabotropic receptors.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8395285

  9. Inhibition of atrial receptor-induced renal responses by stimulation of carotid baroreceptors in anaesthetized dogs.

    PubMed Central

    Karim, F; Majid, D S

    1991-01-01

    1. Dogs were anaesthetized with chloralose and artificially ventilated. The receptors at three pulmonary vein-atrial junctions and in the left atrial appendage were stimulated by distension of small balloons. The carotid sinuses were vascularly isolated and perfused with arterial blood. A volume reservoir was connected to the aorta via the common carotid and femoral arteries to keep the mean aortic pressure constant (78.8 +/- 2.9 mmHg at low and 87.1 +/- 4.3 mmHg at high carotid sinus pressure, CSP). Propranolol and atropine were infused (i.v.) at 17 and 13 micrograms kg-1 min-1 respectively in order to block beta-adrenergic and cholinergic receptor activities. The renal blood flow was measured by an electromagnetic flow meter (wrap-round probe), glomerular filtration rate by creatinine clearance, urinary sodium excretion by flame photometry and osmolar excretion by osmometry. 2. In twelve tests in eight dogs, stimulation of the left atrial receptors for 13 min, at a mean CSP of 68.6 +/- 2.3 mmHg, resulted in significant increases in renal blood flow from 216 +/- 20.0 to 230 +/- 22.1 ml min-1 (100 g renal mass)-1 (P less than 0.005), glomerular filtration rate from 33.9 +/- 3.2 to 42.1 +/- 4.1 ml min-1 100 g-1 (P less than 0.005), filtration fraction from 0.23 +/- 0.02 to 0.26 +/- 0.02 (P less than 0.005), urine flow rate from 0.21 +/- 0.03 to 0.26 +/- 0.03 ml min-1 100 g-1 (P less than 0.001), sodium excretion from 12.9 +/- 4.0 to 16.4 +/- 4.8 mumol min-1 100 g-1 (P less than 0.01), osmolar excretion from 196 +/- 27.8 to 246 +/- 32.9 muosmol min-1 100 g-1 (P less than 0.005), whilst free water clearance decreased from -0.39 +/- 0.07 to -0.50 +/- 0.09 ml min-1 100 g-1 (P less than 0.005). However, the fractional excretion of sodium did not change. 3. In nine tests in seven dogs, stimulation of the left atrial receptors at a constantly high CSP (161 +/- 11.3 mmHg) did not produce significant change in any of the renal variables. 4. The results show that high level

  10. DC electric stimulation upregulates angiogenic factors in endothelial cells through activation of VEGF receptors

    PubMed Central

    Bai, Huai; Forrester, John V.; Zhao, Min

    2015-01-01

    Small direct current (DC) electric fields direct some important angiogenic responses of vascular endothelial cells. Those responses indicate promising use of electric fields to modulate angiogenesis. We sought to determine the regulation of electric fields on transcription and expression of a serial of import angiogenic factors by endothelial cells themselves. Using semi-quantitative PCR and ELISA we found that electric stimulation upregulates the levels of mRNAs and proteins of a number of angiogenic proteins, most importantly VEGF165, VEGF121 and IL-8 in human endothelial cells. The up-regulation of mRNA levels might be specific, as the mRNA encoding bFGF, TGF-beta and eNOS are not affected by DC electric stimulation at 24 h time-point. Inhibition of VEGF receptor (VEGFR1 or VEGFR2) signaling significantly decreased VEGF production and completely abolished IL-8 production. DC electric stimulation selectively regulates production of some growth factors and cytokines important for angiogenesis through a feed-back loop mediated by VEGF receptors. PMID:21524919

  11. Preselection Thymocytes Are More Sensitive to T Cell Receptor Stimulation Than Mature T Cells

    PubMed Central

    Davey, Gayle M.; Schober, Sonya L.; Endrizzi, Bart T.; Dutcher, Angela K.; Jameson, Stephen C.; Hogquist, Kristin A.

    1998-01-01

    During T cell development, thymocytes which are tolerant to self-peptides but reactive to foreign peptides are selected. The current model for thymocyte selection proposes that self-peptide–major histocompatibility complex (MHC) complexes that bind the T cell receptor with low affinity will promote positive selection while those with high affinity will result in negative selection. Upon thymocyte maturation, such low affinity self-peptide–MHC ligands no longer provoke a response, but foreign peptides can incidentally be high affinity ligands and can therefore stimulate T cells. For this model to work, thymocytes must be more sensitive to ligand than mature T cells. Contrary to this expectation, several groups have shown that thymocytes are less responsive than mature T cells to anti-T cell receptor for antigen (TCR)/CD3 mAb stimulation. Additionally, the lower TCR levels on thymocytes, compared with T cells, would potentially correlate with decreased thymocyte sensitivity. Here we compared preselection thymocytes and mature T cells for early activation events in response to peptide–MHC ligands. Remarkably, the preselection thymocytes were more responsive than mature T cells when stimulated with low affinity peptide variants, while both populations responded equally well to the antigenic peptide. This directly demonstrates the increased sensitivity of thymocytes compared with T cells for TCR engagement by peptide–MHC complexes. PMID:9815264

  12. Uveal melanocytes do not respond to or express receptors for alpha-melanocyte-stimulating hormone.

    PubMed

    Li, Li; Hu, Dan-Ning; Zhao, Huiquan; McCormick, Steven A; Nordlund, James J; Boissy, Raymond E

    2006-10-01

    Whereas cutaneous pigmentation increases after exposure to ultraviolet (UV) irradiation, ocular pigmentation does not. This study was designed to examine the evidence that alpha-melanocyte-stimulating hormone (alpha-MSH), which is thought to be the mediator of UV response in the skin, has any role to play in uveal melanocytes. Human uveal melanocytes derived from the choroid and the iris were cultivated by using eyes harvested from adult cadaveric donors and were assessed by Northern blot analysis for growth and melanogenic response to alpha-MSH and expression of the receptor for alpha-MSH (MC1-R). In addition, expression of alpha-MSH was evaluated in ocular tissue by immunocytochemistry. Uveal melanocytes, unlike cutaneous melanocytes in vitro, exhibited no stimulation of proliferation in response to alpha-MSH at dosages ranging from 0.1 to 100 muM. In addition, tyrosine hydroxylase, DOPA oxidase, and protein levels for tyrosinase, TRP-1, and TRP-2 were not influenced by alpha-MSH. Associated with the lack of alpha-MSH response in cultured uveal melanocytes was the absence of expression of the receptor for alpha-MSH (MC1-R), as assessed by Northern blot analysis. Also in contrast to the skin, pigmented ocular tissue lacked expression of the alpha-MSH ligand, as assessed by immunocytochemistry. In conclusion, ocular pigmentation does not appear to be regulated by melanocyte stimulating hormone.

  13. DC electric stimulation upregulates angiogenic factors in endothelial cells through activation of VEGF receptors.

    PubMed

    Bai, Huai; Forrester, John V; Zhao, Min

    2011-07-01

    Small direct current (DC) electric fields direct some important angiogenic responses of vascular endothelial cells. Those responses indicate promising use of electric fields to modulate angiogenesis. We sought to determine the regulation of electric fields on transcription and expression of a serial of import angiogenic factors by endothelial cells themselves. Using semi-quantitative PCR and ELISA we found that electric stimulation upregulates the levels of mRNAs and proteins of a number of angiogenic proteins, most importantly VEGF165, VEGF121 and IL-8 in human endothelial cells. The up-regulation of mRNA levels might be specific, as the mRNA encoding bFGF, TGF-beta and eNOS are not affected by DC electric stimulation at 24h time-point. Inhibition of VEGF receptor (VEGFR1 or VEGFR2) signaling significantly decreased VEGF production and completely abolished IL-8 production. DC electric stimulation selectively regulates production of some growth factors and cytokines important for angiogenesis through a feed-back loop mediated by VEGF receptors.

  14. Enterococcus faecium stimulates human neutrophils via the formyl-peptide receptor 2.

    PubMed

    Bloes, Dominik Alexander; Otto, Michael; Peschel, Andreas; Kretschmer, Dorothee

    2012-01-01

    The human formyl-peptide receptor 2 (FPR2/ALX) senses phenol-soluble modulin (PSM) peptide toxins produced by pathogenic staphylococcal species and plays a crucial role in directing neutrophil influx during staphylococcal infection. However, it has remained unclear if FPR2 responds also to molecules from other bacterial pathogens. Here we analyzed a variety of gram-positive and gram-negative pathogens and found that apart from staphylococci only certain enterococcal strains have the capacity to stimulate FPR2/ALX. Most of the analyzed Enterococcus faecium but only sporadic Enterococcus faecalis strains released FPR2/ALX-stimulating molecules leading to neutrophil calcium ion fluxes, chemotaxis, and complement receptor upregulation. Among ten test strains vancomycin-resistant E. faecium had a significantly higher capacity to stimulate FPR2/ALX than vancomycin-susceptible strains, suggesting an association of strong FPR2/ALX activation with health-care associated strains. The enterococcal FPR2/ALX agonists were found to be peptides or proteins, which appear, however, to be unrelated to staphylococcal PSMs in sequence and physicochemical properties. Enterococci are among the most frequent invasive bacterial pathogens but the basis of enterococcal virulence and immune activation has remained incompletely understood. Our study indicates that previously unrecognized proteinaceous agonists contribute to Enterococcus-host interaction and underscores the importance of FPR2/ALX in host defense against major endogenous bacterial pathogens.

  15. Studies on the structure of the follicle-stimulating hormone receptor using photoaffinity labeling procedures

    SciTech Connect

    Smith, R.A.

    1985-01-01

    The general objective of this project was to study the structure of the follicle stimulating hormone (FSH) receptor using affinity labeling methods. A low density fraction derived from homogenates of bovine testis was found to contain high affinity and low capacity receptors specific for FSH. Electron microscopic examination of the fraction revealed structure resembling multilamellar membranous vesicles (MV). For photoaffinity labeling of the FSH receptors in MV, an azidobenzoyl-/sup 125/I-analog of human FSH was prepared (/sup 125/I-AB-hFSH) and binding of specific FSH receptors was studied. /sup 125/I-AB-hFSH binding of receptors was inhibited in a dose dependent manner by unlabeled hFSH, and binding was not prevented by structurally-related human chorionic gonadotropin (hCG). The formation of photocrosslinked protein of relative molecular mass (M/sub r/) 54,000, 64,000, 76,000, 84,000, 97,000 and 116,000 was found to be inhibited by unlabeled hFSH in a dose related manner, and to be dependent on photoactivation of the FSH derivative. The interpretation of the photoaffinity labeling experiments was that three proteins associated with the FSH receptor were photoaffinity labeled. Analysis by indirect means suggested that the three proteins were assembled to form oligomeric complexes, and based on the intensities and composition of the oligomeric species, spatial relationships of the polypeptides with respect to each other on the membrane surface were deduced. The results of photoaffinity labeling suggest the FSH receptor is composed of three subunits of M/sub r/ 38,000, 48,000, and 81,000 and exists in the membrane in part as a M/sub r/ 330,000 dimer.

  16. Amphetamine decreases behavioral inhibition by stimulation of dopamine D2, but not D3, receptors.

    PubMed

    van Gaalen, Marcel M; Unger, Liliane; Jongen-Rêlo, Ana-Lucia; Schoemaker, Hans; Gross, Gerhard

    2009-09-01

    Behavioral disinhibition is a manifestation of impulsive behavior that is prominent in the psychopathology of various psychiatric disorders such as addiction, attention-deficit hyperactivity disorder, mania, and personality disorders. Impulsivity may be studied by measuring anticipatory responses made before the presentation of a food-predictive, brief light stimulus in a two-choice serial reaction time task. In such serial reaction time tasks, amphetamine has been shown to produce dose-dependent increases in premature responding in a manner dependent on dopamine D(2)-like receptor stimulation. So far, it is unknown whether it is the D(2) or D(3) receptor that is involved in this form of impulsivity. In this study, rats were trained in a two-choice serial reaction time task until baseline performance was stable. Next, effects of the dopamine D(2) preferring antagonist L-741,626 and selective D(3) antagonist SB-277011 were assessed alone and in the presence of amphetamine. Neither L-741,626 nor SB-277011 affected behavioral inhibition, although the latter significantly increased reaction time at 10 mg/kg. Amphetamine dose-dependently increased impulsivity. The effect of amphetamine was attenuated by L-741,626 (3 mg/kg), whereas SB-277011 (3 mg/kg) had no effect. Therefore, amphetamine-induced behavioral disinhibition depends on D(2), but not D(3), receptor stimulation.

  17. Stimulation of prolactin receptor induces STAT-5 phosphorylation and cellular invasion in glioblastoma multiforme

    PubMed Central

    Alkharusi, Amira; Yu, Shengze; Landázuri, Natalia; Zadjali, Fahad; Davodi, Belghis; Nyström, Thomas; Gräslund, Torbjörn; Rahbar, Afsar; Norstedt, Gunnar

    2016-01-01

    Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor in humans and is characterized with poor outcome. In this study, we investigated components of prolactin (Prl) system in cell models of GBM and in histological tissue sections obtained from GBM patients. Expression of Prolactin receptor (PrlR) was detected at high levels in U251-MG, at low levels in U87-MG and barely detectable in U373 cell lines and in 66% of brain tumor tissues from 32 GBM patients by immunohistochemical technique. In addition, stimulation of U251-MG and U87-MG cells but not U373 with Prl resulted in increased STAT5 phosphorylation and only in U251-MG cells with increased cellular invasion. Furthermore, STAT5 phosphorylation and cellular invasion induced in Prl stimulated cells were significantly reduced by using a Prl receptor antagonist that consists of Prl with four amino acid replacements. We conclude that Prl receptor is expressed at different levels in the majority of GBM tumors and that blocking of PrlR in U251-MG cells significantly reduce cellular invasion. PMID:27788487

  18. Activation of the GABA(B) Receptor Prevents Nicotine-Induced Locomotor Stimulation in Mice.

    PubMed

    Lobina, Carla; Carai, Mauro A M; Froestl, Wolfgang; Mugnaini, Claudia; Pasquini, Serena; Corelli, Federico; Gessa, Gian Luigi; Colombo, Giancarlo

    2011-01-01

    Recent studies demonstrated that activation of the GABA(B) receptor, either by means of orthosteric agonists or positive allosteric modulators (PAMs), inhibited different nicotine-related behaviors, including intravenous self-administration and conditioned place preference, in rodents. The present study investigated whether the anti-nicotine effects of the GABA(B) receptor agonist, baclofen, and GABA(B) PAMs, CGP7930, and GS39783, extend to nicotine stimulant effects. To this end, CD1 mice were initially treated with baclofen (0, 1.25, and 2.5 mg/kg, i.p.), CGP7930 (0, 25, and 50 mg/kg, i.g.), or GS39783 (0, 25, and 50 mg/kg, i.g.), then treated with nicotine (0 and 0.05 mg/kg, s.c.), and finally exposed to an automated apparatus for recording of locomotor activity. Pretreatment with doses of baclofen, CGP7930, or GS39783 that did not alter locomotor activity when given with nicotine vehicle fully prevented hyperlocomotion induced by 0.05 mg/kg nicotine. These data extend to nicotine stimulant effects the capacity of baclofen and GABA(B) PAMs to block the reinforcing, motivational, and rewarding properties of nicotine. These data strengthen the hypothesis that activation of the GABA(B) receptor may represent a potentially useful, anti-smoking therapeutic strategy.

  19. Hormone stimulation of androgen receptor mediates dynamic changes in DNA methylation patterns at regulatory elements

    PubMed Central

    Dhiman, Vineet K.; Attwood, Kristopher; Campbell, Moray J.; Smiraglia, Dominic J.

    2015-01-01

    DNA methylation is an epigenetic modification that contributes to stable gene silencing by interfering with the ability of transcriptional regulators to bind to DNA. Recent findings have revealed that hormone stimulation of certain nuclear receptors induces rapid, dynamic changes in DNA methylation patterns alongside transcriptional responses at a subset of target loci, over time. However, the ability of androgen receptor (AR) to dynamically regulate gene transcription is relatively under-studied and its role in the regulation of DNA methylation patterns remains to be elucidated. Here we demonstrate in normal prostate cells that hormone stimulated AR activity results in dynamic changes in the transcription rate and DNA methylation patterns at the AR target genes, TIPARP and SGK1. Time-resolved chromatin immunoprecipitation experiments on the SGK1 locus reveals dynamic recruitment of AR and RNA Polymerase II, as well as the recruitment of proteins involved in the DNA demethylation process, TET1 and TDG. Furthermore, the presence of DNA methylation at dynamic regions inhibits protein binding and transcriptional activity of SGK1. These findings establish AR activity as a contributing factor to the dynamic regulation of DNA methylation patterns at target genes in prostate biology and infer further complexity involved in nuclear receptor mediation of transcriptional regulation. PMID:26646795

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

    SciTech Connect

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

    1987-09-01

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

  1. Immunoglobulin-like domain containing receptor 1 mediates fat-stimulated cholecystokinin secretion

    PubMed Central

    Chandra, Rashmi; Wang, Yu; Shahid, Rafiq A.; Vigna, Steven R.; Freedman, Neil J.; Liddle, Rodger A.

    2013-01-01

    Cholecystokinin (CCK) is a satiety hormone produced by discrete enteroendocrine cells scattered among absorptive cells of the small intestine. CCK is released into blood following a meal; however, the mechanisms inducing hormone secretion are largely unknown. Ingested fat is the major stimulant of CCK secretion. We recently identified a novel member of the lipoprotein remnant receptor family known as immunoglobulin-like domain containing receptor 1 (ILDR1) in intestinal CCK cells and postulated that this receptor conveyed the signal for fat-stimulated CCK secretion. In the intestine, ILDR1 is expressed exclusively in CCK cells. Orogastric administration of fatty acids elevated blood levels of CCK in wild-type mice but not Ildr1-deficient mice, although the CCK secretory response to trypsin inhibitor was retained. The uptake of fluorescently labeled lipoproteins in ILDR1-transfected CHO cells and release of CCK from isolated intestinal cells required a unique combination of fatty acid plus HDL. CCK secretion secondary to ILDR1 activation was associated with increased [Ca2+]i, consistent with regulated hormone release. These findings demonstrate that ILDR1 regulates CCK release through a mechanism dependent on fatty acids and lipoproteins and that absorbed fatty acids regulate gastrointestinal hormone secretion. PMID:23863714

  2. D1/D5 dopamine receptors stimulate intracellular calcium release in primary cultures of neocortical and hippocampal neurons.

    PubMed

    Lezcano, Nelson; Bergson, Clare

    2002-04-01

    D1/D5 dopamine receptors in basal ganglia, hippocampus, and cerebral cortex modulate motor, reward, and cognitive behavior. Previous work with recombinant proteins revealed that in cells primed with heterologous G(q/11)-coupled G-protein-coupled receptor (GPCR) agonists, the typically G(s)-linked D1/D5 receptors can stimulate robust release of calcium from internal stores when coexpressed with calcyon. To learn more about the intracellular signaling mechanisms underlying these D1/D5 receptor regulated behaviors, we explored the possibility that endogenous receptors stimulate internal release of calcium in neurons. We have identified a population of neurons in primary cultures of hippocampus and neocortex that respond to D1/D5 dopamine receptor agonists with a marked increase in intracellular calcium (Ca) levels. The D1/D5 receptor stimulated responses occurred in the absence of extracellular Ca(2+) indicating the rises in Ca involve release from internal stores. In addition, the responses were blocked by D1/D5 receptor antagonists. Further, the D1/D5 agonist-evoked responses were state dependent, requiring priming with agonists of G(q/11)-coupled glutamate, serotonin, muscarinic, and adrenergic receptors or with high external K(+) solution. In contrast, D1/D5 receptor agonist-evoked Ca(2+) responses were not detected in neurons derived from striatum. However, D1/D5 agonists elevated cAMP levels in striatal cultures as effectively as in neocortical and hippocampal cultures. Further, neither forskolin nor 8-Br-cAMP stimulation following priming was able to mimic the D1/D5 agonist-evoked Ca(2+) response in neocortical neurons indicating that increased cAMP levels are not sufficient to stimulate Ca release. Our data suggest that D1-like dopamine receptors likely modulate neocortical and hippocampal neuronal excitability and synaptic function via Ca(2+) as well as cAMP-dependent signaling.

  3. Nonlinear relationship between alpha 1-adrenergic receptor occupancy and norepinephrine-stimulated calcium flux in cultured vascular smooth muscle cells

    SciTech Connect

    Colucci, W.S.; Brock, T.A.; Gimbrone, M.A. Jr.; Alexander, R.W.

    1985-05-01

    To determine the relationship between vascular alpha 1-adrenergic receptor occupancy and receptor-coupled calcium flux, the authors have studied (/sup 3/H)prazosin binding and l-norepinephrine-induced /sup 45/Ca efflux in cultured vascular smooth muscle cells isolated from the rabbit aorta. In a crude cellular homogenate, (/sup 3/H)prazosin bound to a single high affinity site, whereas l-norepinephrine (NE) binding was best described by a two-site model. NE-stimulated /sup 45/Ca efflux was concentration-dependent (EC/sup 50/ = 108 nM) and potently inhibited by prazosin (IC/sup 50/ = 0.15 nM). For the total receptor pool identified by (/sup 3/H)prazosin binding, the relationship between receptor occupancy by NE and NE-stimulated /sup 45/Ca efflux was markedly nonlinear, such that 50% of maximum NE-stimulated efflux occurred with occupancy of only approximately 7% of receptors. These two experimental approaches provide direct evidence for the presence in cultured rabbit aortic smooth muscle cells of a sizable pool of alpha 1-adrenergic receptors in excess of those needed for maximum NE-stimulated /sup 45/Ca efflux. This evidence of ''spare'' receptors, together with the finding of two affinity states of agonist binding, raises the possibility of functional heterogeneity of alpha 1-adrenergic receptors in this system.

  4. Interaction of the Clostridium difficile Binary Toxin CDT and Its Host Cell Receptor, Lipolysis-stimulated Lipoprotein Receptor (LSR)*

    PubMed Central

    Hemmasi, Sarah; Czulkies, Bernd A.; Schorch, Björn; Veit, Antonia; Aktories, Klaus; Papatheodorou, Panagiotis

    2015-01-01

    CDT (Clostridium difficile transferase) is a binary, actin ADP-ribosylating toxin frequently associated with hypervirulent strains of the human enteric pathogen C. difficile, the most serious cause of antibiotic-associated diarrhea and pseudomembranous colitis. CDT leads to the collapse of the actin cytoskeleton and, eventually, to cell death. Low doses of CDT result in the formation of microtubule-based protrusions on the cell surface that increase the adherence and colonization of C. difficile. The lipolysis-stimulated lipoprotein receptor (LSR) is the host cell receptor for CDT, and our aim was to gain a deeper insight into the interplay between both proteins. We show that CDT interacts with the extracellular, Ig-like domain of LSR with an affinity in the nanomolar range. We identified LSR splice variants in the colon carcinoma cell line HCT116 and disrupted the LSR gene in these cells by applying the CRISPR-Cas9 technology. LSR truncations ectopically expressed in LSR knock-out cells indicated that intracellular parts of LSR are not essential for plasma membrane targeting of the receptor and cellular uptake of CDT. By generating a series of N- and C-terminal truncations of the binding component of CDT (CDTb), we found that amino acids 757–866 of CDTb are sufficient for binding to LSR. With a transposon-based, random mutagenesis approach, we identified potential LSR-interacting epitopes in CDTb. This study increases our understanding about the interaction between CDT and its receptor LSR, which is key to the development of anti-toxin strategies for preventing cell entry of the toxin. PMID:25882847

  5. Interaction of the Clostridium difficile Binary Toxin CDT and Its Host Cell Receptor, Lipolysis-stimulated Lipoprotein Receptor (LSR).

    PubMed

    Hemmasi, Sarah; Czulkies, Bernd A; Schorch, Björn; Veit, Antonia; Aktories, Klaus; Papatheodorou, Panagiotis

    2015-05-29

    CDT (Clostridium difficile transferase) is a binary, actin ADP-ribosylating toxin frequently associated with hypervirulent strains of the human enteric pathogen C. difficile, the most serious cause of antibiotic-associated diarrhea and pseudomembranous colitis. CDT leads to the collapse of the actin cytoskeleton and, eventually, to cell death. Low doses of CDT result in the formation of microtubule-based protrusions on the cell surface that increase the adherence and colonization of C. difficile. The lipolysis-stimulated lipoprotein receptor (LSR) is the host cell receptor for CDT, and our aim was to gain a deeper insight into the interplay between both proteins. We show that CDT interacts with the extracellular, Ig-like domain of LSR with an affinity in the nanomolar range. We identified LSR splice variants in the colon carcinoma cell line HCT116 and disrupted the LSR gene in these cells by applying the CRISPR-Cas9 technology. LSR truncations ectopically expressed in LSR knock-out cells indicated that intracellular parts of LSR are not essential for plasma membrane targeting of the receptor and cellular uptake of CDT. By generating a series of N- and C-terminal truncations of the binding component of CDT (CDTb), we found that amino acids 757-866 of CDTb are sufficient for binding to LSR. With a transposon-based, random mutagenesis approach, we identified potential LSR-interacting epitopes in CDTb. This study increases our understanding about the interaction between CDT and its receptor LSR, which is key to the development of anti-toxin strategies for preventing cell entry of the toxin. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. The octadecaneuropeptide ODN stimulates neurosteroid biosynthesis through activation of central-type benzodiazepine receptors.

    PubMed

    Do-Rego, J L; Mensah-Nyagan, A G; Beaujean, D; Leprince, J; Tonon, M C; Luu-The, V; Pelletier, G; Vaudry, H

    2001-01-01

    Neurosteroids may play a major role in the regulation of various neurophysiological and behavioural processes. However, while the biochemical pathways involved in the synthesis of neuroactive steroids in the central nervous system are now elucidated, the mechanisms controlling the activity of neurosteroid-producing cells remain almost completely unknown. In the present study, we have investigated the effect of the octadecaneuropeptide (ODN), an endogenous ligand of benzodiazepine receptors, in the control of steroid biosynthesis in the frog hypothalamus. Glial cells containing ODN-like immunoreactivity were found to send their thick processes in the close vicinity of neurones expressing the steroidogenic enzyme 3 beta-hydroxysteroid dehydrogenase. Exposure of frog hypothalamic explants to graded concentrations of ODN (10(-10)-10(-5) M) produced a dose-dependent increase in the conversion of tritiated pregnenolone into various radioactive steroids, including 17-hydroxypregnenolone, progesterone, 17-hydroxyprogesterone, dehydroepiandrosterone and dihydrotestosterone. The ODN-induced stimulation of neurosteroid biosynthesis was mimicked by the central-type benzodiazepine receptor (CBR) inverse agonists methyl beta-carboline-3-carboxylate (beta-CCM) and methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM). The stimulatory effects of ODN, beta-CCM and DMCM on steroid formation was markedly reduced by the CBR antagonist flumazenil. The ODN-evoked stimulation of neurosteroid production was also significantly attenuated by GABA. Collectively, these data indicate that the endozepine ODN, released by glial cell processes in the vicinity of 3 beta-hydroxysteroid dehydrogenase-containing neurones, stimulates the biosynthesis of neurosteroids through activation of central-type benzodiazepines receptors.

  7. The "TSH Receptor Glo Assay" - A High-Throughput Detection System for Thyroid Stimulation.

    PubMed

    Latif, Rauf; Lau, Zerlina; Cheung, Pamela; Felsenfeld, Dan P; Davies, Terry F

    2016-01-01

    To identify novel small molecules against the TSH receptor, we developed a sensitive transcription-based luciferase high-throughput screening (HTS) system named the TSHR-Glo Assay (TSHR-Glo). This assay uses double-transfected Chinese hamster ovary cells stably expressing the human TSHR and a cAMP-response element (CRE) construct fused to an improved luciferase reporter gene. The assay was highly responsive toward TSH in a dose-dependent manner with a TSH sensitivity of 10(-10)M (10 ± 1.12 μU/ml) and thyroid-stimulating antibodies, a hallmark of Graves' disease, could also be detected. The assay was validated against the standard indicator of HTS performance - the Z-factor (Z') - producing a score of 0.895. Using the TSHR-Glo assay, we screened 48,224 compounds from a diverse chemical library in duplicate plates at a fixed dose of 17 μM. Twenty molecules with the greatest activity out of 62 molecules that were identified by this technique were subsequently screened against the parent luciferase stable cell line in order to eliminate false positive stimulators. Using this approach, we were able to identify specific agonists against the TSH receptor leading to the characterization of several TSH agonist molecules. Hence, the TSHR-Glo assay was a one-step cell-based HTS assay, which was successful in the discovery of novel small molecular agonists and for the detection of stimulating antibodies to the TSH receptor.

  8. FSH stimulates lipid biosynthesis in chicken adipose tissue by upregulating the expression of its receptor FSHR.

    PubMed

    Cui, Huanxian; Zhao, Guiping; Liu, Ranran; Zheng, Maiqing; Chen, Jilan; Wen, Jie

    2012-05-01

    Transcripts and protein for follicle-stimulating hormone receptor (FSHR) were demonstrated in abdominal adipose tissue of female chickens. There was no expression of the Fsh gene, but FSH and FSHR colocalized, suggesting that FSH was receptor bound. Partial correlations indicted that changes in abdominal fat (AF) content were most directly correlated with Fshr mRNA expression, and the latter was directly correlated with tissue FSH content. These relationships were consistent with FSH inducing Fshr mRNA expression and with the finding that FSH influenced the accumulation of AF in chickens, a novel role for the hormone. Chicken preadipocytes responded linearly to doubling concentrations of FSH in Fshr mRNA expression and quantities of FSHR and lipid, without discernable effect on proliferation. Cells exposed to FSH more rapidly acquired adipocyte morphology. Treatment of young chickens with chicken FSH (4 mIU/day, subcutaneous, days 7-13) did not significantly decrease live weight but increased AF weight by 54.61%, AF as a percentage of live weight by 55.45%, and FSHR transcripts in AF by 222.15% (2 h after injection). In cells stimulated by FSH, genes related to lipid metabolism, including Rdh10, Dci, RarB, Lpl, Acsl3, and Dgat2, were expressed differentially, compared with no FSH. Several pathways of retinal and fatty acid metabolism, and peroxisome proliferator-activated receptor (PPAR) signaling changed. In conclusion, FSH stimulates lipid biosynthesis by upregulating Fshr mRNA expression in abdominal adipose tissue of chickens. Several genes involved in fatty acid and retinal metabolism and the PPAR signaling pathway mediate this novel function of FSH.

  9. 5-HT2A receptor-stimulated phosphoinositide hydrolysis in the stimulus effects of hallucinogens.

    PubMed

    Rabin, Richard A; Regina, Meredith; Doat, Mireille; Winter, J C

    2002-05-01

    The role of 5-HT2A-mediated stimulation of phosphoinositide hydrolysis in the discriminative effects of hallucinogens was investigated in PC12 cells stably expressing the rat 5-HT2A receptor (PC12-5-HT2A cells). The hallucinogenic compounds, D-lysergic acid diethylamide (LSD), (-)2,5-dimethoxy-4-methylamphetamine (DOM), psilocybin, N,N-dimethyltryptamine (DMT), 5-methoxy-N,N-dimethyltryptamine (MDMT) and N,N-diethyltryptamine (DET), all caused a concentration-dependent increase in the generation of [3H]inositol phosphates. The nonhallucinogenic compounds, 6-fluoro-N,N-diethyltryptamine (6-F-DET), lisuride and quipazine, also displayed significant efficacy in stimulating phosphoinositide hydrolysis, while 2-bromo-lysergic acid diethylamide (BOL), which is not a hallucinogen, did not alter inositol phosphate generation. The beta-carbolines, harmaline and harmane, also did not alter phosphoinositide hydrolysis. Comparison of these results with previous drug discrimination studies indicated the apparent lack of correlation between the degree of substitution in LSD- and DOM-trained animals and efficacy in stimulating phosphoinositide hydrolysis. The present study indicates that 5-HT2A-mediated stimulation of phosphoinositide hydrolysis does not appear to be the sole critical signaling mechanism involved in the discriminative effects of hallucinogens.

  10. Caloric restriction stimulates autophagy in rat cortical neurons through neuropeptide Y and ghrelin receptors activation

    PubMed Central

    Carmo-Silva, Sara; Botelho, Mariana; de Almeida, Luís Pereira; Cavadas, Cláudia

    2016-01-01

    Caloric restriction is an anti-aging intervention known to extend lifespan in several experimental models, at least in part, by stimulating autophagy. Caloric restriction increases neuropeptide Y (NPY) in the hypothalamus and plasma ghrelin, a peripheral gut hormone that acts in hypothalamus to modulate energy homeostasis. NPY and ghrelin have been shown to be neuroprotective in different brain areas and to induce several physiological modifications similar to those induced by caloric restriction. However, the effect of NPY and ghrelin in autophagy in cortical neurons is currently not known. Using a cell culture of rat cortical neurons we investigate the involvement of NPY and ghrelin in caloric restriction-induced autophagy. We observed that a caloric restriction mimetic cell culture medium stimulates autophagy in rat cortical neurons and NPY or ghrelin receptor antagonists blocked this effect. On the other hand, exogenous NPY or ghrelin stimulate autophagy in rat cortical neurons. Moreover, NPY mediates the stimulatory effect of ghrelin on autophagy in rat cortical neurons. Since autophagy impairment occurs in aging and age-related neurodegenerative diseases, NPY and ghrelin synergistic effect on autophagy stimulation may suggest a new strategy to delay aging process. PMID:27441412

  11. Adaptation and cross-adaptation to odor stimulation of olfactory receptors in the tiger salamander

    PubMed Central

    1979-01-01

    We have used the effects of self- and cross-adaptation on the unitary responses of olfactory receptors of the tiger salamander to odor stimulation to investigate the stimulus-specific components of these responses and to provide information about the cross-cell variations in the numbers and numbers of types of constitutent receptive sites. An olfactometer delivered sequential odorous pulses, either juxtaposed or separated by a variable time delay. We used four pairs of odorants judged to be similar within a given pair. The unitary response to the test stimulation relative to that of the conditioning stimulation varied from being unchanged to being completely eliminated. We sometimes observed substantial poststimulus increases in the firing rate following stimulation with juxtaposed odorous pulse. Except in the case of one odorant pair, cross-adaptation occurred both with juxtaposed pulses and with pulses separated in time. With the methyl butyrate/ethyl butyrate odorant pair, however, statistically significant cross-adaptation appeared only with juxtaposed pulses. We propose a simple model to aid in explaining these phenomena. The experimental observations in conjunction with this model are used to obtain estimates of the maximal and minimal number of receptive site types available for interaction with the chosen odorants. PMID:479820

  12. Tryptophan Metabolism Along the Kynurenine Pathway Downstream of Toll-like Receptor Stimulation in Peripheral Monocytes.

    PubMed

    Orhan, F; Bhat, M; Sandberg, K; Ståhl, S; Piehl, F; Svensson, C; Erhardt, S; Schwieler, L

    2016-11-01

    Tryptophan degradation along the kynurenine pathway is of central importance for the immune function. Toll-like receptors (TLRs), representing the first line of immune defence against pathogens, are expressed in various cell types. The most abundant expression is found on monocytes, macrophages and dendritic cells. The aim of this study was to investigate whether stimulation with different TLR ligands induces the kynurenine pathway in human peripheral monocytes. Cell supernatants were analysed using a liquid chromatography/mass spectrometry to measure kynurenine, kynurenic acid (KYNA), quinolinic acid (QUIN) and tryptophan. Stimulation of TLR-2, TLR-3, TLR-4, TLR-7/8 and TLR-9 was found to induce the production of kynurenine, but only stimulation of TLR-3 increased levels of further downstream metabolites, such as KYNA and QUIN. Stimulation of TLR-1, TLR-5 and TLR-6 did not induce the kynurenine pathway. Taken together, this study provides novel evidence demonstrating that TLR activation induces a pattern of downstream tryptophan degradation along the kynurenine pathway in monocytes. The results of this study may implicate that TLRs can be used as new drug targets for the regulation of aberrant tryptophan metabolism along this pathway, a potential therapeutic strategy that may be of importance in several disorders. © 2016 The Foundation for the Scandinavian Journal of Immunology.

  13. Caffeine and propranolol block the increase in rat pineal melatonin production produced by stimulation of adenosine receptors.

    PubMed

    Babey, A M; Palmour, R M; Young, S N

    1994-07-18

    The adenosine agonist 5'-N-ethylcarboxamidoadenosine (NECA) injected i.p. during the light period increased rat pineal melatonin levels and this increase was blocked by simultaneous administration of the non-selective adenosine receptor antagonist caffeine. A single dose of the adenosine A1 agonist cyclopentyladenosine had no effect on nocturnal melatonin production. The NECA-stimulated increase was also blocked by the beta-adrenergic receptor antagonist propranolol. Given alone, neither caffeine nor propranolol had any effect on melatonin levels. The results point to an intermediate role for beta-adrenergic receptors in the adenosine-stimulated increase of melatonin production.

  14. Cannabinoid-induced stimulation of motor activity in planaria through an opioid receptor-mediated mechanism.

    PubMed

    Buttarelli, Francesca R; Pontieri, Francesco E; Margotta, Vito; Palladini, Guido

    2002-01-01

    Planaria, the most primitive example of centralization and cephalization of the nervous system along phylogeny, shows specific stereotyped behavioral patterns following exposure to drugs acting on neural transmission. In this study, the authors investigated the effects of exposure to the synthetic cannabinoid receptor agonist WTN55212.2 on motor activity in planaria. WTN55212.2 produced dose-dependent stimulation of motor behavior. High doses of the drug caused stereotyped activities identical to those seen previously with opioid agonists. These effects were antagonized by coexposure to cannabinoid or opioid receptor antagonists. The results indicate that functional interactions between cannabinoid and opioid systems are highly conserved along phylogeny, at least at the behavioral level.

  15. Identification of vertebrate volatiles stimulating olfactory receptors on tarsus I of the tick Amblyomma variegatum Fabricius (Ixodidae). I. Receptors within the Haller's organ capsule.

    PubMed

    Steullet, P; Guerin, P M

    1994-01-01

    Gas chromatography-coupled electrophysiological recordings (GC-EL) from olfactory sensilla within the capsule of Haller's organ of the tick Amblyomma variegatum indicate the presence of a number of stimulants in rabbit and bovine odours, and in steer skin wash. Some of these stimulants were fully identified by gas chromatography-mass spectrometry analysis and by matching electrophysiological activity of synthetic analogues as: 1) hexanal, 2-heptenal, nonanal, furfural, benzaldehyde, and 2-hydroxybenzaldehyde (in all extracts); 2) heptanal, 2-, 3-, and 4-methylbenzaldehyde, and gamma-valerolactone (only in bovine and rabbit odour). Careful examination of the electrophysiological responses permit characterization of 6 receptor types: 1) a benzaldehyde receptor, 2) a 2-hydroxybenzaldehyde receptor, 3) three types of receptors responding differently to aliphatic aldehydes, and 4) a lactone receptor.

  16. Misfolding Ectodomain Mutations of the Lutropin Receptor Increase Efficacy of Hormone Stimulation

    PubMed Central

    Charmandari, E.; Guan, R.; Zhang, M.; Silveira, L. G.; Fan, Q. R.; Chrousos, G. P.; Sertedaki, A. C.; Latronico, A. C.

    2016-01-01

    We demonstrate 2 novel mutations of the LHCGR, each homozygous, in a 46,XY patient with severe Leydig cell hypoplasia. One is a mutation in the signal peptide (p.Gln18_Leu19ins9; referred to here as SP) that results in an alteration of the coding sequence of the N terminus of the mature mutant receptor. The other mutation (p.G71R) is also within the ectodomain. Similar to many other inactivating mutations, the cell surface expression of recombinant human LHR(SP,G71R) is greatly reduced due to intracellular retention. However, we made the unusual discovery that the intrinsic efficacy for agonist-stimulated cAMP in the reduced numbers of receptors on the cell surface was greatly increased relative to the same low number of cell surface wild-type receptor. Remarkably, this appears to be a general attribute of misfolding mutations in the ectodomains, but not serpentine domains, of the gonadotropin receptors. These findings suggest that there must be a common, shared mechanism by which disparate mutations in the ectodomain that cause misfolding and therefore reduced cell surface expression concomitantly confer increased agonist efficacy to those receptor mutants on the cell surface. Our data further suggest that, due to their increased agonist efficacy, extremely small changes in cell surface expression of misfolded ectodomain mutants cause larger than expected alterations in the cellular response to agonist. Therefore, for inactivating LHCGR mutations causing ectodomain misfolding, the numbers of cell surface mutant receptors on fetal Leydig cells of 46,XY individuals exert a more exquisite effect on the relative severity of the clinical phenotypes than already appreciated. PMID:26554443

  17. Tranexamic acid induces kaolin intake stimulating a pathway involving tachykinin neurokinin 1 receptors in rats.

    PubMed

    Kakiuchi, Hitoshi; Kawarai-Shimamura, Asako; Kuwagata, Makiko; Orito, Kensuke

    2014-01-15

    Tranexamic acid suppresses post-partum haemorrhage and idiopathic menorrhagia through its anti-fibrinolytic action. Although it is clinically useful, it is associated with high risks of side effects such as emesis. Understanding the mechanisms underlying tranexamic acid-induced emesis is very important to explore appropriate anti-emetic drugs for the prevention and/or suppression of emesis. In this study, we examined the receptors involved in tranexamic acid-induced kaolin intake in rats, which reflects the drug's clinical emetogenic potential in humans. Further, we examined the brain regions activated by administration of tranexamic acid and elucidated pivotal pathways of tranexamic acid-induced kaolin intake. We examined the effects of ondansetron, a 5-hydroxytryptamine 3 receptor antagonist, domperidone, a dopamine 2 receptor antagonist, and aprepitant, a tachykinin neurokinin 1 (NK1) receptor antagonist, on tranexamic acid-induced kaolin intake in rats. Then, we determined the brain regions that showed increased numbers of c-Fos immunoreactive cells. Finally, we examined the effects of an antagonist(s) that reduced tranexamic acid-induced kaolin intake on the increase in c-Fos immunoreactive cells. Aprepitant significantly decreased tranexamic acid-induced kaolin intake. However, neither ondansetron nor domperidone decreased kaolin intake. Tranexamic acid significantly increased c-Fos immunoreactive cells by approximately 5.5-fold and 22-fold in the area postrema and nucleus of solitary tract, respectively. Aprepitant decreased the number of c-Fos immunoreactive cells in both areas. Tranexamic acid induced kaolin intake possibly via stimulation of tachykinin NK1 receptors in rats. The tachykinin NK1 receptor could be targeted to prevent and/or suppress emesis in patients receiving tranexamic acid.

  18. Involvement of prefrontal AMPA receptors in encounter stimulation-induced hyperactivity in isolation-reared mice.

    PubMed

    Araki, Ryota; Ago, Yukio; Hasebe, Shigeru; Nishiyama, Saki; Tanaka, Tatsunori; Oka, Satoshi; Takuma, Kazuhiro; Matsuda, Toshio

    2014-06-01

    We recently showed that social encounter stimulation induces hyperactivity in mice reared in social isolation from early life and this is associated with the transient activation of prefrontal dopaminergic and serotonergic systems. In the present study, we examined the effect of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor antagonist 2, 3-dioxo-6-nitro-1, 2, 3, 4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide (NBQX) on encounter-induced behavioural and neurochemical changes to study the role of the receptor in abnormal behaviours in isolation-reared mice. The encounter to an intruder mouse induced hyperactivity with transient increases in prefrontal dopamine and serotonin levels in isolation-reared mice. NBQX attenuated the encounter-induced hyperactivity and the associated neurochemical changes in isolation-reared mice. In addition, NBQX reduced aggressive behaviour and cognitive impairment in isolation-reared mice, but did not affect depressive-like behaviour or spontaneous hyper-locomotion in these animals. The AMPA receptor agonist (S)-AMPA increased prefrontal dopamine and serotonin release, and this effect was higher in isolation-reared mice than in the group-reared mice, suggesting higher prefrontal AMPA receptor activity in isolation-reared mice. Furthermore, isolation rearing increased the expression of AMPA receptor subunits (GluR1, GluR2 and GluR3) and GluR1 Ser845 phosphorylation in the prefrontal cortex, but not in the hippocampus or nucleus accumbens. Taken together, these results suggest that an increase in AMPA receptor activity in the prefrontal cortex contributes to some, but not all, abnormal behaviours in isolation-reared mice.

  19. Effects of serotonin 2A/1A receptor stimulation on social exclusion processing

    PubMed Central

    Preller, Katrin H.; Pokorny, Thomas; Hock, Andreas; Kraehenmann, Rainer; Stämpfli, Philipp; Seifritz, Erich; Scheidegger, Milan; Vollenweider, Franz X.

    2016-01-01

    Social ties are crucial for physical and mental health. However, psychiatric patients frequently encounter social rejection. Moreover, an increased reactivity to social exclusion influences the development, progression, and treatment of various psychiatric disorders. Nevertheless, the neuromodulatory substrates of rejection experiences are largely unknown. The preferential serotonin (5-HT) 2A/1A receptor agonist, psilocybin (Psi), reduces the processing of negative stimuli, but whether 5-HT2A/1A receptor stimulation modulates the processing of negative social interactions remains unclear. Therefore, this double-blind, randomized, counterbalanced, cross-over study assessed the neural response to social exclusion after the acute administration of Psi (0.215 mg/kg) or placebo (Pla) in 21 healthy volunteers by using functional magnetic resonance imaging (fMRI) and resting-state magnetic resonance spectroscopy (MRS). Participants reported a reduced feeling of social exclusion after Psi vs. Pla administration, and the neural response to social exclusion was decreased in the dorsal anterior cingulate cortex (dACC) and the middle frontal gyrus, key regions for social pain processing. The reduced neural response in the dACC was significantly correlated with Psi-induced changes in self-processing and decreased aspartate (Asp) content. In conclusion, 5-HT2A/1A receptor stimulation with psilocybin seems to reduce social pain processing in association with changes in self-experience. These findings may be relevant to the normalization of negative social interaction processing in psychiatric disorders characterized by increased rejection sensitivity. The current results also emphasize the importance of 5-HT2A/1A receptor subtypes and the Asp system in the control of social functioning, and as prospective targets in the treatment of sociocognitive impairments in psychiatric illnesses. PMID:27091970

  20. Effects of serotonin 2A/1A receptor stimulation on social exclusion processing.

    PubMed

    Preller, Katrin H; Pokorny, Thomas; Hock, Andreas; Kraehenmann, Rainer; Stämpfli, Philipp; Seifritz, Erich; Scheidegger, Milan; Vollenweider, Franz X

    2016-05-03

    Social ties are crucial for physical and mental health. However, psychiatric patients frequently encounter social rejection. Moreover, an increased reactivity to social exclusion influences the development, progression, and treatment of various psychiatric disorders. Nevertheless, the neuromodulatory substrates of rejection experiences are largely unknown. The preferential serotonin (5-HT) 2A/1A receptor agonist, psilocybin (Psi), reduces the processing of negative stimuli, but whether 5-HT2A/1A receptor stimulation modulates the processing of negative social interactions remains unclear. Therefore, this double-blind, randomized, counterbalanced, cross-over study assessed the neural response to social exclusion after the acute administration of Psi (0.215 mg/kg) or placebo (Pla) in 21 healthy volunteers by using functional magnetic resonance imaging (fMRI) and resting-state magnetic resonance spectroscopy (MRS). Participants reported a reduced feeling of social exclusion after Psi vs. Pla administration, and the neural response to social exclusion was decreased in the dorsal anterior cingulate cortex (dACC) and the middle frontal gyrus, key regions for social pain processing. The reduced neural response in the dACC was significantly correlated with Psi-induced changes in self-processing and decreased aspartate (Asp) content. In conclusion, 5-HT2A/1A receptor stimulation with psilocybin seems to reduce social pain processing in association with changes in self-experience. These findings may be relevant to the normalization of negative social interaction processing in psychiatric disorders characterized by increased rejection sensitivity. The current results also emphasize the importance of 5-HT2A/1A receptor subtypes and the Asp system in the control of social functioning, and as prospective targets in the treatment of sociocognitive impairments in psychiatric illnesses.

  1. Acetylcholine stimulates cortical precursor cell proliferation in vitro via muscarinic receptor activation and MAP kinase phosphorylation.

    PubMed

    Ma, W; Maric, D; Li, B S; Hu, Q; Andreadis, J D; Grant, G M; Liu, Q Y; Shaffer, K M; Chang, Y H; Zhang, L; Pancrazio, J J; Pant, H C; Stenger, D A; Barker, J L

    2000-04-01

    Increasing evidence has shown that some neurotransmitters act as growth-regulatory signals during brain development. Here we report a role for the classical neurotransmitter acetylcholine (ACh) to stimulate proliferation of neural stem cells and stem cell-derived progenitor cells during neural cell lineage progression in vitro. Neuroepithelial cells in the ventricular zone of the embryonic rat cortex were found to express the m2 subtype of the muscarinic receptor. Neural precursor cells dissociated from the embryonic rat cortical neuroepithelium were expanded in culture with basic fibroblast growth factor (bFGF). reverse transcriptase-polymerase chain reaction (RT-PCR) revealed the presence of m2, m3 and m4 muscarinic receptor subtype transcripts, while immunocytochemistry demonstrated m2 protein. ACh and carbachol induced an increase in cytosolic Ca2+ and membrane currents in proliferating (BrdU+) cells, both of which were abolished by atropine. Exposure of bFGF-deprived precursor cells to muscarinic agonists not only increased both cell number and DNA synthesis, but also enhanced differentiation of neurons. These effects were blocked by atropine, indicating the involvement of muscarinic ACh receptors. The growth-stimulating effects were also antagonized by a panel of inhibitors of second messengers, including 1,2-bis-(O-aminophenoxy)-ethane-N,N,N', N'-tetraacetic acid (BAPTA-AM) to chelate cytosolic Ca2+, EGTA to complex extracellular Ca2+, pertussis toxin, which uncouples certain G-proteins, the protein kinase C inhibitor H7 and the mitogen-activated protein kinase (MAPK) inhibitor PD98059. Muscarinic agonists activated MAPK, which was significantly inhibited by atropine and the same panel of inhibitors. Thus, muscarinic receptors expressed by neural precursors transduce a growth-regulatory signal during neurogenesis via pathways involving pertussis toxin-sensitive G-proteins, Ca2+ signalling, protein kinase C activation, MAPK phosphorylation and DNA synthesis.

  2. Opioid peptide receptor stimulation reverses beta-adrenergic effects in rat heart cells.

    PubMed

    Xiao, R P; Pepe, S; Spurgeon, H A; Capogrossi, M C; Lakatta, E G

    1997-02-01

    Opioid peptide receptor (OPR) agonists are co-released with the beta-adrenergic receptor (beta-AR) agonist norepinephrine (NE) from nerve terminals in the heart during sympathetic stimulation. Whereas recent studies indicate that OPR and beta-AR coexist on the surface of cardiac myocytes, whether significant "cross talk" occurs between OPR and beta-AR signaling cascades within heart cells is unknown. In the present study we demonstrate a marked effect of delta-OPR stimulation to modulate beta-adrenergic responses in single isolated rat ventricular myocytes. Nanomolar concentrations (10(-8) M) of the OPR agonist leucine enkephalin (LE), a naturally occurring delta-opioid peptide, inhibited NE-induced increases in sarcolemmal L-type Ca2+ current, cytosolic Ca2+ transient, and contraction. The antiadrenergic effect of LE was pertussis toxin sensitive and abolished by naloxone, an opioid receptor antagonist. In contrast, LE was unable to inhibit the positive inotropic effects induced by equipotent concentrations of 8-(4 chlorophenylthio)-adenosine 3',5'-cyclic monophosphate, a cell-permeant adenosine 3',5'-cyclic monophosphate analog, or by the non-receptor-induced increase in contraction by elevated bathing Ca2+ concentration. These results indicate that an interaction of the OPR and beta-AR systems occurs proximal to activation of the adenosine 3',5'-cyclic monophosphate-dependent protein kinase of the beta-AR intracellular signaling pathway. This modulation of beta-adrenergic effects by OPR activation at the myocyte level may have important implications in the regulation of cardiac Ca2+ metabolism and contractility, particularly during the myocardial response to stress.

  3. CB1 receptor stimulation in specific brain areas differently modulate anxiety-related behaviour.

    PubMed

    Rubino, T; Guidali, C; Vigano, D; Realini, N; Valenti, M; Massi, P; Parolaro, D

    2008-01-01

    There is a general consensus that the effects of cannabinoid agonists on anxiety seem to be biphasic, with low doses being anxiolytic and high doses ineffective or possibly anxiogenic. Besides the behavioural effects of cannabinoids on anxiety, very few papers have dealt with the neuroanatomical sites of these effects. We investigated the effect on rat anxiety behavior of local administration of THC in the prefrontal cortex, basolateral amygdala and ventral hippocampus, brain regions belonging to the emotional circuit and containing high levels of CB1 receptors. THC microinjected at low doses in the prefrontal cortex (10 microg) and ventral hippocampus (5 microg) induced in rats an anxiolytic-like response tested in the elevated plus-maze, whilst higher doses lost the anxiolytic effect and even seemed to switch into an anxiogenic profile. Low THC doses (1 microg) in the basolateral amygdala produced an anxiogenic-like response whereas higher doses were ineffective. All these effects were CB1-dependent and closely linked to modulation of CREB activation. Specifically, THC anxiolytic activity in the prefrontal cortex and ventral hippocampus was paralleled by an increase in CREB activation, whilst THC anxiogenic response in the basolateral amygdala was paralleled by a decrease in CREB activation. Our results suggest that while a mild activation of CB1 receptors in the prefrontal cortex and ventral hippocampus attenuates anxiety, a slight CB1 receptor stimulation in the amygdala results in an anxiogenic-like response. The molecular underpinnings of these effects involve a direct stimulation of CB1 receptors ending in pCREB modulation and/or a possible alteration in the fine tuning of local neuromodulator release.

  4. Drug Hypersensitivity: How Drugs Stimulate T Cells via Pharmacological Interaction with Immune Receptors.

    PubMed

    Pichler, Werner J; Adam, Jacqueline; Watkins, Stephen; Wuillemin, Natascha; Yun, James; Yerly, Daniel

    2015-01-01

    Small chemicals like drugs tend to bind to proteins via noncovalent bonds, e.g. hydrogen bonds, salt bridges or electrostatic interactions. Some chemicals interact with other molecules than the actual target ligand, representing so-called 'off-target' activities of drugs. Such interactions are a main cause of adverse side effects to drugs and are normally classified as predictable type A reactions. Detailed analysis of drug-induced immune reactions revealed that off-target activities also affect immune receptors, such as highly polymorphic human leukocyte antigens (HLA) or T cell receptors (TCR). Such drug interactions with immune receptors may lead to T cell stimulation, resulting in clinical symptoms of delayed-type hypersensitivity. They are assigned the 'pharmacological interaction with immune receptors' (p-i) concept. Analysis of p-i has revealed that drugs bind preferentially or exclusively to distinct HLA molecules (p-i HLA) or to distinct TCR (p-i TCR). P-i reactions differ from 'conventional' off-target drug reactions as the outcome is not due to the effect on the drug-modified cells themselves, but is the consequence of reactive T cells. Hence, the complex and diverse clinical manifestations of delayed-type hypersensitivity are caused by the functional heterogeneity of T cells. In the abacavir model of p-i HLA, the drug binding to HLA may result in alteration of the presenting peptides. More importantly, the drug binding to HLA generates a drug-modified HLA, which stimulates T cells directly, like an allo-HLA. In the sulfamethoxazole model of p-i TCR, responsive T cells likely require costimulation for full T cell activation. These findings may explain the similarity of delayed-type hypersensitivity reactions to graft-versus-host disease, and how systemic viral infections increase the risk of delayed-type hypersensitivity reactions.

  5. PGE2 signaling through the EP4 receptor on fibroblasts upregulates RANKL and stimulates osteolysis.

    PubMed

    Tsutsumi, Ryosuke; Xie, Chao; Wei, Xiaochao; Zhang, Minjie; Zhang, Xinping; Flick, Lisa M; Schwarz, Edward M; O'Keefe, Regis J

    2009-10-01

    Periprosthetic osteolysis is the most common cause of aseptic loosening in total joint arthroplasty. The role of inflammatory mediators such as prostaglandin E2 (PGE2) and osteoclast promoting factors including RANKL in the pathogenesis of osteolysis has been well characterized. However, the PGE2 receptor (EP1, EP2, or EP4), and cell type in which it is expressed, which is responsible for PGE2 induction of RANKL during wear debris-induced osteolysis, has yet to be elucidated. To address this, we used mice genetically deficient in these EP receptors to assess PGE2 and wear debris responses in vitro and in vivo. Wear debris-induced osteolysis and RANKL expression were observed at similar levels in WT, EP1(-/-), and EP2(-/-) mice, indicating that these receptors do not mediate PGE2 signals in this process. A conditional knockout approach was used to eliminate EP4 expression in FSP1(+) fibroblasts that are the predominant source of RANKL. In the absence of EP4, fibroblasts do not express RANKL after stimulation with particles or PGE2, nor do they exhibit high levels of osteoclasts and osteolysis. These results show that periprosthetic fibroblasts are important mediators of osteolysis through the expression of RANKL, which is induced after PGE2 signaling through the EP4 receptor.

  6. Prostaglandin receptor EP2 mediates PGE2 stimulated hypercalcemia in mice in vivo.

    PubMed

    Li, Xiaodong; Tomita, Masato; Pilbeam, Carol C; Breyer, Richard M; Raisz, Lawrence G

    2002-04-01

    Prostaglandin E2 (PGE2) can stimulate bone resorption by a cyclic AMP-dependent pathway. Two PGE2 receptors, EP2 and EP4 have been shown to play a role in PGE2 stimulation of osteoclast formation. In primary osteoblastic cell cultures from EP2 wild type (EP2 +/+) mice, PGE2 (0.1 microM) increased cyclic AMP production 3.5-fold, but PGE2 had no effect on cells from mice in which the EP2 receptor had been deleted (EP2 -/-). To examine the role of the EP2 receptor in the resorption response in vivo we injected PGE2 in EP2 -/- mice, and compared them with EP2 +/+ mice. Injection of PGE2 (3 mg/kg, four times daily for three days) in 9- to 12-month-old male mice on a 129 SvEv background increased serum calcium from 9.8 +/- 0.5 to 10.7 +/- 0.3 mg/dl (P < 0.01) in EP2 +/+ mice but not in EP2 -/- mice (10.1 +/- 0.3 vs. 10.2 +/- 0.3 mg/dl). PGE2 injection (6 mg/kg twice a day for three days) in 3-4 month old male mice on a C57 BL/6 X 129 SvEv background increased calcium from 8.2 +/- 0.1 to 9.0 +/- 0.3 mg/dl (P < 0.05) in EP2 +/+ mice but had no effect in EP2-/- mice (8.4 +/- 0.1 vs. 8.3 +/- 0.2 mg/dl). Injection of PGE2 over the calvariae of EP2 +/+ and EP2-/- mice increased the expression of receptor activator of nuclear factor kappaB ligand (RANKL) both locally and in the tibia, but RANKL responses were lower in EP2 -/- mice. We conclude that EP2 receptor plays a role in the hypercalcemic response to PGE2. This impaired response in EP2 -/- mice may be due to decreased ability to stimulate cyclic AMP and in part, to a smaller increase in the expression of RANKL mRNA.

  7. Protection against ventricular fibrillation via cholinergic receptor stimulation and the generation of nitric oxide

    PubMed Central

    Kalla, Manish; Chotalia, Minesh; Coughlan, Charles; Hao, Guoliang; Crabtree, Mark J.; Tomek, Jakub; Bub, Gil; Paterson, David J.

    2016-01-01

    Key points Animal studies suggest an anti‐fibrillatory action of the vagus nerve on the ventricle, although the exact mechanism is controversial.Using a Langendorff perfused rat heart, we show that the acetylcholine analogue carbamylcholine raises ventricular fibrillation threshold (VFT) and flattens the electrical restitution curve.The anti‐fibrillatory action of carbamylcholine was prevented by the nicotinic receptor antagonist mecamylamine, inhibitors of neuronal nitric oxide synthase (nNOS) and soluble guanylyl cyclase (sGC), and can be mimicked by the nitric oxide (NO) donor sodium nitroprusside.Carbamylcholine increased NO metabolite content in the coronary effluent and this was prevented by mecamylamine.The anti‐fibrillatory action of both carbamylcholine and sodium nitroprusside was ultimately dependent on muscarinic receptor stimulation as all effects were blocked by atropine.These data demonstrate a protective effect of carbamylcholine on VFT that depends upon both muscarinic and nicotinic receptor stimulation, where the generation of NO is likely to be via a neuronal nNOS–sGC dependent pathway. Abstract Implantable cardiac vagal nerve stimulators are a promising treatment for ventricular arrhythmia in patients with heart failure. Animal studies suggest the anti‐fibrillatory effect may be nitric oxide (NO) dependent, although the exact site of action is controversial. We investigated whether a stable analogue of acetylcholine could raise ventricular fibrillation threshold (VFT), and whether this was dependent on NO generation and/or muscarinic/nicotinic receptor stimulation. VFT was determined in Langendorff perfused rat hearts by burst pacing until sustained VF was induced. Carbamylcholine (CCh, 200 nmol l–1, n = 9) significantly (P < 0.05) reduced heart rate from 292 ± 8 to 224 ± 6 b.p.m. Independent of this heart rate change, CCh caused a significant increase in VFT (control 1.5 ± 0.3 mA, CCh 2.4 ± 0.4 mA, wash 1.1

  8. Effect of Combination of Non-Invasive Spinal Cord Electrical Stimulation and Serotonin Receptor Activation in Patients with Chronic Spinal Cord Lesion.

    PubMed

    Moshonkina, T R; Shapkova, E Yu; Sukhotina, I A; Emeljannikov, D V; Gerasimenko, Yu P

    2016-10-01

    We analyzed the efficiency of percutaneous electrical stimulation of the spinal cord and serotonin receptor activation in rehabilitation of paralyzed patients. Four-week course of spinal cord electrical stimulation combined with mechanotherapy produced positive shifts in the status of chronically paralyzed patients. Serotonin receptor activation potentiated the effect of spinal cord stimulation and can be regarded as an additional neurorehabilitation option.

  9. Stimulation of inorganic-phosphate incorporation into phosphatidylinositol in rat thoracic aorta mediated through V1-vasopressin receptors.

    PubMed Central

    Takhar, A P; Kirk, C J

    1981-01-01

    Vasopressin stimulates the incorporation of [32P]Pi into phosphatidylinositol but not into other phospholipids in rat thoracic aorta strips. The relative abilities of three vasopressin analogues to stimulate phosphatidylinositol labelling in rat aorta are similar to their relative pressor potencies in vivo and to their relative potencies in stimulating the metabolism of rat hepatocytes, but very different from their relative antidiuretic potencies. The vasopressor antagonist [1-(beta-mercapto-beta, beta-cyclopentamethylenepropionic acid),8-arginine]vasopressin competitively inhibits [Arg8]vasopressin-stimulated phosphatidylinositol labelling in rat aorta with a pA2 of 8.1. It is concluded that the Ca2+-mobilizing vasopressin receptors (V1-receptors) of the rat aorta stimulate phosphatidylinositol metabolism, probably by enhancing phosphatidylinositol breakdown. PMID:6272723

  10. Stimulation of Sigma-1 Receptor Ameliorates Depressive-like Behaviors in CaMKIV Null Mice.

    PubMed

    Moriguchi, Shigeki; Sakagami, Hiroyuki; Yabuki, Yasushi; Sasaki, Yuzuru; Izumi, Hisanao; Zhang, Chen; Han, Feng; Fukunaga, Kohji

    2015-12-01

    Sigma-1 receptor (Sig-1R) is a molecular chaperone regulating calcium efflux from the neuronal endoplasmic reticulum to the mitochondria. Calcium/calmodulin-dependent protein kinase IV (CaMKIV) null mice exhibit depressive-like behaviors and impaired neurogenesis as assessed by bromodeoxyuridine (BrdU) incorporation into newborn cells of the hippocampal dentate gyrus (DG). Here, we demonstrate that chronic stimulation of Sig-1R by treatment with the agonist SA4503 or the SSRI fluvoxamine for 14 days improves depressive-like behaviors in CaMKIV null mice. By contrast, treatment with paroxetine, which lacks affinity for Sig-1R, did not alter these behaviors. Reduced numbers of BrdU-positive cells and decreased brain-derived neurotrophic factor (BDNF) mRNA expression and protein kinase B (Akt; Ser-473) phosphorylation seen in the DG of CaMKIV null mice were significantly rescued by chronic Sig-1R stimulation. Interestingly, reduced ATP production observed in the DG of CaMKIV null mice was improved by chronic Sig-1R stimulation. Such stimulation also improved hippocampal long-term potentiation (LTP) induction and maintenance, which are impaired in the DG of CaMKIV null mice. LTP rescue was closely associated with both increases in calcium/calmodulin-dependent protein kinase II (CaMKII) autophosphorylation and GluA1 (Ser-831) phosphorylation. Taken together, Sig-1R stimulation by SA4503 or fluvoxamine treatment increased hippocampal neurogenesis, which is closely associated with amelioration of depressive-like behaviors in CaMKIV null mice.

  11. Angiotensin type 2 receptor stimulation increases renal function in female, but not male, spontaneously hypertensive rats.

    PubMed

    Hilliard, Lucinda M; Chow, Charis L E; Mirabito, Katrina M; Steckelings, U Muscha; Unger, Thomas; Widdop, Robert E; Denton, Kate M

    2014-08-01

    Accumulating evidence suggests that the protective pathways of the renin-angiotensin system are enhanced in women, including the angiotensin type 2 receptor (AT2R), which mediates vasodilatory and natriuretic effects. To provide insight into the sex-specific ability of pharmacological AT2R stimulation to modulate renal function in hypertension, we examined the influence of the AT2R agonist, compound 21 (100-300 ng/kg per minute), on renal function in 18- to 19-week-old anesthetized male and female spontaneously hypertensive rats. AT2R stimulation significantly increased renal blood flow in female hypertensive rats (PTreatment<0.001), without influencing arterial pressure. For example, at 300 ng/kg per minute of compound 21, renal blood flow increased by 14.3±1.8% from baseline. Furthermore, at 300 ng/kg per minute of compound 21, a significant increase in urinary sodium excretion was observed in female hypertensive rats (+180±59% from baseline; P<0.05 versus vehicle-treated rats). This was seen in the absence of any major change in glomerular filtration rate, indicating that the natriuretic effects of AT2R stimulation were likely the result of altered renal tubular function. Conversely, we did not observe any significant effect of AT2R stimulation on renal hemodynamic or excretory function in male hypertensive rats. Finally, gene expression studies confirmed greater renal AT2R expression in female than in male hypertensive rats. Taken together, acute AT2R stimulation enhanced renal vasodilatation and sodium excretion without concomitant alterations in glomerular filtration rate in female hypertensive rats. Chronic studies of AT2R agonist therapy on renal function and arterial pressure in hypertensive states are now required to establish the suitability of AT2R as a therapeutic target for cardiovascular disease, particularly in women. © 2014 American Heart Association, Inc.

  12. Peroxisome proliferator-activated receptors as stimulants of angiogenesis in cardiovascular disease and diabetes.

    PubMed

    Desouza, Cyrus V; Rentschler, Lindsey; Fonseca, Vivian

    2009-09-25

    The incidence of diabetes is directly related to the incidence of obesity, which is at epidemic proportions in the US. Cardiovascular disease is a common complication of diabetes, which results in high morbidity and mortality. Peroxisome proliferator-activated receptors (PPARs) are a group of nuclear hormone receptors that regulate lipid and glucose metabolism. PPAR-α agonists such as fenofibrate and PPAR-γ agonists such as the thiozolidinediones have been used to treat dyslipidemia and insulin resistance in diabetes. Over the past few years research has discovered the role of PPARs in the regulation of inflammation, proliferation, and angiogenesis. Clinical trials looking at the effect of PPAR agonists on cardiovascular outcomes have produced controversial results. Studies looking at angiogenesis and proliferation in various animal models and cell lines have shown a wide variation in results. This may be due to the differential effects of PPARs on proliferation and angiogenesis in various tissues and pathologic states. This review discusses the role of PPARs in stimulating angiogenesis. It also reviews the settings in which stimulation of angiogenesis may be either beneficial or harmful.

  13. Hypertensive response following stimulation of opiate receptors in the caudal ventrolateral medulla.

    PubMed

    Willette, R N; Punnen, S; Krieger, A J; Sapru, H N

    1984-04-01

    In urethane-anesthetized rats, vasodepressor neuron pools were located bilaterally in and adjacent to the A1 area of the ventrolateral medulla by injecting the neuroexcitatory amino acid, L-glutamate. Ventrolateral vasodepressor areas included the caudalateral part of the nucleus reticularis gigantocellularis, the rostrolateral part of the nucleus reticularis ventralis, and the dorsal nucleus reticularis lateralis. In the ventrolateral vasodepressor areas L-glutamate elicited a transient fall in blood pressure (BP) and heart rate (HR). The opiate agonist (D-ala2-met5)-enkephalinamide (DAME) was used to stimulate opiate receptors in vasodepressor sites, identified with L-glutamate. In these sites, bilateral injections (0.1 microliter/site) of DAME caused a dose-related (2.5-500.0 ng) increase in blood pressure and heart rate, as well as exaggeration of the response to occlusion of the carotid. The effects of DAME on blood pressure were completely abolished by alpha-adrenergic blockade (phentolamine, 2 mg/kg, i.v.) and all effects of DAME were reversed by the administration of naloxone HCl (1 mg/kg, i.v.). Naloxone reversal was accompanied by an unexpected "rebound" hypertension. Saline had no significant effects when injected, or administered intravenously, in the absence or presence of DAME. It was concluded that stimulation of opiate receptors in the ventrolateral vasodepressor areas activated sympathetic outflow. An enkephalinergic system in this area of the brain stem may serve to modulate blood pressure, heart rate and cardiovascular reflexes.

  14. SYNAPTIC TRANSLATION OF STRIATAL-ENRICHED TYROSINE PHOSPHATASE (STEP) AFTER β1-ADRENERGIC RECEPTOR STIMULATION

    PubMed Central

    Hu, Yaer; Zhang, Yang; Venkitaramani, Deepa V.; Lombroso, Paul J.

    2009-01-01

    The β-adrenergic system is implicated in long-term synaptic plasticity in the central nervous system, a process that requires protein synthesis. To identify proteins that are translated in response to β-adrenergic receptor stimulation and the pathways that regulate this process, we investigated the effects of isoproterenol on the translation of striatal-enriched protein tyrosine phosphatase (STEP) in both cortico-striatal slices and primary neuronal cultures. Isoproterenol stimulation induced a rapid dose-dependent increase in STEP expression. Anisomycin blocked the increase in STEP expression while actinomycin D had no effect, suggesting a translation-dependent mechanism. Isoproterenol-induced STEP translation required activation of β1 receptors. Application of the MEK inhibitor SL327 blocked both isoproterenol-induced activation of pERK and subsequent STEP translation. Inhibitors of PI3K (LY294002) or mTOR (rapamycin) also completely blocked STEP translation. These results suggest that co-activation of both the ERK and PI3K-Akt-mTOR pathways are required for STEP translation. As the substrates of STEP include ERK itself, these results suggest that STEP is translated upon β-adrenergic activation as part of a negative feedback mechanism. PMID:17623046

  15. Association of the thyroid stimulating hormone receptor gene (TSHR) with Graves' disease.

    PubMed

    Brand, Oliver J; Barrett, Jeffrey C; Simmonds, Matthew J; Newby, Paul R; McCabe, Christopher J; Bruce, Christopher K; Kysela, Boris; Carr-Smith, Jackie D; Brix, Thomas; Hunt, Penny J; Wiersinga, Wilmar M; Hegedüs, Laszlo; Connell, John; Wass, John A H; Franklyn, Jayne A; Weetman, Anthony P; Heward, Joanne M; Gough, Stephen C L

    2009-05-01

    Graves' disease (GD) is a common autoimmune disease (AID) that shares many of its susceptibility loci with other AIDs. The thyroid stimulating hormone receptor (TSHR) represents the primary autoantigen in GD, in which autoantibodies bind to the receptor and mimic its ligand, thyroid stimulating hormone, causing the characteristic clinical phenotype. Although early studies investigating the TSHR and GD proved inconclusive, more recently we provided convincing evidence for association of the TSHR region with disease. In the current study, we investigated a combined panel of 98 SNPs, including 70 tag SNPs, across an extended 800 kb region of the TSHR to refine association in a cohort of 768 GD subjects and 768 matched controls. In total, 28 SNPs revealed association with GD (P < 0.05), with strongest SNP associations at rs179247 (chi(2) = 32.45, P = 8.90 x 10(-8), OR = 1.53, 95% CI = 1.32-1.78) and rs12101255 (chi(2) = 30.91, P = 1.95 x 10(-7), OR = 1.55, 95% CI = 1.33-1.81), both located in intron 1 of the TSHR. Association of the most associated SNP, rs179247, was replicated in 303 GD families (P = 7.8 x 10(-4)). In addition, we provide preliminary evidence that the disease-associated genotypes of rs179247 (AA) and rs12101255 (TT) show reduced mRNA expression ratios of flTSHR relative to two alternate TSHR mRNA splice variants.

  16. Flavaglines Stimulate Transient Receptor Potential Melastatin Type 6 (TRPM6) Channel Activity

    PubMed Central

    Verkaart, Sjoerd A. J.; Lameris, Anke L.; Basmadjian, Christine; Zhao, Qian; Désaubry, Laurent; Bindels, René J. M.; Hoenderop, Joost G. J.

    2015-01-01

    Magnesium (Mg2+) is essential for enzymatic activity, brain function and muscle contraction. Blood Mg2+ concentrations are tightly regulated between 0.7 and 1.1 mM by Mg2+ (re)absorption in kidney and intestine. The apical entry of Mg2+ in (re)absorbing epithelial cells is mediated by the transient receptor potential melastatin type 6 (TRPM6) ion channel. Here, flavaglines are described as a novel class of stimulatory compounds for TRPM6 activity. Flavaglines are a group of natural and synthetic compounds that target the ubiquitously expressed prohibitins and thereby affect cellular signaling. By whole-cell patch clamp analyses, it was demonstrated that nanomolar concentrations of flavaglines increases TRPM6 activity by ∼2 fold. The stimulatory effects were dependent on the presence of the alpha-kinase domain of TRPM6, but did not require its phosphotransferase activity. Interestingly, it was observed that two natural occurring TRPM6 mutants with impaired insulin-sensitivity, TRPM6-p.Val1393Ile and TRPM6-p.Lys1584Glu, are not sensitive to flavagline stimulation. In conclusion, we have identified flavaglines as potent activators of TRPM6 activity. Our results suggest that flavaglines stimulate TRPM6 via the insulin receptor signaling pathway. PMID:25774985

  17. Flavaglines Stimulate Transient Receptor Potential Melastatin Type 6 (TRPM6) Channel Activity.

    PubMed

    Blanchard, Maxime G; de Baaij, Jeroen H F; Verkaart, Sjoerd A J; Lameris, Anke L; Basmadjian, Christine; Zhao, Qian; Désaubry, Laurent; Bindels, René J M; Hoenderop, Joost G J

    2015-01-01

    Magnesium (Mg2+) is essential for enzymatic activity, brain function and muscle contraction. Blood Mg2+ concentrations are tightly regulated between 0.7 and 1.1 mM by Mg2+ (re)absorption in kidney and intestine. The apical entry of Mg2+ in (re)absorbing epithelial cells is mediated by the transient receptor potential melastatin type 6 (TRPM6) ion channel. Here, flavaglines are described as a novel class of stimulatory compounds for TRPM6 activity. Flavaglines are a group of natural and synthetic compounds that target the ubiquitously expressed prohibitins and thereby affect cellular signaling. By whole-cell patch clamp analyses, it was demonstrated that nanomolar concentrations of flavaglines increases TRPM6 activity by ∼2 fold. The stimulatory effects were dependent on the presence of the alpha-kinase domain of TRPM6, but did not require its phosphotransferase activity. Interestingly, it was observed that two natural occurring TRPM6 mutants with impaired insulin-sensitivity, TRPM6-p.Val1393Ile and TRPM6-p.Lys1584Glu, are not sensitive to flavagline stimulation. In conclusion, we have identified flavaglines as potent activators of TRPM6 activity. Our results suggest that flavaglines stimulate TRPM6 via the insulin receptor signaling pathway.

  18. Multiple sclerosis lymphocytes upregulate A2A adenosine receptors that are antiinflammatory when stimulated.

    PubMed

    Vincenzi, Fabrizio; Corciulo, Carmen; Targa, Martina; Merighi, Stefania; Gessi, Stefania; Casetta, Ilaria; Gentile, Mauro; Granieri, Enrico; Borea, Pier Andrea; Varani, Katia

    2013-08-01

    Multiple sclerosis (MS) is an autoimmune-mediated inflammatory disease characterized by multifocal areas of demyelination. Experimental evidence indicates that A2A adenosine receptors (ARs) play a pivotal role in the inhibition of inflammatory processes. The aim of this study was to investigate the contribution of A2A ARs in the inhibition of key pro-inflammatory mediators for the pathogenesis of MS. In lymphocytes from MS patients, A1, A2A, A2B, and A3 ARs were analyzed by using RT-PCR, Western blotting, immunofluorescence, and binding assays. Moreover the effect of A2A AR stimulation on proinflammatory cytokine release such as TNF-α, IFN-γ, IL-6, IL-1β, IL-17, and on lymphocyte proliferation was evaluated. The capability of an A2A AR agonist on the modulation of very late antigen (VLA)-4 expression and NF-κB was also explored. A2A AR upregulation was observed in lymphocytes from MS patients in comparison with healthy subjects. The stimulation of these receptors mediated a significant inhibition of TNF-α, IFN-γ, IL-6, IL-1β, IL-17, and cell proliferation as well as VLA-4 expression and NF-κB activation. This new evidence highlights that A2A AR agonists could represent a novel therapeutic tool for MS treatment as suggested by the antiinflammatory role of A2A ARs in lymphocytes from MS patients.

  19. Potentiation of Brain Stimulation Reward by Morphine: Effects of Neurokinin-1 Receptor Antagonism

    PubMed Central

    Robinson, J.E.; Fish, E.W.; Krouse, M.C.; Thorsell, A.; Heilig, M.; Malanga, C.J.

    2012-01-01

    Rationale The abuse potential of opioids may be due to their reinforcing and rewarding effects, which may be attenuated by neurokinin-1 receptor (NK1R) antagonists. Objective To measure the effects of opioid and neurokinin-1 (NK1R) receptor blockade on the potentiation of brain stimulation reward (BSR) by morphine using the intracranial self-stimulation (ICSS) method. Methods Adult male C57BL/6J mice (n = 15) were implanted with unipolar stimulating electrodes in the lateral hypothalamus and trained to respond for varying frequencies of rewarding electrical stimulation. The BSR threshold (θ0) and maximum response rate (MAX) were determined before and after intraperitoneal administration of saline, morphine (1.0 - 17.0 mg/kg), or the NK1R antagonists L-733,060 (1.0 - 17.0 mg/kg) and L-703,606 (1.0 - 17.0 mg/kg). In morphine antagonism experiments, naltrexone (0.1 – 1.0 mg/kg) or 10.0 mg/kg L-733,060 or L-703,606 was administered 15 minutes before morphine (1.0 - 10.0 mg/kg) or saline. Results Morphine dose-dependently decreased θ0 (maximum effect = 62% of baseline) and altered MAX when compared to saline. L-703,606 and L-733,060 altered θ0 without affecting MAX. 10.0 mg/kg L-733,060 and L-703,606, which did not affect θ0 or MAX, attenuated the effects of 3.0 and 10.0 mg/kg morphine. 1.0 and 0.3 mg/kg naltrexone blocked the effects of 10.0 mg/kg morphine. Naltrexone given before saline did not affect θ0 or MAX. Conclusions The decrease in θ0 by morphine reflects its rewarding effects, which were attenuated by NK1R and opioid receptor blockade. These results demonstrate the importance of substance P signaling during limbic reward system activation by opioids. PMID:21909635

  20. Phencyclidine-induced social withdrawal results from deficient stimulation of cannabinoid CB₁ receptors: implications for schizophrenia.

    PubMed

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

    2013-08-01

    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.

  1. Differential involvement of IFN-beta in Toll-like receptor-stimulated dendritic cell activation.

    PubMed

    Hoshino, Katsuaki; Kaisho, Tsuneyasu; Iwabe, Tomio; Takeuchi, Osamu; Akira, Shizuo

    2002-10-01

    Toll-like receptor (TLR) can activate dendritic cells (DC) through common signaling pathways requiring a cytoplasmic adapter, MyD88. However, the signaling is differentially regulated among TLR family members. TLR4 can activate MyD88-deficient bone marrow-derived DC (BMDC), and lead to induction of IFN-inducible genes and up-regulation of co-stimulatory molecules such as CD40, implying that the MyD88-independent signaling pathway functions downstream of TLR4. Because these effects can also be induced by type I IFN, we have analyzed whether type I IFN is involved in TLR4-induced responses. In response to lipopolysaccharide (LPS), IFN-beta gene expression was augmented in both wild-type and MyD88-deficient BMDC. Expression of all IFN-inducible genes except immune-responsive gene 1 (IRG1) was abolished and CD40 up-regulation was decreased in LPS-stimulated BMDC lacking either IFN-alpha/beta receptor (IFN-alpha/betaR) or signal transducer and activator of transcription 1 (STAT-1). Similar to the LPS response, TLR9 signaling can also induce expression of IFN-beta and IFN-inducible genes, and up-regulation of CD40. However, all these effects were MyD88 dependent. Thus, in TLR4 signaling, IFN-beta expression can be induced either by the MyD88-dependent or -independent pathway, whereas, in TLR9 signaling, it is dependent on MyD88. In CpG DNA-stimulated DC, expression of IFN-inducible genes except IRG1 was dependent on type I IFN signaling as in LPS-stimulated DC. However, in contrast to TLR4 signaling, TLR9 signaling requires type I IFN signaling for CD40 up-regulation. Taken together, this study demonstrates differential involvement of type I IFN in TLR4- and TLR9-induced effects on DC.

  2. A Role for Sigma Receptors in Stimulant Self Administration and Addiction.

    PubMed

    Katz, Jonathan L; Su, Tsung-Ping; Hiranita, Takato; Hayashi, Teruo; Tanda, Gianluigi; Kopajtic, Theresa; Tsai, Shang-Yi

    2011-01-01

    Sigma(1) receptors (σ(1)Rs) represent a structurally unique class of intracellular proteins that function as chaperones. σ(1)Rs translocate from the mitochondria-associated membrane to the cell nucleus or cell membrane, and through protein-protein interactions influence several targets, including ion channels, G-protein-coupled receptors, lipids, and other signaling proteins. Several studies have demonstrated that σR antagonists block stimulant-induced behavioral effects, including ambulatory activity, sensitization, and acute toxicities. Curiously, the effects of stimulants have been blocked by σR antagonists tested under place-conditioning but not self-administration procedures, indicating fundamental differences in the mechanisms underlying these two effects. The self administration of σR agonists has been found in subjects previously trained to self administer cocaine. The reinforcing effects of the σR agonists were blocked by σR antagonists. Additionally, σR agonists were found to increase dopamine concentrations in the nucleus accumbens shell, a brain region considered important for the reinforcing effects of abused drugs. Although the effects of the σR agonist, DTG, on dopamine were obtained at doses that approximated those that maintained self administration behavior those of another agonist, PRE-084 required higher doses. The effects of DTG were antagonized by non-selective or a preferential σ(2)R antagonist but not by a preferential σ(1)R antagonist. The effects of PRE-084 on dopamine were insensitive to σR antagonists. The data suggest that the self administration of σR agonists is independent of dopamine and the findings are discussed in light of a hypothesis that cocaine has both intracellular actions mediated by σRs, as well as extracellular actions mediated through conventionally studied mechanisms. The co-activation and potential interactions among these mechanisms, in particular those involving the intracellular chaperone σRs, may

  3. Hypotensive and sympathoinhibitory responses to selective central AT2 receptor stimulation in spontaneously hypertensive rats.

    PubMed

    Brouwers, Sofie; Smolders, Ilse; Wainford, Richard D; Dupont, Alain G

    2015-07-01

    The type 2 angiotensin receptor (AT2R) has been suggested to counterbalance the type 1 angiotensin receptor (AT1R) in the central regulation of blood pressure and sympathetic tone. In the present study we investigated the blood pressure responses to stimulation of central AT2Rs by the selective agonist Compound 21 in conscious spontaneously hypertensive rats (SHRs) and normotensive Wistar Kyoto rats (WKY rats). We also assessed the impact on noradrenaline [norepinephrine (NE)] plasma levels, autonomic function, spontaneous baroreflex sensitivity, and the possible involvement of the nitric oxide (NO) pathway and the AT1Rs. Chronic intracerebroventricular Compound 21 infusion lowered blood pressure and NE plasma levels in both rat strains. The night-time hypotensive effect was greater in SHRs compared with WKY rats. Compound 21 improved spontaneous baroreflex sensitivity more in SHRs than in WKY rats. These effects were abolished by co-administration of the AT2R antagonist PD123319 or the NO synthase inhibitor Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME). Central AT1R blockade did not enhance the hypotensive response to Compound 21. Chronic selective stimulation of central AT2Rs lowers blood pressure through sympathoinhibition, and improves spontaneous baroreflex sensitivity more in SHRs than in WKY rats. These responses appear to require a functioning central NO pathway, but are not modified by central AT1R blockade. Collectively, the data demonstrate specific beneficial effects of stimulation of central AT2Rs in hypertension associated with increased sympathetic tone, and suggest that central AT2Rs may represent a potential new therapeutic target for the treatment of neurogenic hypertension.

  4. cap alpha. /sub 2/-Adrenergic receptor-mediated sensitization of forskolin-stimulated cyclic AMP production

    SciTech Connect

    Jones, S.B.; Toews, M.L.; Turner, J.T.; Bylund, D.B.

    1987-03-01

    Preincubation of HT29 human colonic adenocarcinoma cells with ..cap alpha../sub 2/-adrenergic agonists resulted in a 10- to 20-fold increase in forskolin-stimulated cyclic AMP production as compared to cells preincubated without agonist. Similar results were obtained using either a (/sup 3/H)adenine prelabeling assay or a cyclic AMP radioimmunoassay to measure cyclic AMP levels. This phenomenon, which is termed sensitization, is ..cap alpha../sub 2/-adrenergic receptor-mediated and rapid in onset and reversal. Yohimbine, an ..cap alpha../sub 2/-adrenergic receptor-selective antagonist, blocked norepinephrine-induced sensitization, whereas prazosin (..cap alpha../sub 1/-adrenergic) and sotalol (..beta..-adrenergic) did not. The time for half-maximal sensitization was 5 min and the half-time for reversal was 10 min. Only a 2-fold sensitization of cyclic AMP production stimulated by vasoactive intestinal peptide was observed, indicating that sensitization is relatively selective for forskolin. Sensitization reflects an increased production of cyclic AMP and not a decreased degradation of cyclic AMP, since incubation with a phosphodiesterase inhibitor and forskolin did not mimic sensitization. Increasing the levels of cyclic AMP during the preincubation had no effect on sensitization, indicating that sensitization is not caused by decreased cyclic AMP levels during the preincubation. This rapid and dramatic sensitization of forskolin-stimulated cyclic AMP production is a previously unreported effect that can be added to the growing list of ..cap alpha../sub 2/-adrenergic responses that are not mediated by a decrease in cyclic AMP.

  5. A Role for Sigma Receptors in Stimulant Self Administration and Addiction

    PubMed Central

    Katz, Jonathan L.; Su, Tsung-Ping; Hiranita, Takato; Hayashi, Teruo; Tanda, Gianluigi; Kopajtic, Theresa; Tsai, Shang-Yi

    2011-01-01

    Sigma1 receptors (σ1Rs) represent a structurally unique class of intracellular proteins that function as chaperones. σ1Rs translocate from the mitochondria-associated membrane to the cell nucleus or cell membrane, and through protein-protein interactions influence several targets, including ion channels, G-protein-coupled receptors, lipids, and other signaling proteins. Several studies have demonstrated that σR antagonists block stimulant-induced behavioral effects, including ambulatory activity, sensitization, and acute toxicities. Curiously, the effects of stimulants have been blocked by σR antagonists tested under place-conditioning but not self-administration procedures, indicating fundamental differences in the mechanisms underlying these two effects. The self administration of σR agonists has been found in subjects previously trained to self administer cocaine. The reinforcing effects of the σR agonists were blocked by σR antagonists. Additionally, σR agonists were found to increase dopamine concentrations in the nucleus accumbens shell, a brain region considered important for the reinforcing effects of abused drugs. Although the effects of the σR agonist, DTG, on dopamine were obtained at doses that approximated those that maintained self administration behavior those of another agonist, PRE-084 required higher doses. The effects of DTG were antagonized by non-selective or a preferential σ2R antagonist but not by a preferential σ1R antagonist. The effects of PRE-084 on dopamine were insensitive to σR antagonists. The data suggest that the self administration of σR agonists is independent of dopamine and the findings are discussed in light of a hypothesis that cocaine has both intracellular actions mediated by σRs, as well as extracellular actions mediated through conventionally studied mechanisms. The co-activation and potential interactions among these mechanisms, in particular those involving the intracellular chaperone σRs, may lead to the

  6. Follicle stimulating hormone receptor in mesenchymal stem cells integrates effects of glycoprotein reproductive hormones

    PubMed Central

    Tourkova, Irina L.; Witt, Michelle R.; Li, La; Larrouture, Quitterie; Liu, Li; Luo, Jianhua; Robinson, Lisa J.; Blair, Harry C.

    2014-01-01

    Previously we reported that follicle stimulating hormone (FSH) affects bone degradation in human cells and in FSH-R null mice. Here we describe a FSH-R knockout bone formation phenotype. We used mesenchymal stem cells (MSCs), osteoblast precursors that express follicle stimulating hormone receptor (FSH-R), to determine whether FSH regulates bone formation. FSH stimulates MSC cell adhesion 1–3 h and proliferation at 24 h after addition. On the basis of phylogenetic and clinical precedents, we also examined effects of pregnant levels of human chorionic gonadotropin (hCG) on MSCs. We found effects similar to those of FSH, and RNAi knockdown of FSH-R abrogated both FSH and hCG effects on MSCs. In contrast to effects on MSCs, neither FSH nor hCG had significant effects on osteoblast maturation. Also in MSCs, short term treatment by FSH and hCG altered signaling pathways for proliferation, including Erk1/2 phosphorylation. Our results show augmentation of MSC proliferation by either FSH at menopausal levels or hCG at normal pregnant levels. We conclude that FSH-R participates in regulation of MSC precursor pools in response to either FSH or hCG, integrating the effects of these two glycoprotein hormones. PMID:25118101

  7. Follicle stimulating hormone receptor in mesenchymal stem cells integrates effects of glycoprotein reproductive hormones.

    PubMed

    Tourkova, Irina L; Witt, Michelle R; Li, La; Larrouture, Quitterie; Liu, Li; Luo, Jianhua; Robinson, Lisa J; Blair, Harry C

    2015-01-01

    Previously we reported that follicle stimulating hormone (FSH) affects bone degradation in human cells and in follicle stimulating hormone receptor (FSH-R) null mice. Here we describe a FSH-R knockout bone-formation phenotype. We used mesenchymal stem cells (MSCs), osteoblast precursors that express FSH-R, to determine whether FSH regulates bone formation. FSH stimulates MSC cell adhesion 1-3 h and proliferation at 24 h after addition. On the basis of phylogenetic and clinical precedents, we also examined effects of pregnant levels of human chorionic gonadotropin (hCG) on MSCs. We found effects similar to those of FSH, and RNAi knockdown of FSH-R abrogated both FSH and hCG effects on MSCs. In contrast to effects on MSCs, neither FSH nor hCG had significant effects on osteoblast maturation. Also in MSCs, short-term treatment by FSH and hCG altered signaling pathways for proliferation, including Erk1/2 phosphorylation. Our results show augmentation of MSC proliferation by either FSH at menopausal levels or hCG at normal pregnant levels. We conclude that FSH-R participates in regulation of MSC precursor pools in response to either FSH or hCG, integrating the effects of these two glycoprotein hormones.

  8. Pulmonary rapidly adapting receptor stimulation does not increase airway resistance in anesthetized rabbits.

    PubMed

    Yu, J; Zhang, J F; Roberts, A M; Collins, L C; Fletcher, E C

    1999-09-01

    In open-chest artificially ventilated rabbits, removal followed by replacement of positive end-expiratory pressure (PEEP maneuver) favors stimulation of airway rapidly adapting receptors (RARs). The purpose of the present study was to determine whether activation of RARs can cause bronchoconstriction. We measured airway pressure, airflow, and tidal volume, and calculated dynamic lung compliance and total lung resistance. PEEP maneuver increased airway pressure swings (16.4 +/- 4% above control; p = 0.0016) and decreased compliance (to 84.8 +/- 2.8% of control; p = 0.0002) without changing resistance (108.0 +/- 4.4% of control; p = 0.85). On the other hand, the resistance increased greatly (93 +/- 13%, p < 0.01) after intravenous injection of acetylcholine or electrical stimulation of vagal efferents, indicating that our system could detect increases in the resistance. In a separate group, we stimulated RARs by stroking the trachea with a cotton tip (tickling), tickling produced cough, manifested by increased pressure and flow without resistance changing. These changes were abolished after paralysis with succinylcholine. Because we did not detect an increase in airflow resistance during activation of RARs by the PEEP maneuver and tickling, we conclude that increase in resistance may not be an important reflex component of airway RARs.

  9. Cell surface adrenergic receptor stimulation modifies the endothelial response to SIRS. Systemic Inflammatory Response Syndrome.

    PubMed

    Tighe, D; Moss, R; Bennett, D

    1996-11-01

    The complex pathway seen in patients with the systemic inflammatory response syndrome (SIRS) does not readily respond to mediator blockade. All such trials conducted in SIRS patients have shown no benefit in reducing mortality. We have shown experimentally that in sepsis, the administration of beta 2-adrenoceptor agonists reduces hepatic cellular injury, whereas administration of an alpha 1-adrenoceptor agonist increases hepatic cellular injury. Inflammatory mediators can cause a dose-related reversible change in target endothelial cells (ECs). There is a substantial body of literature describing the anti-inflammatory effects of beta 2-adrenoceptor agonists. They reduce both the increased permeability and the production of inflammatory mediators from ECs. Cellular transduction processes are involved when adrenergic receptor agonists modify either the anti-inflammatory or proinflammatory response to sepsis in ECs. Inflammatory mediators and alpha 1-adrenoceptor agonists stimulate their trimeric G protein-linked receptors to produce diacylglycerol (DAG) and increase the intracellular concentration of calcium. DAG is involved in the production of both inflammatory proteins and lipids. In addition, mitogen-activated protein kinase (MAPK) is activated which is also involved in the production of inflammatory proteins and lipids. beta 2-adrenoceptor agonists activate their trimeric G protein-linked receptors to produce the stimulatory G protein (Gs). Gs stimulates adenyl cyclase to form cyclic adenosine monophosphate (cAMP) and activate protein kinase A (PKA). PKA is involved in activating gene transcription agents to produce anti-inflammatory proteins such as interleukin-10. PKA also inhibits phospholipase C and MAPK. Although promising, the use of beta-adrenoceptor agonists or agonists that increase cellular cAMP to activate the cells' endogenous anti-inflammatory pathway requires further study.

  10. Recombinant canine single chain insulin analogues: insulin receptor binding capacity and ability to stimulate glucose uptake.

    PubMed

    Adams, Jamie P; Holder, Angela L; Catchpole, Brian

    2014-12-01

    Virtually all diabetic dogs require exogenous insulin therapy to control their hyperglycaemia. In the UK, the only licensed insulin product currently available is a purified porcine insulin preparation. Recombinant insulin is somewhat problematic in terms of its manufacture, since the gene product (preproinsulin) undergoes substantial post-translational modification in pancreatic β cells before it becomes biologically active. The aim of the present study was to develop recombinant canine single chain insulin (SCI) analogues that could be produced in a prokaryotic expression system and which would require minimal processing. Three recombinant SCI constructs were developed in a prokaryotic expression vector, by replacing the insulin C-peptide sequence with one encoding a synthetic peptide (GGGPGKR), or with one of two insulin-like growth factor (IGF)-2 C-peptide coding sequences (human: SRVSRRSR; canine: SRVTRRSSR). Recombinant proteins were expressed in the periplasmic fraction of Escherichia coli and assessed for their ability to bind to the insulin and IGF-1 receptors, and to stimulate glucose uptake in 3T3-L1 adipocytes. All three recombinant SCI analogues demonstrated preferential binding to the insulin receptor compared to the IGF-1 receptor, with increased binding compared to recombinant canine proinsulin. The recombinant SCI analogues stimulated glucose uptake in 3T3-L1 adipocytes compared to negligible uptake using recombinant canine proinsulin, with the canine insulin/cIGF-2 chimaeric SCI analogue demonstrating the greatest effect. Thus, biologically-active recombinant canine SCI analogues can be produced relatively easily in bacteria, which could potentially be used for treatment of diabetic dogs.

  11. Activation of nicotinic receptors on GABAergic amacrine cells in the rabbit retina indirectly stimulates dopamine release.

    PubMed

    Neal, M J; Cunningham, J R; Matthews, K L

    2001-01-01

    The retina possesses subpopulations of amacrine cells, which utilize different transmitters, including acetylcholine (ACh), GABA, and dopamine. We have examined interactions between these neurones by studying the effects of nicotinic agonists on GABA and dopamine release. Isolated rabbit retinas were incubated with [3H]dopamine and then superfused. Fractions of the superfusate (2 min) were collected and the [3H]dopamine in each sample was measured. Endogenous GABA release was examined by incubating retinas in a small chamber. At 5-min intervals, the medium was changed and the GABA measured by high-pressure liquid chromatography (HPLC). Exposure of the retina to nicotine, epibatidine, and other nicotinic agonists increased the release of both GABA and dopamine. The effects of nicotine and epibatidine were blocked by mecamylamine, confirming an action on nicotinic receptors. The action of epibatidine on dopamine release was unaffected by glutamate antagonists but was blocked by picrotoxin and gabazine. These results suggested that nicotine might increase dopamine release indirectly by stimulating the release of GABA, which in turn inhibited the release of an inhibitory transmitter acting tonically on the dopaminergic amacrines. Exposure of the retina to GABA caused a small increase in dopamine release. This hypothetical inhibitory transmitter was not GABA, an opioid, adenosine, glycine, nociceptin, a cannabinoid, or nitric oxide because appropriate antagonists did not affect the resting release of dopamine. However, metergoline, a 5HT1/5HT2 receptor antagonist, and ketanserin, a 5HT2A receptor antagonist, but not the 5HT1A antagonist WAY100635, increased the resting release of dopamine and blocked the effects of nicotine. The 5HT1A/5HT7 agonist 8-hydroxy DPAT inhibited both the nicotine and GABA-evoked release of dopamine. We conclude that nicotinic agonists directly stimulate the release of GABA, but the evoked release of dopamine is indirect, and arises from GABA

  12. Methylphenidate and μ opioid receptor interactions: a pharmacological target for prevention of stimulant abuse.

    PubMed

    Zhu, Jinmin; Spencer, Thomas J; Liu-Chen, Lee-Yuan; Biederman, Joseph; Bhide, Pradeep G

    2011-01-01

    Methylphenidate (MPH) is one of the most commonly used and highly effective treatments for attention deficit hyperactivity disorder (ADHD) in children and adults. As the therapeutic use of MPH has increased, so has its abuse and illicit street-use. Yet, the mechanisms associated with development of MPH-associated abuse and dependence are not well understood making it difficult to develop methods to help its mitigation. As a result, many ADHD patients especially children and youth, that could benefit from MPH treatment do not receive it and risk lifelong disabilities associated with untreated ADHD. Therefore, understanding the mechanisms associated with development of MPH addiction and designing methods to prevent it assume high public health significance. Using a mouse model we show that supra-therapeutic doses of MPH produce rewarding effects (surrogate measure for addiction in humans) in a conditioned place preference paradigm and upregulate μ opioid receptor (MOPR) activity in the striatum and nucleus accumbens, brain regions associated with reward circuitry. Co-administration of naltrexone, a non-selective opioid receptor antagonist, prevents MPH-induced MOPR activation and the rewarding effects. The MPH-induced MOPR activation and rewarding effect require activation of the dopamine D1 but not the D2-receptor. These findings identify the MOPR as a potential target for attenuating rewarding effects of MPH and suggest that a formulation that combines naltrexone with MPH could be a useful pharmaceutical approach to alleviate abuse potential of MPH and other stimulants.

  13. Role of macrophage-stimulating protein and its receptor, RON tyrosine kinase, in ciliary motility.

    PubMed Central

    Sakamoto, O; Iwama, A; Amitani, R; Takehara, T; Yamaguchi, N; Yamamoto, T; Masuyama, K; Yamanaka, T; Ando, M; Suda, T

    1997-01-01

    Macrophage-stimulating protein (MSP) is an 80-kD serum protein with homology to hepatocyte growth factor (HGF). Its receptor, RON tyrosine kinase, is a new member of the HGF receptor family. The MSP-RON signaling pathway has been implicated in the functional regulation of mononuclear phagocytes. However, the function of this pathway in other types of cells has not been elucidated. Here we show that in contrast to the HGF receptor, which was expressed at the basolateral surface, RON was localized at the apical surface of ciliated epithelia in the airways and oviduct. In addition, MSP was found in the bronchoalveolar space at biologically significant concentrations. MSP bound to RON on normal human bronchial epithelial cells with a high affinity (Kd = 0.5 nM) and induced autophosphorylation of RON. Activation of RON by MSP led to a significant increase in ciliary beat frequency of human nasal cilia. These findings indicate that the ciliated epithelium of the mucociliary transport apparatus is a novel target of MSP. Ciliary motility is critical for mucociliary transport. Our findings suggest that the MSP-RON signaling pathway is a novel regulatory system of mucociliary function and might be involved in the host defense and fertilization. PMID:9045873

  14. Gangliosides stimulate bradykinin B2 receptors to promote calmodulin kinase II-mediated neuronal differentiation.

    PubMed

    Kanatsu, Yoshinori; Chen, Nai Hong; Mitoma, Junya; Nakagawa, Tetsuto; Hirabayashi, Yoshio; Higashi, Hideyoshi

    2012-07-01

    Gangliosides mediate neuronal differentiation and maturation and are indispensable for the maintenance of brain function and survival. As part of our ongoing efforts to understand signaling pathways related to ganglioside function, we recently demonstrated that neuronal cells react to exogenous gangliosides GT1b and GD1b. Both of these gangliosides are enriched in the synapse-forming area of the brain and induce Ca(2+) release from intracellular stores, activation of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and activation of cdc42 to promote reorganization of cytoskeletal actin and dendritic differentiation. Here, we show that bradykinin B2 receptors transduce these reactions as a mediator for ganglioside glycan signals. The B2 antagonist Hoe140 inhibited ganglioside-induced CaMKII activation, actin reorganization and early development of axon- and dendrite-like processes of primary cultured hippocampal neurons. Furthermore, we confirmed by yeast reporter assay that major b-series gangliosides, GT1b, GD1b and GD3, stimulated B2 bradykinin receptors. We hypothesize that this B2 receptor-mediated ganglioside signal transduction pathway is one mechanism that modulates neuronal differentiation and maturation.

  15. The Genetics of the Thyroid Stimulating Hormone Receptor: History and Relevance

    PubMed Central

    Yin, Xiaoming; Latif, Rauf

    2010-01-01

    Background The thyroid stimulating hormone receptor (TSHR) is the key regulator of thyrocyte function. The gene for the TSHR on chromosome 14q31 has been implicated as coding for the major autoantigen in the autoimmune hyperthyroidism of Graves' disease (GD) to which T cells and autoantibodies are directed. Summary The TSHR is a seven-transmembrane domain receptor that undergoes complex posttranslational processing. In this brief review, we look at the genetics of this important autoantigen and its influence on a variety of tissue functions in addition to its role in the induction of GD. Conclusions There is convincing evidence that the TSH receptor gene confers increased susceptibility for GD, but not Hashimoto's thyroiditis. GD is associated with polymorphisms in the intron 1 gene region. How such noncoding nucleotide changes influence disease susceptibility remains uncertain, but is likely to involve TSHR splicing variants and/or microRNAs arising from this gene region. Whether such influences are confined to the thyroid gland or whether they influence cell function in the many extrathyroidal sites of TSHR expression remains unknown. PMID:20578897

  16. An mRNA expression analysis of stimulation and blockade of 5-HT7 receptors during memory consolidation.

    PubMed

    Pérez-García, Georgina; Gonzalez-Espinosa, Claudia; Meneses, Alfredo

    2006-04-25

    Despite the compelling support for 5-hydroxytryptamine (5-HT) receptors participation in learning and memory in mammal species, the molecular basis had been largely absent from any discussion of its mechanistic underpinnings. Here, we report that reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed that there was a higher level of expression of the investigated 5-HT receptor mRNAs in autoshaping-trained relative to untrained groups. Actually, pharmacological naïve untrained and autoshaping-trained rats showed significant differences, the latter groups expressing, in decreasing order, 5-HT1A < 5-HT6 < 5-HT4 < or = 5-HT7 receptors mRNA in prefrontal cortex and hippocampus. In order to determine more precisely mRNA expression and memory consolidation, we combined selective 5-HT7 receptors stimulation or blockade in the same animals, and brain areas individually analyzed. 5-HT7 receptors were strongly expressed in all the three brain areas of vehicle-trained rats relative to untrained group. The potential selective 5-HT7 receptor agonist AS 19 enhanced memory consolidation, attenuated mRNA receptors expression, and the facilitatory memory effect was reversed by SB-269970. Finally, pharmacological stimulation of 5-HT7 receptors reversed scopolamine- or dizocilpine-induced amnesia and receptor down-regulation.

  17. Regulation of Phospholipase D Activity and Phosphatidic Acid Production after Purinergic (P2Y6) Receptor Stimulation*

    PubMed Central

    Scott, Sarah A.; Xiang, Yun; Mathews, Thomas P.; Cho, Hyekyung P.; Myers, David S.; Armstrong, Michelle D.; Tallman, Keri A.; O'Reilly, Matthew C.; Lindsley, Craig W.; Brown, H. Alex

    2013-01-01

    Phosphatidic acid (PA) is a lipid second messenger located at the intersection of several lipid metabolism and cell signaling events including membrane trafficking, survival, and proliferation. Generation of signaling PA has long been primarily attributed to the activation of phospholipase D (PLD). PLD catalyzes the hydrolysis of phosphatidylcholine into PA. A variety of both receptor-tyrosine kinase and G-protein-coupled receptor stimulations have been shown to lead to PLD activation and PA generation. This study focuses on profiling the PA pool upon P2Y6 receptor signaling manipulation to determine the major PA producing enzymes. Here we show that PLD, although highly active, is not responsible for the majority of stable PA being produced upon UDP stimulation of the P2Y6 receptor and that PA levels are tightly regulated. By following PA flux in the cell we show that PLD is involved in an initial increase in PA upon receptor stimulation; however, when PLD is blocked, the cell compensates by increasing PA production from other sources. We further delineate the P2Y6 signaling pathway showing that phospholipase Cβ3 (PLCβ3), PLCδ1, DGKζ and PLD are all downstream of receptor activation. We also show that DGKζ is a novel negative regulator of PLD activity in this system that occurs through an inhibitory mechanism with PKCα. These results further define the downstream events resulting in PA production in the P2Y6 receptor signaling pathway. PMID:23723068

  18. New melanocortin 1 receptor binding motif based on the C-terminal sequence of alpha-melanocyte-stimulating hormone.

    PubMed

    Schiöth, Helgi B; Muceniece, Ruta; Mutule, Ilga; Wikberg, Jarl E S

    2006-10-01

    The C-terminal tripeptide of the alpha-melanocyte stimulating hormone (alpha-MSH11-13) possesses strong antiinflammatory activity without known cellular target. In order to better understand the structural requirements for function of such motif, we designed, synthesized and tested out Trp- and Tyr-containing analogues of the alpha-MSH11-13. Seven alpha-MSH11-13 analogues were synthesized and characterized for their binding to the melanocortin receptors recombinantly expressed in insect (Sf9) cells, infected with baculovirus carrying corresponding MC receptor DNA. We also tested these analogues on B16-F1 mouse melanoma cells endogenously expressing the MC1 receptor for binding and for ability to increase cAMP levels as well as on COS-7 cells transfected with the human MC receptors. The data indicate that HS401 (Ac-Tyr-Lys-Pro-Val-NH2) and HS402 (Ac-Lys-Pro-Val-Tyr-NH2) selectively bound to the MC1 receptor and stimulated cAMP generation in a concentration dependent way while the other Tyr- and Trp-containing alpha-MSH11-13 analogues neither bound to MC receptors nor stimulated cAMP. We have thus identified new MC receptor binding motif derived from the C-terminal sequence of alpha-MSH. The tetrapeptides have novel properties as the both act via MC-ergic pathways and also carry the anti-inflammatory alpha-MSH11-13 message sequence.

  19. Arginine vasopressin stimulates mesangial cell proliferation by activating the epidermal growth factor receptor.

    PubMed

    Ghosh, P M; Mikhailova, M; Bedolla, R; Kreisberg, J I

    2001-06-01

    The potent vasoconstrictor arginine vasopressin (AVP) is also a mitogen for mesangial cells. Treatment with AVP decreased transit time through the cell cycle. AVP-stimulated mesangial cell growth by activating both the Ras mitogen-activated protein kinase (MAPK) and the phosphatidylinositol 3-kinase (PI3K) cell signaling pathways. Both the selective PI3K inhibitor LY-294002 and the MAPK kinase (MEK) inhibitor PD-98059 inhibited AVP-stimulated mesangial cell proliferation. However, LY-294002 was more potent, indicating an important role for PI3K activation in AVP-stimulated mesangial cell proliferation. AVP appeared to exert its effect on MAPK and PI3K activation, as well as on cell proliferation, by activating the epidermal growth factor receptor (EGF-R). Pretreatment with the tyrphostin-derived EGF-R antagonist AG-1478 inhibited mesangial cell proliferation as well as the activation of extracellular signal-regulated kinase 1/2 (ERK1/2 or p42/p44(MAPK)), and p70S6 kinase, a downstream effector of PI3K, providing evidence that MAPK and PI3K activation, respectively, occurred downstream of EGF-R activation. Treatment with rapamycin, an inhibitor of the p70S6 kinase activator mTOR, also resulted in growth inhibition, further suggesting the importance of the PI3K signaling pathway in AVP-induced proliferation. AVP treatment appeared to transactivate EGF-R by inducing tyrosine phosphorylation of the Ca(2+)/protein kinase C (PKC)-dependent nonreceptor tyrosine kinase, Pyk2, leading to Pyk2/c-Src association and c-Src activation. This was followed by association of c-Src with EGF-R and EGF-R activation. These data suggested that AVP-stimulated Pyk2 tyrosine phosphorylation to activate c-Src, thereby leading to EGF-R transactivation.

  20. Benzylpiperizine-based party pills' impact on the Auckland City Hospital Emergency Department Overdose Database (2002-2004) compared with ecstasy (MDMA or methylene dioxymethamphetamine), gamma hydroxybutyrate (GHB), amphetamines, cocaine, and alcohol.

    PubMed

    Theron, Lynn; Jansen, Karl; Miles, Jennifer

    2007-02-16

    To examine the impact of 'party pills' (PP; herbal highs) on the Auckland City Hospital Emergency Department Overdose Database 2002-2004, and to present figures for five other substances in that database. Auckland City Hospital's Emergency Department's overdose database was reviewed for 2002, 2003, and 2004 for 'herbal ingestions' and 'party pills' (PP), ecstasy, methamphetamine, GHB, cocaine, and alcohol. Adverse effects attributed to PP were examined. In 2002, 1 patient presented with PP ingestion; 4 presented in 2003 and 21 in 2004 respectively (p<0.001). Of these 21 patients in 2004, 5 had allegedly ingested PP only and none required medical admission. PP only contributed to 1.58% of the overdose database for 2004. 'Party pills' appeared to have a minor impact on the overdose database at Auckland City Hospital between 2002 and 2004. There was a significant decrease in GHB presentations from 2003 to 2004 (p<0.001), but no significant fall in stimulant overdose presentations.

  1. Stimulation of 5-HT1B receptors decreases cocaine- and sucrose-seeking behavior.

    PubMed

    Acosta, Jazmin I; Boynton, Floren A; Kirschner, Kenneth F; Neisewander, Janet L

    2005-02-01

    Serotonin systems have been implicated in incentive motivation for cocaine, yet little is known about the role of 5-HT(1B) receptors in these processes. We used the extinction/reinstatement model to examine the effects of the 5-HT(1B/1A) receptor agonist, RU24969, on reinstatement of extinguished cocaine-seeking behavior. Rats trained to self-administer cocaine subsequently underwent extinction. They were then tested twice for cue and cocaine-primed reinstatement of extinguished cocaine-seeking behavior, receiving saline pretreatment 1 day and their assigned dose of RU24969 (0.3, 1.0, 3.0 mg/kg) the other day. Rats were later trained on a schedule of sucrose reinforcement in novel chambers and then tested for effects of RU24969 on cue reinstatement of sucrose-seeking behavior and locomotion. RU24969 decreased cue and cocaine reinstatement of cocaine-seeking behavior and cue reinstatement of sucrose-seeking behavior. Locomotion was increased only at the highest RU24969 dose (3 mg/kg). A subsequent experiment demonstrated that the effects of RU24969 (1 mg/kg) on extinguished cocaine-seeking behavior were reversed by the 5-HT(1B) antagonist GR127935 (3 mg/kg). These findings suggest that the effects of RU24969 on cue and cocaine reinstatement of cocaine-seeking behavior are 5-HT(1B) receptor-mediated. Overall, the results suggest that stimulation of 5-HT(1B) receptors may produce a general decrease in motivation.

  2. Positive AMPA receptor modulation rapidly stimulates BDNF release and increases dendritic mRNA translation.

    PubMed

    Jourdi, Hussam; Hsu, Yu-Tien; Zhou, Miou; Qin, Qingyu; Bi, Xiaoning; Baudry, Michel

    2009-07-08

    Brain-derived neurotrophic factor (BDNF) stimulates local dendritic mRNA translation and is involved in formation and consolidation of memory. 2H,3H,6aH-pyrrolidino[2'',1''-3',2']1,3-oxazino[6',5'-5,4]-benzo[e]1,4-dioxan-10-one (CX614), one of the best-studied positive AMPA receptor modulators (also known as ampakines), increases BDNF mRNA and protein and facilitates long-term potentiation (LTP) induction. Several other ampakines also improve performance in various behavioral and learning tasks. Since local dendritic protein synthesis has been implicated in LTP stabilization and in memory consolidation, this study investigated whether CX614 could influence synaptic plasticity by upregulating dendritic protein translation. CX614 treatment of primary neuronal cultures and acute hippocampal slices rapidly activated the translation machinery and increased local dendritic protein synthesis. CX614-induced activation of translation was blocked by K252a [(9S,10R,12R)-2,3,9,10,11,12-hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylic acid methyl ester], CNQX, APV, and TTX, and was inhibited in the presence of an extracellular BDNF scavenger, TrkB-Fc. The acute effect of CX614 on translation was mediated by increased BDNF release as demonstrated with a BDNF scavenging assay using TrkB-Fc during CX614 treatment of cultured primary neurons and was blocked by nifedipine, ryanodine, and lack of extracellular Ca(2+) in acute hippocampal slices. Finally, CX614, like BDNF, rapidly increased dendritic translation of an exogenous translation reporter. Together, our results demonstrate that positive modulation of AMPA receptors rapidly stimulates dendritic translation, an effect mediated by BDNF secretion and TrkB receptor activation. They also suggest that increased BDNF secretion and stimulation of local protein synthesis contribute to the effects of ampakines on synaptic plasticity.

  3. What is the effect of nicotinic acetylcholine receptor stimulation on osteoarthritis in a rodent animal model?

    PubMed Central

    Bock, Kilian; Plaass, Christian; Coger, Vincent; Peck, Claas-Tido; Reimers, Kerstin; Stukenborg-Colsman, Christina; Claassen, Leif

    2016-01-01

    Objectives: Despite the rising number of patients with osteoarthritis, no sufficient chondroprotective and prophylactic therapy for osteoarthritis has been established yet. The purpose of this study was to verify whether stimulation of the nicotinic acetylcholine receptor via nicotine has a beneficial effect on cartilage degeneration in the development of osteoarthritis and is capable of reducing the expression of proinflammatory cytokines and cartilage degrading enzymes in synovial membranes after osteoarthritis induction. Methods: Experimental osteoarthritis was induced in Lewis rats using a standardized osteoarthritis model with monoiodoacetate. A total of 16 Lewis rats were randomized into four groups: control, sham + nicotine application, osteoarthritis, and osteoarthritis + nicotine application. Nicotine (0.625 mg/kg twice daily) was administered intraperitoneally for 42 days. We analyzed histological sections, radiological images and the expression of the proinflammatory cytokines, such as interleukin-1β, tumor necrosis factor-α and interleukin-6, and of matrix metalloproteases 3, 9 and 13 and tissue inhibitors of metalloprotease-1 in synovial membranes via quantitative polymerase chain reaction. Results: Histological and x-ray examination revealed cartilage degeneration in the osteoarthritis group compared to control or sham + nicotine groups (histological control vs osteoarthritis: p = 0.002 and x-ray control vs osteoarthritis: p = 0.004). Nicotine treatment reduced the cartilage degeneration without significant differences. Osteoarthritis induction led to a higher expression of proinflammatory cytokines and matrix metalloproteases as compared to control groups. This effect was attenuated after nicotine administration. The differences of proinflammatory cytokines and matrix metalloproteases did not reach statistical significance. Conclusion: With the present small-scale study, we could not prove a positive effect of nicotinic

  4. GABAB receptor stimulation by baclofen and taurine enhances excitatory amino acid induced phosphatidylinositol turnover in neonatal rat cerebellum.

    PubMed

    Smith, S S; Li, J

    1991-10-28

    Excitatory amino acid stimulation of phosphatidylinositol (PI) hydrolysis has been associated with development of the CNS. Normally minimally ineffective in stimulating PI hydrolysis in the neonatal rat cerebellum, N-methyl-D-aspartate (NMDA) increased levels of PI hydrolysis 82.3 +/- 5.5% above basal values in the presence of 1 microM baclofen, a gamma-aminobutyric acidB (GABAB) receptor agonist. This effect was observed at day 7 but not in adult cerebellum. The effect of baclofen could be mimicked by low dose GABA and taurine, actions which were blocked by prior application of a specific GABAB antagonist. Therefore, the ability of NMDA to stimulate PI hydrolysis in neonatal cerebellar tissue may be regulated by the degree of GABAB receptor stimulation.

  5. Stimulating and blocking thyroid-stimulating hormone (TSH) receptor autoantibodies from patients with Graves' disease and autoimmune hypothyroidism have very similar concentration, TSH receptor affinity, and binding sites.

    PubMed

    Morgenthaler, Nils G; Ho, Su Chin; Minich, Waldemar B

    2007-03-01

    The distinct biological properties of TSH receptor (TSH-R) autoantibodies (TRAbs) from patients with Graves' disease (GD) are yet unexplained on the molecular level. Here we compare serum concentration, affinity to the TSH-R, and binding sites on the TSH-R of stimulating (TSAb) and blocking (TBAb) TRAbs. Four-step affinity purification using human recombinant TSH-R was performed with 22 TRAb-positive sera from GD patients (11 with only TSAb and 11 with only TBAb) and five control sera. Antibody concentration, TSH binding inhibition (TBII), and TSAb/TBAb activity of the purified TRAb were assessed. Labeled purified TRAbs were used for displacement studies with TRAb and an additional 30 patients and 10 control sera. TRAbs could be purified to 80-93% purity with recovery of the TBII and TSAb and TBAb activity. No TRAbs could be purified from healthy individuals. The mean +/- SD concentration of TRAb was 17.3 +/- 5.4 microg/IU for the TSAb sera (range, 9.6-25.9) and 18.2 +/- 8.5 microg/IU for the TBAb sera (range, 4.6-29.2), respectively (P = 0.79). Affinity was in the picomolar range for both TRAb subtypes with mean +/- sd dissociation constant of 167 +/- 109 pM (60-410 pM) for TSAb and 253 +/- 132 pM (80-410 pM) for TBAb (P = 0.12). Purified and labeled TSAb and TBAb showed a very similar binding pattern to the TSH-R in displacement studies with unlabeled TSAb/TBAb or unpurified patients sera, indicating binding sites on the TSH-R in close proximity to each other. TSAbs and TBAbs in the serum of patients with GD have similar characteristics. They are of low concentration with high affinity and have also similar binding epitopes on the TSH-R.

  6. Caffeine stimulates locomotor activity in the mammalian spinal cord via adenosine A1 receptor-dopamine D1 receptor interaction and PKA-dependent mechanisms.

    PubMed

    Acevedo, JeanMarie; Santana-Almansa, Alexandra; Matos-Vergara, Nikol; Marrero-Cordero, Luis René; Cabezas-Bou, Ernesto; Díaz-Ríos, Manuel

    2016-02-01

    Caffeine is a potent psychostimulant that can have significant and widely variable effects on the activity of multiple neuronal pathways. The most pronounced caffeine-induced behavioral effect seen in rodents is to increase locomotor activity which has been linked to a dose-dependent inhibition of A1 and A(2A) receptors. The effects of caffeine at the level of the lumbar spinal central pattern generator (CPG) network for hindlimb locomotion are lacking. We assessed the effects of caffeine to the locomotor function of the spinal CPG network via extracellular ventral root recordings using the isolated neonatal mouse spinal cord preparation. Addition of caffeine and of an A1 receptor antagonist significantly decreased the cycle period accelerating the ongoing locomotor rhythm, while decreasing burst duration reversibly in most preparations suggesting the role of A1 receptors as the primary target of caffeine. Caffeine and an A1 receptor antagonist failed to stimulate ongoing locomotor activity in the absence of dopamine or in the presence of a D1 receptor antagonist supporting A1/D1 receptor-dependent mechanism of action. The use of caffeine or an A1 receptor blocker failed to stimulate an ongoing locomotor rhythm in the presence of a blocker of the cAMP-dependent protein kinase (PKA) supporting the need of this intracellular pathway for the modulatory effects of caffeine to occur. These results support a stimulant effect of caffeine on the lumbar spinal network controlling hindlimb locomotion through the inhibition of A1 receptors and subsequent activation of D1 receptors via a PKA-dependent intracellular mechanism.

  7. Stimulation of human formyl peptide receptors by calpain inhibitors: homology modeling of receptors and ligand docking simulation.

    PubMed

    Fujita, Hisakazu; Kato, Takayuki; Watanabe, Norifumi; Takahashi, Tatsuji; Kitagawa, Seiichi

    2011-12-15

    Calpain inhibitors, including peptide aldehydes (N-acetyl-Leu-Leu-Nle-CHO and N-acetyl-Leu-Leu-Met-CHO) and α-mercapto-acrylic acid derivatives (PD150606 and PD151746), have been shown to stimulate phagocyte functions via activation of human formyl peptide receptor (hFPR) and/or hFPR-like 1 (hFPRL1). Using the homology modeling of the receptors and the ligand docking simulation, here we show that these calpain inhibitors could bind to the putative N-formyl-Met-Leu-Phe (fMLF) binding site on hFPR and/or hFPRL1. The studies with HEK-293 cells stably expressing hFPR or hFPRL1 showed that the concentrations of calpain inhibitors required to induce an increase in cytoplasmic free Ca(2+) ([Ca(2+)](i)) was much higher (>100 folds) than those of fMLF and Trp-Lys-Tyr-Met-Val-D-Met (WKYMVm). HEK-293 cells expressing hFPR or hFPRL1 with the mutated fMLF binding site never exhibited the [Ca(2+)](i) response to calpain inhibitors. When the optimal concentrations of each stimulus were used, pretreatment of cells with fMLF or WKYMVm abolished an increase in [Ca(2+)](i) induced by calpain inhibitors as well as the same stimulus, whereas pretreatment of cells with calpain inhibitors significantly suppressed, but never abolished, the [Ca(2+)](i) response induced by fMLF or WKYMVm, suggesting that the binding affinity of the inhibitors to the putative fMLF binding site may be lower than that of fMLF or WKYMVm. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Electrical Stimulation Decreases Coupling Efficiency Between Beta-Adrenergic Receptors and Cyclic AMP Production in Cultured Muscle Cells

    NASA Technical Reports Server (NTRS)

    Young, R. B.; Bridge, K. Y.

    1999-01-01

    Electrical stimulation of skeletal muscle cells in culture is an effective way to simulate the effects of muscle contraction and its effects on gene expression in muscle cells. Expression of the beta-adrenergic receptor and its coupling to cyclic AMP synthesis are important components of the signaling system that controls muscle atrophy and hypertrophy, and the goal of this project was to determine if electrical stimulation altered the beta-adrenergic response in muscle cells. Chicken skeletal muscle cells that had been grown for seven days in culture were subjected to electrical stimulation for an additional two days at a pulse frequency of 0.5 pulses/sec and a pulse duration of 200 msec. At the end of this two-day stimulation period, beta-adrenergic receptor population was measured by the binding of tritium-labeled CGP-12177 to muscle cells, and coupling to cAMP synthesis was measured by Radioimmunoassay (RIA) after treating the cells for 10 min with the potent (beta)AR agonist, isoproterenol. The number of beta adrenergic receptors and the basal levels of intracellular cyclic AMP were not affected by electrical stimulation. However, the ability of these cells to synthesize cyclic AMP was reduced by approximately 50%. Thus, an enhanced level of contraction reduces the coupling efficiency of beta-adrenergic receptors for cyclic AMP production.

  9. Electrical Stimulation Decreases Coupling Efficiency Between Beta-Adrenergic Receptors and Cyclic AMP Production in Cultured Muscle Cells

    NASA Technical Reports Server (NTRS)

    Young, R. B.; Bridge, K. Y.

    1999-01-01

    Electrical stimulation of skeletal muscle cells in culture is an effective way to simulate the effects of muscle contraction and its effects on gene expression in muscle cells. Expression of the beta-adrenergic receptor and its coupling to cyclic AMP synthesis are important components of the signaling system that controls muscle atrophy and hypertrophy, and the goal of this project was to determine if electrical stimulation altered the beta-adrenergic response in muscle cells. Chicken skeletal muscle cells that had been grown for seven days in culture were subjected to electrical stimulation for an additional two days at a pulse frequency of 0.5 pulses/sec and a pulse duration of 200 msec. At the end of this two-day stimulation period, beta-adrenergic receptor population was measured by the binding of tritium-labeled CGP-12177 to muscle cells, and coupling to cAMP synthesis was measured by Radioimmunoassay (RIA) after treating the cells for 10 min with the potent (beta)AR agonist, isoproterenol. The number of beta adrenergic receptors and the basal levels of intracellular cyclic AMP were not affected by electrical stimulation. However, the ability of these cells to synthesize cyclic AMP was reduced by approximately 50%. Thus, an enhanced level of contraction reduces the coupling efficiency of beta-adrenergic receptors for cyclic AMP production.

  10. Basal and adenosine receptor-stimulated levels of cAMP are reduced in lymphocytes from alcoholic patients

    SciTech Connect

    Diamond, I.; Wrubel, B.; Estrin, W.; Gordon, A.

    1987-03-01

    Alcoholism causes serious neurologic disease that may be due, in part, to the ability of ethanol to interact with neural cell membranes and change neuronal function. Adenosine receptors are membrane-bound proteins that appear to mediate some of the effects of ethanol in the brain. Human lymphocytes also have adenosine receptors, and their activation causes increases in cAMP levels. To test the hypothesis that basal and adenosine receptor-stimulated cAMP levels in lymphocytes might be abnormal in alcoholism, the authors studied lymphocytes from 10 alcoholic subjects, 10 age- and sex-matched normal individuals, and 10 patients with nonalcoholic liver disease. Basal and adenosine receptor-stimulated cAMP levels were reduced 75% in lymphocytes from alcoholic subjects. Also, there was a 76% reduction in ethanol stimulation of cAMP accumulation in lymphocytes from alcoholics. Similar results were demonstrable in isolated T cells. Unlike other laboratory tests examined, these measurements appeared to distinguish alcoholics from normal subjects and from patients with nonalcoholic liver disease. Reduced basal and adenosine receptor-stimulated levels of cAMP in lymphocytes from alcoholics may reflect a change in cell membranes due either to chronic alcohol abuse or to a genetic predisposition unique to alcoholic subjects.

  11. Primary neural involvement in renal haemodynamic and functional responses to prolonged stimulation of atrial receptors in anaesthetized dogs.

    PubMed

    Majid, D S; Karim, F

    1995-07-01

    To determine the precise contributory role of neural and humoral factors in the efferent mechanism of the atrial receptor-renal reflex, we have examined the effects of prolonged (45 min) stimulation of left atrial receptors on renal haemodynamics and function simultaneously in both kidneys (right kidney intact and left kidney denervated) of anaesthetized dogs. Aortic pressure in these dogs was held constant by means of an arterial reservoir connected to the aorta; heart rate changes were prevented by blocking beta 1-adrenoceptor activity with atenolol (2 mg kg-1 i.v.). Localized stimulation of atrial receptors in six dogs increased renal blood flow (6 +/- 2%), creatinine clearance (11 +/- 4%), urine flow (9 +/- 3%), sodium excretion (14 +/- 7%) and osmolal excretion (10 +/- 4%), and decreased free water clearance (14 +/- 7%) in intact kidneys, but led to no changes in denervated kidneys. In an additional four dogs, cooling the vagus nerves to 6-7 degrees C or cutting them in the neck abolished the renal responses to stimulation of atrial receptors in these stabilized preparations. These data clearly demonstrate that the renal responses to prolonged stimulation of atrial receptors are primarily mediated via myelinated vagal afferents and renal sympathetic efferents.

  12. Human cardiac beta1- or beta2-adrenergic receptor stimulation and the negative chronotropic effect of low-dose pirenzepine.

    PubMed

    Jakubetz, J; Schmuck, S; Wochatz, G; Ruhland, B; Poller, U; Radke, J; Brodde, O E

    2000-05-01

    The M1-muscarinic receptor antagonist pirenzepine in low doses (<1 mg intravenously) decreases heart rate. We investigated whether these effects of pirenzepine differ in volunteers with activated cardiac beta1-adrenergic receptors versus activated cardiac beta2-adrenergic receptors. In 17 male volunteers (25 +/- 1 years) we studied effects of pirenzepine infusion (0.5 mg intravenous bolus followed by continuous infusion of 0.15 microg/kg/min) on heart rate and heart rate-corrected duration of electromechanical systole (QS2c, as a measure of inotropism) that had been stimulated by activation of cardiac beta1-adrenergic receptors (bicycle exercise in the supine position for 60 minutes at 25 W) or cardiac beta2-adrenergic receptors (continuous intravenous infusion of 100 ng/kg/min terbutaline). Bicycle exercise and terbutaline infusion significantly increased heart rate and shortened QS2c. When pirenzepine was infused 20 minutes after the beginning of the exercise or terbutaline infusion, heart rate decreased in both settings by approximately the same extent (approximately -10 to -14 beats/min), although exercise and terbutaline infusion continued; however, QS2c was not affected. Pirenzepine (0.05 to 1 mg intravenous bolus)-induced decrease in heart rate was abolished after 6 days of transdermal scopolamine treatment of volunteers. Low-dose pirenzepine decreased heart rate by muscarinic receptor stimulation, because this was blocked by scopolamine. Moreover, low-dose pirenzepine did not differentiate between cardiac beta1- or beta2-adrenergic receptor stimulation; however, low-dose pirenzepine did not affect cardiac contractility as measured by QS2c. Low-dose pirenzepine therefore exerted a unique pattern of action in the human heart: it decreased heart rate (basal and beta1- and/or beta2-adrenergic receptor-stimulated) without affecting contractility.

  13. Gamma hydroxybutyrate (GHB), gamma butyrolactone (GBL) and 1,4-butanediol (1,4-BD; BDO): A literature review with a focus on UK fatalities related to non-medical use.

    PubMed

    Corkery, John M; Loi, Barbara; Claridge, Hugh; Goodair, Christine; Corazza, Ornella; Elliott, Simon; Schifano, Fabrizio

    2015-06-01

    Misuse of gamma hydroxybutrate (GHB) and gamma butyrolactone (GBL) has increased greatly since the early 1990s, being implicated in a rising number of deaths. This paper reviews knowledge on GHB and derivatives, and explores the largest series of deaths associated with their non-medical use. Descriptive analyses of cases associated with GHB/GBL and 1,4-butanediol (1,4-BD) use extracted from the UK's National Programme on Substance Abuse Deaths database. From 1995 to September 2013, 159 GHB/GBL-associated fatalities were reported. Typical victims: White (92%); young (mean age 32 years); male (82%); with a drug misuse history (70%). Most deaths (79%) were accidental or related to drug use, the remainder (potential) suicides. GHB/GBL alone was implicated in 37%; alcohol 14%; other drugs 28%; other drugs and alcohol 15%. Its endogenous nature and rapid elimination limit toxicological detection. Post-mortem blood levels: mean 482 (range 0-6500; SD 758)mg/L. Results suggest significant caution is needed when ingesting GHB/GBL, particularly with alcohol, benzodiazepines, opiates, stimulants, and ketamine. More awareness is needed about risks associated with consumption. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. The structure and organization of the human follicle-stimulating hormone receptor (FSHR) gene

    SciTech Connect

    Gromoll, J; Pekel, E.; Nieschlag, E.

    1996-07-15

    The structure and organization of the human follicle-stimulating hormone receptor (FSHR) gene were determined by either screening a phage library of human genomic DNA or applying the long PCR technique to amplify different exon pairs with their corresponding introns. The FSHR gene spans a region of 54 kb and consists of 10 exons and 9 introns. Most of the extracellular domain is encoded by 9 exons, ranging in length between 69 and 251 bp; the C-terminal part of the extracellular domain, the transmembrane domain, and the intracellular domain are encoded by the large exon 10 (1234 bp). Overall the gene encodes 695 amino acids. The structure of the human FSHR displays a striking similarity to that of the previously characterized rat FSHR gene, with a high degree of conservation in exon sizes and exon/intron junctions. 20 refs., 2 tabs.

  15. Graves' Disease Mechanisms: The Role of Stimulating, Blocking, and Cleavage Region TSH Receptor Antibodies

    PubMed Central

    Morshed, S. A.; Davies, T. F.

    2016-01-01

    The immunologic processes involved in Graves' disease (GD) have one unique characteristic – the autoantibodies to the TSH receptor (TSHR) – which have both linear and conformational epitopes. Three types of TSHR antibodies (stimulating, blocking, and cleavage) with different functional capabilities have been described in GD patients, which induce different signaling effects varying from thyroid cell proliferation to thyroid cell death. The establishment of animal models of GD by TSHR antibody transfer or by immunization with TSHR antigen has confirmed its pathogenic role and, therefore, GD is the result of a breakdown in TSHR tolerance. Here we review some of the characteristics of TSHR antibodies with a special emphasis on new developments in our understanding of what were previously called “neutral” antibodies and which we now characterize as autoantibodies to the “cleavage” region of the TSHR ectodomain. PMID:26361259

  16. Current Concepts of Follicle-Stimulating Hormone Receptor Gene Regulation1

    PubMed Central

    George, Jitu W.; Dille, Elizabeth A.; Heckert, Leslie L.

    2010-01-01

    Follicle-stimulating hormone (FSH), a pituitary glycoprotein hormone, is an integral component of the endocrine axis that regulates gonadal function and fertility. To transmit its signal, FSH must bind to its receptor (FSHR) located on Sertoli cells of the testis and granulosa cells of the ovary. Thus, both the magnitude and the target of hormone response are controlled by mechanisms that determine FSHR levels and cell-specific expression, which are supported by transcription of its gene. The present review examines the status of FSHR/Fshr gene regulation, emphasizing the importance of distal sequences in FSHR/Fshr transcription, new insights gained from the influx of genomics data and bioinformatics, and emerging trends that offer direction in deciphering the FSHR/Fshr regulatory landscape. PMID:20739665

  17. Targeting the thyroid-stimulating hormone receptor with small molecule ligands and antibodies

    PubMed Central

    Davies, Terry F; Latif, Rauf

    2015-01-01

    Introduction The thyroid-stimulating hormone receptor (TSHR) is the essential molecule for thyroid growth and thyroid hormone production. Since it is also a key autoantigen in Graves’ disease and is involved in thyroid cancer pathophysiology, the targeting of the TSHR offers a logical model for disease control. Areas covered We review the structure and function of the TSHR and the progress in both small molecule ligands and TSHR antibodies for their therapeutic potential. Expert opinion Stabilization of a preferential conformation for the TSHR by allosteric ligands and TSHR antibodies with selective modulation of the signaling pathways is now possible. These tools may be the next generation of therapeutics for controlling the pathophysiological consequences mediated by the effects of the TSHR in the thyroid and other extrathyroidal tissues. PMID:25768836

  18. Activating parabrachial cannabinoid CB1 receptors selectively stimulates feeding of palatable foods in rats.

    PubMed

    DiPatrizio, Nicholas V; Simansky, Kenny J

    2008-09-24

    The endocannabinoid system is emerging as an integral component in central and peripheral regulation of feeding and energy balance. Our investigation analyzed behavioral roles for cannabinoid mechanisms of the pontine parabrachial nucleus (PBN) in modulating intake of presumably palatable foods containing fat and/or sugar. The PBN serves to gate neurotransmission associated with, but not limited to, the gustatory properties of food. Immunofluorescence and in vitro [(35)S]GTPgammaS autoradiography of rat tissue sections containing the PBN revealed the presence of cannabinoid receptors and their functional capability to couple to their G-proteins after incubation with the endocannabinoid 2-arachidonoyl glycerol (2-AG). The selective cannabinoid 1 receptor (CB(1)R) antagonist AM251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] prevented the response, demonstrating CB(1)R mediation of 2-AG-induced coupling. Microinfusions of 2-AG into the PBN in behaving rats robustly stimulated feeding of pellets high in content of fat and sucrose (HFS), pure sucrose, and pure fat (Crisco), during the first 30 min after infusion. In contrast, 2-AG failed to increase consumption of standard chow, even when the feeding regimen was manipulated to match baseline intakes of HFS. Orexigenic responses to 2-AG were attenuated by AM251, again indicating CB(1)R mediation of 2-AG actions. Furthermore, responses were regionally specific, because 2-AG failed to alter intake when infused into sites approximately 500 mum caudal to infusions that successfully stimulated feeding. Our data suggest that hedonically positive sensory properties of food enable endocannabinoids at PBN CB(1)Rs to initiate increases in eating, and, more generally, these pathways may serve a larger role in brain functions controlling behavioral responses for natural reward.

  19. Colony-stimulating factor 1 receptor (CSF1R) inhibitors in cancer therapy.

    PubMed

    Cannarile, Michael A; Weisser, Martin; Jacob, Wolfgang; Jegg, Anna-Maria; Ries, Carola H; Rüttinger, Dominik

    2017-07-18

    The tumor-permissive and immunosuppressive characteristics of tumor-associated macrophages (TAM) have fueled interest in therapeutically targeting these cells. In this context, the colony-stimulating factor 1 (CSF1)/colony-stimulating factor 1 receptor (CSF1R) axis has gained the most attention, and various approaches targeting either the ligands or the receptor are currently in clinical development. Emerging data on the tolerability of CSF1/CSF1R-targeting agents suggest a favorable safety profile, making them attractive combination partners for both standard treatment modalities and immunotherapeutic agents. The specificity of these agents and their potent blocking activity has been substantiated by impressive response rates in diffuse-type tenosynovial giant cell tumors, a benign connective tissue disorder driven by CSF1 in an autocrine fashion. In the malignant disease setting, data on the clinical activity of immunotherapy combinations with CSF1/CSF1R-targeting agents are pending. As our knowledge of macrophage biology expands, it becomes apparent that the complex phenotypic and functional properties of macrophages are heavily influenced by a continuum of survival, differentiation, recruitment, and polarization signals within their specific tissue environment. Thus, the role of macrophages in regulating tumorigenesis and the impact of depleting and/or reprogramming TAM as therapeutic approaches for cancer patients may vary greatly depending on organ-specific characteristics of these cells. We review the currently available clinical safety and efficacy data with CSF1/CSF1R-targeting agents and provide a comprehensive overview of ongoing clinical studies. Furthermore, we discuss the local tissue macrophage and tumor-type specificities and their potential impact on CSF1/CSF1R-targeting treatment strategies for the future.

  20. STIMULATION OF TARSAL RECEPTORS OF THE BLOWFLY BY ALIPHATIC ALDEHYDES AND KETONES

    PubMed Central

    Chadwick, L. E.; Dethier, V. G.

    1949-01-01

    Rejection of eight aldehydes, eight ketones, five secondary alcohols, and 3-pentanol has been studied in the blowfly Phormia regina Meigen. The data agree with results previously reported for normal alcohols and several series of glycols in showing a logarithmic increase in stimulating effect with increasing chain length. The order of increasing effectiveness among the different species of compounds thus far investigated is the following: polyglycols, diols, secondary alcohols, iso-alcohols, normal alcohols, ketones, iso-aldehydes, normal aldehydes. Curves relating the logarithms of threshold concentration to the logarithms of chain length for diols, alcohols, aldehydes, and ketones show inflections in the 3 to 6 carbon range. Above and below the region of inflection the curves are nearly rectilinear. The slopes for the upper limbs (smaller molecules) are of the order of –2; for the lower limbs, about –10. Comparisons of the threshold data with numerical values for molecular weights, molecular areas and volumes, oil-water distribution coefficients, activity coefficients, standard free energies, vapor pressures, boiling points, melting points, dipole moments, dielectric constants, and degree of association are discussed briefly, and it is concluded that none of the comparisons serves to bring the data from the several series and from the two portions of each series into a single homogeneous system. A qualitative comparison with water solubilities shows fewer discrepancies. It is suggested that the existence of a combination of aqueous and lipoid phases at the receptor surface would fit best with what is presently known about the relationship between chemical structure and stimulating effect in contact chemoreception. In this hypothesis the smaller and more highly water-soluble compounds are envisaged as gaining access to the receptors partly through the aqueous phase, the larger molecules predominantly through the lipoid phase. PMID:18114559

  1. Nitric oxide production is stimulated by bitter taste receptors ubiquitously expressed in the sinonasal cavity

    PubMed Central

    Yan, Carol H.; Hahn, Samuel; McMahon, Derek; Bonislawski, David; Kennedy, David W.; Adappa, Nithin D.; Palmer, James N.; Jiang, Peihua; Lee, Robert J.

    2017-01-01

    Background: Bitter taste receptors (T2R) have recently been demonstrated to contribute to sinonasal innate immunity. One T2R, T2R38, regulates mucosal defense against gram-negative organisms through nitric oxide (NO) production, which enhances mucociliary clearance and directly kills bacteria. To determine whether additional T2Rs contribute to this innate defense, we evaluated two other sinonasal T2Rs (T2R4 and T2R16) for regulation of NO production and expression within the human sinonasal cavity. Methods: Primary human sinonasal cultures were stimulated with ligands specific to T2R4 and T2R16, colchicine and D-salicin, respectively. Cellular NO production was measured by intracellular 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate fluorescence. For T2R expression mapping, sinonasal tissue was obtained from patients who underwent sinus surgery of the middle turbinate, maxillary sinus, ethmoid sinus, or sphenoid sinus. The expression of T2R4, T2R16, and T2R38 was evaluated by using immunofluorescence with validated antibodies. Results: Similar to T2R38, T2R4 and T2R16 trigger NO production in a dose-dependent manner by using the canonical taste signaling pathway in response to stimulation with their respective ligands. All three receptors were expressed in the cilia of human epithelial cells of all regions in the sinonasal cavity. Conclusion: These three T2Rs signaled through the same NO-mediated antimicrobial pathway and were ubiquitously expressed in the sinonasal epithelium. Additional T2Rs besides T2R38 may play a role in sinonasal immune defense. Mapping of T2R expression demonstrated the potential widespread role of T2Rs in sinonasal defense, whereas the genetics of these T2Rs may contribute to our understanding of specific endotypes of chronic rhinosinusitis and develop into novel therapeutic targets. PMID:28452704

  2. Nitric oxide production is stimulated by bitter taste receptors ubiquitously expressed in the sinonasal cavity.

    PubMed

    Yan, Carol H; Hahn, Samuel; McMahon, Derek; Bonislawski, David; Kennedy, David W; Adappa, Nithin D; Palmer, James N; Jiang, Peihua; Lee, Robert J; Cohen, Noam A

    2017-03-01

    Bitter taste receptors (T2R) have recently been demonstrated to contribute to sinonasal innate immunity. One T2R, T2R38, regulates mucosal defense against gram-negative organisms through nitric oxide (NO) production, which enhances mucociliary clearance and directly kills bacteria. To determine whether additional T2Rs contribute to this innate defense, we evaluated two other sinonasal T2Rs (T2R4 and T2R16) for regulation of NO production and expression within the human sinonasal cavity. Primary human sinonasal cultures were stimulated with ligands specific to T2R4 and T2R16, colchicine and D-salicin, respectively. Cellular NO production was measured by intracellular 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate fluorescence. For T2R expression mapping, sinonasal tissue was obtained from patients who underwent sinus surgery of the middle turbinate, maxillary sinus, ethmoid sinus, or sphenoid sinus. The expression of T2R4, T2R16, and T2R38 was evaluated by using immunofluorescence with validated antibodies. Similar to T2R38, T2R4 and T2R16 trigger NO production in a dose-dependent manner by using the canonical taste signaling pathway in response to stimulation with their respective ligands. All three receptors were expressed in the cilia of human epithelial cells of all regions in the sinonasal cavity. These three T2Rs signaled through the same NO-mediated antimicrobial pathway and were ubiquitously expressed in the sinonasal epithelium. Additional T2Rs besides T2R38 may play a role in sinonasal immune defense. Mapping of T2R expression demonstrated the potential widespread role of T2Rs in sinonasal defense, whereas the genetics of these T2Rs may contribute to our understanding of specific endotypes of chronic rhinosinusitis and develop into novel therapeutic targets.

  3. Androgen stimulates endothelial cell proliferation via an androgen receptor/VEGF/cyclin A-mediated mechanism

    PubMed Central

    Cai, Jingjing; Hong, Yuan; Weng, Chunyan; Tan, Chen; Imperato-McGinley, Julianne

    2011-01-01

    Growing evidences support that androgen displays beneficial effects on cardiovascular functions although the mechanism of androgen actions remains to be elucidated. Modulation of endothelial cell growth and function is a potential mechanism of androgen actions. We demonstrated in the present study that androgens [dihydrotestosterone (DHT) and testosterone], but not 17β-estradiol, produced a time- and dose-dependent induction of cell proliferation in primary human aortic endothelial cells (HAECs) as evident by increases in viable cell number and DNA biosynthesis. Real-time qRT-PCR analysis showed that DHT induced androgen receptor (AR), cyclin A, cyclin D1, and vascular endothelial growth factor (VEGF) gene expression in a dose- and time-dependent manner. The addition of casodex, a specific AR antagonist, or transfection of a specific AR siRNA blocked DHT-induced cell proliferation and target gene expression, indicating that the DHT effects are mediated via AR. Moreover, coadministration of SU5416 to block VEGF receptors, or transfection of a specific VEGF-A siRNA to knockdown VEGF expression, produced a dose-dependent blockade of DHT induction of cell proliferation and cyclin A gene expression. Interestingly, roscovitine, a selective cyclin-dependent kinase inhibitor, also blocked the DHT stimulation of cell proliferation with a selective inhibition of DHT-induced VEGF-A expression. These results indicate that androgens acting on AR stimulate cell proliferation through upregulation of VEGF-A, cyclin A, and cyclin D1 in HAECs, which may be beneficial to cardiovascular functions since endothelial cell proliferation could assist the repair of endothelial injury/damage in cardiovascular system. PMID:21257919

  4. Activation of transmembrane bile acid receptor TGR5 stimulates insulin secretion in pancreatic β cells

    PubMed Central

    Kumar, Divya P.; Rajagopal, Senthilkumar; Mahavadi, Sunila; Mirshahi, Faridoddin; Grider, John R.; Murthy, Karnam S.; Sanyal, Arun J.

    2012-01-01

    Bile acids act as signaling molecules and stimulate the G protein coupled receptor, TGR5, in addition to nuclear farnesoid X receptor to regulate lipid, glucose and energy metabolism. Bile acid induced activation of TGR5 in the enteroendocrine cells promotes glucagon like peptide-1 (GLP-1) release, which has insulinotropic effect in the pancreatic β cells. In the present study, we have identified the expression of TGR5 in pancreatic β cell line MIN6 and also in mouse and human pancreatic islets. TGR5 selective ligands, oleanolic acid (OA) and INT-777 selectively activated Gαs and caused an increase in intracellular cAMP and Ca2+. OA and INT-777 also increased phosphoinositide (PI) hydrolysis and the increase was blocked by NF449 (a selective Gαs inhibitor) or U73122 (PI hydrolysis inhibitor). OA, INT-777 and lithocholic acid increased insulin release in MIN6 and human islets and the increase was inhibited by treatment with NF449, U73122 or BAPTA-AM (chelator of calcium), but not with myristoylated PKI (PKA inhibitor), suggesting that the release is dependent on Gs/cAMP/Ca2+ pathway. 8-pCPT-2′-O-Me-cAMP, a cAMP analogue, which activates Epac, but not PKA also stimulated PI hydrolysis. In conclusion, our study demonstrates that the TGR5 expressed in the pancreatic β cells regulates insulin secretion and highlights the importance of ongoing therapeutic strategies targeting TGR5 in the control of glucose homeostasis. PMID:23022524

  5. Inosine Released from Dying or Dead Cells Stimulates Cell Proliferation via Adenosine Receptors.

    PubMed

    Chen, Jin; Chaurio, Ricardo A; Maueröder, Christian; Derer, Anja; Rauh, Manfred; Kost, Andriy; Liu, Yi; Mo, Xianming; Hueber, Axel; Bilyy, Rostyslav; Herrmann, Martin; Zhao, Yi; Muñoz, Luis E

    2017-01-01

    Many antitumor therapies induce apoptotic cell death in order to cause tumor regression. Paradoxically, apoptotic cells are also known to promote wound healing, cell proliferation, and tumor cell repopulation in multicellular organisms. We aimed to characterize the nature of the regenerative signals concentrated in the micromilieu of dead and dying cells. Cultures of viable melanoma B16F10 cells, mouse fibroblasts, and primary human fibroblast-like synoviocytes (FLS) in the presence of dead and dying cells, their supernatants (SNs), or purified agonists and antagonists were used to evaluate the stimulation of proliferation. Viable cell quantification was performed by either flow cytometry of harvested cells or by crystal violet staining of adherent cells. High-performance liquid chromatography and liquid chromatography coupled with mass spectrometry of cell SNs were deployed to identify the nature of growth-promoting factors. Coimplantation of living cells in the presence of SNs collected from dead and dying cells and specific agonists was used to evaluate tumor growth in vivo. The stimulation of proliferation of few surviving cells by bystander dead cells was confirmed for melanoma cells, mouse fibroblasts, and primary FLS. We found that small soluble molecules present in the protein-free fraction of SNs of dead and dying cells were responsible for the promotion of proliferation. The nucleoside inosine released by dead and dying cells acting via adenosine receptors was identified as putative inducer of proliferation of surviving tumor cells after irradiation and heat treatment. Inosine released by dead and dying cells mediates tumor cell proliferation via purinergic receptors. Therapeutic strategies surmounting this pathway may help to reduce the rate of recurrence after radio- and chemotherapy.

  6. Muscarinic acetylcholine receptor-mediated stimulation of retinal ganglion cell photoreceptors.

    PubMed

    Sodhi, Puneet; Hartwick, Andrew T E

    2016-09-01

    Melanopsin-dependent phototransduction in intrinsically photosensitive retinal ganglion cells (ipRGCs) involves a Gq-coupled phospholipase C (PLC) signaling cascade. Acetylcholine, released in the mammalian retina by starburst amacrine cells, can also activate Gq-PLC pathways through certain muscarinic acetylcholine receptors (mAChRs). Using multielectrode array recordings of rat retinas, we demonstrate that robust spiking responses can be evoked in neonatal and adult ipRGCs after bath application of the muscarinic agonist carbachol. The stimulatory action of carbachol on ipRGCs was a direct effect, as confirmed through calcium imaging experiments on isolated ipRGCs in purified cultures. Using flickering (6 Hz) yellow light stimuli at irradiances below the threshold for melanopsin activation, spiking responses could be elicited in ipRGCs that were suppressed by mAChR antagonism. Therefore, this work identified a novel melanopsin-independent pathway for stimulating sustained spiking in ganglion cell photoreceptors. This mAChR-mediated pathway could enhance ipRGC spiking responses in conditions known to evoke retinal acetylcholine release, such as those involving flickering or moving visual stimuli. Furthermore, this work identifies a pharmacological approach for light-independent ipRGC stimulation that could be targeted by mAChR agonists.

  7. Statins stimulate the production of a soluble form of the receptor for advanced glycation end products

    PubMed Central

    Quade-Lyssy, Patricia; Kanarek, Anna Maria; Baiersdörfer, Markus; Postina, Rolf; Kojro, Elzbieta

    2013-01-01

    The beneficial effects of statin therapy in the reduction of cardiovascular pathogenesis, atherosclerosis, and diabetic complications are well known. The receptor for advanced glycation end products (RAGE) plays an important role in the progression of these diseases. In contrast, soluble forms of RAGE act as decoys for RAGE ligands and may prevent the development of RAGE-mediated disorders. Soluble forms of RAGE are either produced by alternative splicing [endogenous secretory RAGE (esRAGE)] or by proteolytic shedding mediated by metalloproteinases [shed RAGE (sRAGE)]. Therefore we analyzed whether statins influence the production of soluble RAGE. Lovastatin treatment of either mouse alveolar epithelial cells endogenously expressing RAGE or HEK cells overexpressing RAGE caused induction of RAGE shedding, but did not influence secretion of esRAGE from HEK cells overexpressing esRAGE. Lovastatin-induced secretion of sRAGE was also evident after restoration of the isoprenylation pathway, demonstrating a correlation of sterol biosynthesis and activation of RAGE shedding. Lovastatin-stimulated induction of RAGE shedding was completely abolished by a metalloproteinase ADAM10 inhibitor. We also demonstrate that statins stimulate RAGE shedding at low physiologically relevant concentrations. Our results show that statins, due to their cholesterol-lowering effects, increase the soluble RAGE level by inducing RAGE shedding, and by doing this, might prevent the development of RAGE-mediated pathogenesis. PMID:23966666

  8. Promotion of adipogenesis by an EP2 receptor agonist via stimulation of angiogenesis in pulmonary emphysema.

    PubMed

    Tsuji, Takao; Yamaguchi, Kazuhiro; Kikuchi, Ryota; Itoh, Masayuki; Nakamura, Hiroyuki; Nagai, Atsushi; Aoshiba, Kazutetsu

    2014-08-01

    Body weight loss is a common manifestation in patients with chronic obstructive pulmonary disease (COPD), particularly those with severe emphysema. Adipose angiogenesis is a key mediator of adipogenesis and use of pro-angiogenic agents may serve as a therapeutic option for lean COPD patients. Since angiogenesis is stimulated by PGE2, we examined whether ONO-AE1-259, a selective E-prostanoid (EP) 2 receptor agonist, might promote adipose angiogenesis and adipogenesis in a murine model of elastase-induced pulmonary emphysema (EIE mice). Mice were intratracheally instilled with elastase or saline, followed after 4 weeks by intraperitoneal administration of ONO-AE1-259 for 4 weeks. The subcutaneous adipose tissue (SAT) weight decreased in the EIE mice, whereas in the EIE mice treated with ONO-AE1-259, the SAT weight was largely restored, which was associated with significant increases in SAT adipogenesis, angiogenesis, and VEGF protein production. In contrast, ONO-AE1-259 administration induced no alteration in the weight of the visceral adipose tissue. These results suggest that in EIE mice, ONO-AE1-259 stimulated adipose angiogenesis possibly via VEGF production, and thence, adipogenesis. Our data pave the way for the development of therapeutic interventions for weight loss in emphysema patients, e.g., use of pro-angiogenic agents targeting the adipose tissue vascular component.

  9. The angiotensin II-AT1 receptor stimulates reactive oxygen species within the cell nucleus

    SciTech Connect

    Pendergrass, Karl D.; Gwathmey, TanYa M.; Michalek, Ryan D.; Grayson, Jason M.; Chappell, Mark C.

    2009-06-26

    We and others have reported significant expression of the Ang II Type 1 receptor (AT1R) on renal nuclei; thus, the present study assessed the functional pathways and distribution of the intracellular AT1R on isolated nuclei. Ang II (1 nM) stimulated DCF fluorescence, an intranuclear indicator of reactive oxygen species (ROS), while the AT1R antagonist losartan or the NADPH oxidase (NOX) inhibitor DPI abolished the increase in ROS. Dual labeling of nuclei with antibodies against nucleoporin 62 (Nup62) and AT1R or the NADPH oxidase isoform NOX4 revealed complete overlap of the Nup62 and AT1R (99%) by flow cytometry, while NOX4 was present on 65% of nuclei. Treatment of nuclei with a PKC agonist increased ROS while the PKC inhibitor GF109203X or PI3 kinase inhibitor LY294002 abolished Ang II stimulation of ROS. We conclude that the Ang II-AT1R-PKC axis may directly influence nuclear function within the kidney through a redox sensitive pathway.

  10. The G protein-coupled bile acid receptor, TGR5, stimulates gallbladder filling.

    PubMed

    Li, Tingting; Holmstrom, Sam R; Kir, Serkan; Umetani, Michihisa; Schmidt, Daniel R; Kliewer, Steven A; Mangelsdorf, David J

    2011-06-01

    TGR5 is a G protein-coupled bile acid receptor present in brown adipose tissue and intestine, where its agonism increases energy expenditure and lowers blood glucose. Thus, it is an attractive drug target for treating human metabolic disease. However, TGR5 is also highly expressed in gallbladder, where its functions are less well characterized. Here, we demonstrate that TGR5 stimulates the filling of the gallbladder with bile. Gallbladder volume was increased in wild-type but not Tgr5(-/-) mice by administration of either the naturally occurring TGR5 agonist, lithocholic acid, or the synthetic TGR5 agonist, INT-777. These effects were independent of fibroblast growth factor 15, an enteric hormone previously shown to stimulate gallbladder filling. Ex vivo analyses using gallbladder tissue showed that TGR5 activation increased cAMP concentrations and caused smooth muscle relaxation in a TGR5-dependent manner. These data reveal a novel, gallbladder-intrinsic mechanism for regulating gallbladder contractility. They further suggest that TGR5 agonists should be assessed for effects on human gallbladder as they are developed for treating metabolic disease.

  11. β3-Adrenergic receptor stimulation induces E-selectin-mediated adipose tissue inflammation.

    PubMed

    Roth Flach, Rachel J; Matevossian, Anouch; Akie, Thomas E; Negrin, Kimberly A; Paul, Marina T; Czech, Michael P

    2013-01-25

    Inflammation induced by wound healing or infection activates local vascular endothelial cells to mediate leukocyte rolling, adhesion, and extravasation by up-regulation of leukocyte adhesion molecules such as E-selectin and P-selectin. Obesity-associated adipose tissue inflammation has been suggested to cause insulin resistance, but weight loss and lipolysis also promote adipose tissue immune responses. While leukocyte-endothelial interactions are required for obesity-induced inflammation of adipose tissue, it is not known whether lipolysis-induced inflammation requires activation of endothelial cells. Here, we show that β(3)-adrenergic receptor stimulation by CL 316,243 promotes adipose tissue neutrophil infiltration in wild type and P-selectin-null mice but not in E-selectin-null mice. Increased expression of adipose tissue cytokines IL-1β, CCL2, and TNF-α in response to CL 316,243 administration is also dependent upon E-selectin but not P-selectin. In contrast, fasting increases adipose-resident macrophages but not neutrophils, and does not activate adipose-resident endothelium. Thus, two models of lipolysis-induced inflammation induce distinct immune cell populations within adipose tissue and exhibit distinct dependences on endothelial activation. Importantly, our results indicate that β(3)-adrenergic stimulation acts through up-regulation of E-selectin in adipose tissue endothelial cells to induce neutrophil infiltration.

  12. Nicotinic acetylcholine receptor alpha 7 stimulation dampens splenic myelopoiesis and inhibits atherogenesis in Apoe(-/-) mice.

    PubMed

    Al-Sharea, Annas; Lee, Man K S; Whillas, Alexandra; Flynn, Michelle C; Chin-Dusting, Jaye; Murphy, Andrew J

    2017-10-01

    Monocyte levels predict cardiovascular outcomes and play a causal role in atherogenesis. Monocytes can be produced in the spleen and track to the atherosclerotic lesion in significant numbers. The cholinergic system has been shown to have anti-inflammatory actions in the spleen. We aimed to explore whether therapeutic stimulation of the nicotinic acetylcholine receptor alpha 7 (nAChRα7) can suppress atherogenesis. Apoe(-/-) mice were placed on a Western-type diet and treated with bi-daily injections of the nAChRα7 agonist GTS-21 or vehicle every 2-3 days for 8 weeks. GTS-21 caused a reduction in atherosclerosis in the aortic arch and proximal aorta. This also resulted in less plaque macrophages. Moreover, GTS-21 reduced the abundance of blood monocytes, which was caused by inhibition of inflammatory cytokines and extramedullary hematopoiesis in the spleen, along with splenic monocytes. Stimulation of nAChRα7 with GTS-21 reduced atherosclerosis, which was associated with dampened splenic myelopoiesis. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Mineralocorticoid Receptor Stimulation Improves Cognitive Function and Decreases Cortisol Secretion in Depressed Patients and Healthy Individuals

    PubMed Central

    Otte, Christian; Wingenfeld, Katja; Kuehl, Linn K; Kaczmarczyk, Michael; Richter, Steffen; Quante, Arnim; Regen, Francesca; Bajbouj, Malek; Zimmermann-Viehoff, Frank; Wiedemann, Klaus; Hinkelmann, Kim

    2015-01-01

    Memory and executive function are often impaired in patients with major depression, while cortisol secretion is increased. Mineralocorticoid receptors (MR) are abundantly expressed in the hippocampus and in the prefrontal cortex, brain areas critical for memory, executive function, and cortisol inhibition. Here, we investigated whether MR stimulation with fludrocortisone (1) improves memory and executive function and (2) decreases cortisol secretion in depressed patients and healthy individuals. Twenty-four depressed patients without medication and 24 age-, sex-, and education-matched healthy participants received fludrocortisone (0.4 mg) or placebo in a randomized, double-blind, within-subject cross-over design. We measured verbal memory, visuospatial memory, executive function, psychomotor speed, and salivary cortisol secretion during cognitive testing between 1400 and 1700 hours. For verbal memory and executive function, we found better performance after fludrocortisone compared with placebo across groups. No treatment effect on other cognitive domains emerged. Depressed patients performed worse than healthy individuals in psychomotor speed and executive function. No group effect or group × treatment interaction emerged on other cognitive domains. Fludrocortisone decreased cortisol secretion across groups and there was a significant correlation between cortisol inhibition and verbal memory performance. Our data suggest a crucial role of MR in verbal memory and executive function and demonstrate the possibility to improve cognition in depressed patients and healthy individuals through MR stimulation. PMID:25035081

  14. Glucocorticoid Receptor β Stimulates Akt1 Growth Pathway by Attenuation of PTEN*

    PubMed Central

    Stechschulte, Lance A.; Wuescher, Leah; Marino, Joseph S.; Hill, Jennifer W.; Eng, Charis; Hinds, Terry D.

    2014-01-01

    Glucocorticoids (GCs) are known inhibitors of proliferation and are commonly prescribed to cancer patients to inhibit tumor growth and induce apoptosis via the glucocorticoid receptor (GR). Because of alternative splicing, the GR exists as two isoforms, GRα and GRβ. The growth inhibitory actions of GCs are mediated via GRα, a hormone-induced transcription factor. The GRβ isoform, however, lacks helix 12 of the ligand-binding domain and cannot bind GCs. While we have previously shown that GRβ mRNA is responsive to insulin, the role of GRβ in insulin signaling and growth pathways is unknown. In the present study, we show that GRβ suppresses PTEN expression, leading to enhanced insulin-stimulated growth. These characteristics were independent of the inhibitory qualities that have been reported for GRβ on GRα. Additionally, we found that GRβ increased phosphorylation of Akt basally, which was further amplified following insulin treatment. In particular, GRβ specifically targets Akt1 in growth pathways. Our results demonstrate that the GRβ/Akt1 axis is a major player in insulin-stimulated growth. PMID:24817119

  15. Mineralocorticoid receptor stimulation improves cognitive function and decreases cortisol secretion in depressed patients and healthy individuals.

    PubMed

    Otte, Christian; Wingenfeld, Katja; Kuehl, Linn K; Kaczmarczyk, Michael; Richter, Steffen; Quante, Arnim; Regen, Francesca; Bajbouj, Malek; Zimmermann-Viehoff, Frank; Wiedemann, Klaus; Hinkelmann, Kim

    2015-01-01

    Memory and executive function are often impaired in patients with major depression, while cortisol secretion is increased. Mineralocorticoid receptors (MR) are abundantly expressed in the hippocampus and in the prefrontal cortex, brain areas critical for memory, executive function, and cortisol inhibition. Here, we investigated whether MR stimulation with fludrocortisone (1) improves memory and executive function and (2) decreases cortisol secretion in depressed patients and healthy individuals. Twenty-four depressed patients without medication and 24 age-, sex-, and education-matched healthy participants received fludrocortisone (0.4 mg) or placebo in a randomized, double-blind, within-subject cross-over design. We measured verbal memory, visuospatial memory, executive function, psychomotor speed, and salivary cortisol secretion during cognitive testing between 1400 and 1700 hours. For verbal memory and executive function, we found better performance after fludrocortisone compared with placebo across groups. No treatment effect on other cognitive domains emerged. Depressed patients performed worse than healthy individuals in psychomotor speed and executive function. No group effect or group × treatment interaction emerged on other cognitive domains. Fludrocortisone decreased cortisol secretion across groups and there was a significant correlation between cortisol inhibition and verbal memory performance. Our data suggest a crucial role of MR in verbal memory and executive function and demonstrate the possibility to improve cognition in depressed patients and healthy individuals through MR stimulation.

  16. Promoting MPhi transepithelial migration by stimulating the epithelial cell P2Y(2) receptor.

    PubMed

    Langlois, Christine; Gendron, Fernand-Pierre

    2009-10-01

    In intestine, neutrophils are recruited in response to bacterial infiltration and their anti-cellular activities contribute to inflammatory bowel diseases. In contrast, little is known regarding the recruitment of MPhi to the intestinal epithelium. Extracellular adenosine and uridine 5'-triphosphate (ATP and UTP) can function as leukocyte chemoattractants. We investigated the effects of these nucleotides on the ability of intestinal epithelial cells (IEC) to promote MPhi transepithelial migration and adhesion. ATP and UTP promoted the migration of neutrophil-like PLB-985 cells and MPhi across a Caco-2 monolayer. The MPhi-like U-937 cells adhered to nucleotide-stimulated IEC monolayers. In mice with intestinal inflammation, there were infiltrating CD68(+) MPhi in the colonic epithelium and CD68(+) MPhi present at the apical surface of colonocytes. We determined that ATP and UTP activated the P2Y(2) receptor P (P2Y(2)R) to increase ICAM-1 expression, which mediated the adhesion of MPhi to the apical surface of IEC. Intriguingly, stimulation of IEC with nucleotides did not increase the adhesion of neutrophils. However, in the presence of adherent MPhi, there was adhesion of neutrophils, suggesting that MPhi may serve as anchors for neutrophil adhesion. These studies provide insight into the inflammatory mechanisms that contribute to inflammatory bowel diseases and identify potential therapeutic targets for the treatment of gastrointestinal disorders.

  17. Constitutive cholesterol-dependent endocytosis of melanocortin-4 receptor (MC4R) is essential to maintain receptor responsiveness to α-melanocyte-stimulating hormone (α-MSH).

    PubMed

    McDaniel, Faith K; Molden, Brent M; Mohammad, Sameer; Baldini, Giovanna; McPike, Lakisha; Narducci, Paola; Granell, Susana; Baldini, Giulia

    2012-06-22

    Melanocortin-4 receptor (MC4R) is a G-protein-coupled receptor expressed in the hypothalamus where it controls feeding behavior. MC4R cycles constitutively and is internalized at the same rate in the presence or absence of stimulation by the agonist, melanocyte-stimulating hormone (α-MSH). This is different from other G-protein-coupled receptors, such as β(2)-adrenergic receptor (β(2)AR), which internalizes more rapidly in response to agonist stimulation. Here, it is found that in immortalized neuronal Neuro2A cells expressing exogenous receptors, constitutive endocytosis of MC4R and agonist-dependent internalization of β(2)AR were equally sensitive to clathrin depletion. Inhibition of MC4R endocytosis by clathrin depletion decreased the number of receptors at the cell surface that were responsive to the agonist, α-MSH, by 75%. Mild membrane cholesterol depletion also inhibited constitutive endocytosis of MC4R by ∼5-fold, while not affecting recycling of MC4R or agonist-dependent internalization of β(2)AR. Reduced cholesterol did not change the MC4R dose-response curve to α-MSH, but it decreased the amount of cAMP generated per receptor number indicating that a population of MC4R at the cell surface becomes nonfunctional. The loss of MC4R function increased over time (25-50%) and was partially reversed by mutations at putative phosphorylation sites (T312A and S329A). This was reproduced in hypothalamic GT1-7 cells expressing endogenous MC4R. The data indicate that constitutive endocytosis of MC4R is clathrin- and cholesterol-dependent. MC4R endocytosis is required to maintain MC4R responsiveness to α-MSH by constantly eliminating from the plasma membrane a pool of receptors modified at Thr-312 and Ser-329 that have to be cycled to the endosomal compartment to regain function.

  18. Selective Endothelin-B Receptor Stimulation Increases Vascular Endothelial Growth Factor in the Rat Brain during Postnatal Development.

    PubMed

    Leonard, M G; Prazad, P; Puppala, B; Gulati, A

    2015-11-01

    Endothelin, vascular endothelial growth factor and nerve growth factor play important roles in development of the central nervous system. ET(B) receptors have been shown to promote neurovascular remodeling in the adult ischemic brain through an increase in VEGF and NGF. It is possible that ET(B) receptors may be involved in postnatal development of the brain through VEGF and NGF. In the present study, the brains of male rat pups on postnatal days 1, 7, 14 and 28 were analyzed for expression of ET(B) receptors, VEGF and NGF. In order to determine the effect of ET(B) receptor stimulation, a separate group of pups were administered saline or ET(B) receptor agonist, IRL-1620, on day 21, and their brains were analyzed on day 28. The intensity of ET(B) receptor and VEGF staining in the vasculature as well as the number of blood vessels of normal pups increased with age and was significantly higher on postnatal day 14 compared to day 1 and day 7. In contrast, both ET(B) and NGF staining intensity in the cortex and subventricular zones decreased (P<0.01) at postnatal day 14 compared to earlier time points. Stimulation of ET(B) receptors resulted in a significant increase in VEGF and ET(B) intensity both in the vasculature and the brain (P<0.05), however, IRL-1620 did not produce any change in NGF expression. Results indicate that ET(B) receptors appear to play a role in the development of the CNS and selective stimulation of ET(B) receptors enhances VEGF but not NGF in the postnatal rat brain. © Georg Thieme Verlag KG Stuttgart · New York.

  19. 8-iso-PGE2 stimulates anion efflux from airway epithelial cells via the EP4 prostanoid receptor.

    PubMed

    Joy, Andrew P; Cowley, Elizabeth A

    2008-02-01

    Isoprostanes are biologically active molecules, produced when reactive oxygen species mediate the peroxidation of membrane polyunsaturated fatty acids. Previous work has demonstrated that the isoprostane 8-iso-prostaglandin E(2) (PGE(2)) stimulates cystic fibrosis transmembrane conductance regulator (CFTR)-mediated transepithelial anion secretion across the human airway epithelial cell line, Calu-3. Since isoprostanes predominantly achieve their effects via binding to prostanoid receptors, we hypothesized that this 8-iso-PGE(2) stimulation of CFTR activity was the result of the isoprostane binding to a prostanoid receptor. Using RT-PCR, immunoblotting, and immunofluorescence, we here demonstrate that Calu-3 cells express the EP(1-4) and FP receptors, and localize these proteins in polarized cell monolayers. Using iodide efflux as a marker for CFTR-mediated Cl(-) efflux, we investigate whether prostanoid receptor agonists elicit a functional response from Calu-3 cells. Application of the agonists PGE(2), misoprostol (EP(2), EP(3), and EP(4)) and PGE(1)-OH (EP(3) and EP(4)) stimulate iodide efflux; however, iloprost, butaprost, sulprostone, and fluoprostenol (agonists of the EP(1), EP(2), EP(3), and FP receptors, respectively) have no effect. The iodide efflux seen with 8-iso-PGE(2) is abolished by the EP(4) receptor antagonist AH23848, the CFTR inhibitor 172, and inhibition of PKA and the PI3K pathway. In conclusion, we demonstrate that although Calu-3 cells possess numerous prostanoid receptors, only the EP(4) subtype appears capable of eliciting a functional iodide efflux response, which is mediated via the EP(4) receptor. We propose that 8-iso-PGE(2), acting via EP(4) receptor, could play an important role in the CFTR-mediated response to oxidant stress, and which would be compromised in the CF airways.

  20. Individual differences in amygdala reactivity following nicotinic receptor stimulation in abstinent smokers

    PubMed Central

    Sutherland, Matthew T.; Carroll, Allison J.; Jo Salmeron, Betty; Ross, Thomas J.; Hong, L. Elliot; Stein, Elliot A.

    2012-01-01

    Hyperactive amygdala functioning may underlie emotional dysregulation during smoking abstinence and represents one neurobiological target for pharmacological cessation aids. Available pharmacotherapies (e.g., nicotine replacement and varenicline) aid only a subset of individuals with smoking cessation and therefore elucidating the neurobiological impact of these medications is critical to expedite improved interventions. In a fMRI study employing a within-subject, double-blind, placebo-controlled design, we assessed task performance and amygdala functioning during an emotional face matching paradigm following administration of nicotine and varenicline to 24 abstinent smokers and 20 nonsmokers. All participants underwent ~17 days of varenicline and placebo pill administration and were scanned, on different days under each condition, wearing a transdermal nicotine or placebo patch. During the amygdala reactivity paradigm, nicotinic acetylcholine receptor (nAChR) stimulation by nicotine and varenicline decreased reaction time (RT) in abstinent smokers but not in nonsmokers. When considering all smokers as a single homogenous group, no drug-induced effects on amygdala reactivity were detected. However, in an exploratory analysis we parsed participants into subgroups according to individual differences in the propensity to demonstrate stable performance augmentation following nAChR stimulation (stable RT-improvers [SI] vs. variable RT-improvers [VI]). Using this exploratory approach, drugs appeared to modulate amygdala reactivity in only one smoker subgroup but not in either nonsmoker subgroup. Specifically, in the SI-smoker cohort abstinence-induced elevated amygdala reactivity was down-regulated by nAChR stimulation. In contrast, varenicline and nicotine did not modulate amygdala functioning in the VI-smoker cohort who displayed moderate levels of amygdala reactivity in the absence of drug administration. These results suggest that pharmacotherapies most robustly

  1. Mechanisms of amphibian macrophage development: characterization of the Xenopus laevis colony-stimulating factor-1 receptor.

    PubMed

    Grayfer, Leon; Edholm, Eva-Stina; Robert, Jacques

    2014-01-01

    Macrophage-lineage cells are indispensable to vertebrate homeostasis and immunity. In turn, macrophage development is largely regulated through colony-stimulating factor-1 (CSF1) binding to its cognate receptor (CSF1R). To study amphibian monopoiesis, we identified and characterized the X. laevis CSF1R cDNA transcript. Quantitative analysis revealed that CSF1R tissue gene expression increased with X. laevis development, with greatest transcript levels detected in the adult lung, spleen and liver tissues. Notably, considerable levels of CSF1R mRNA were also detected in the regressing tails of metamorphosing animals, suggesting macrophage involvement in this process, and in the adult bone marrow; corroborating the roles for this organ in Xenopus monopoiesis. Following animal infections with the ranavirus Frog Virus 3 (FV3), both tadpole and adult X. laevis exhibited increased kidney CSF1R gene expression. Conversely, while FV3-infected tadpoles increased their spleen and liver CSF1R mRNA levels, the FV3-challenged adults did not. Notably, FV3 induced elevated bone marrow CSF1R expression, and while stimulation of tadpoles with heat-killed E. coli had no transcriptional effects, bacterial stimulation of adult frogs resulted in significantly increased spleen, liver and bone marrow CSF1R expression. We produced the X. laevis CSF1R in recombinant form (rXlCSF1R) and determined, via in vitro cross-linking studies, that two molecules of rXlCSF1R bound the dimeric rXlCSF1. Finally, administration of rXlCSF1R abrogated the rXlCSF1-induced tadpole macrophage recruitment and differentiation as well as bacterial and FV3-elicited peritoneal leukocyte accumulation. This work marks a step towards garnering greater understanding of the unique mechanisms governing amphibian macrophage biology.

  2. Glucose inhibition of epinephrine stimulation of hepatic gluconeogenesis by blockade of the alpha-receptor function.

    PubMed

    Kneer, N M; Bosch, A L; Clark, M G; Lardy, H A

    1974-11-01

    For isolated rat hepatocytes, glucagon, 3':5'-cyclic AMP, 3':5'-cyclic GMP, and epinephrine stimulate the rate of gluconeogenesis from substrates not involving pathways of mitochondrial metabolism. From estimation of the rates of glucose formation, fructose 6-phosphate phosphorylation, and lactate and pyruvate formation it is concluded that epinephrine and 3':5'-cyclic GMP stimulate gluconeogenesis from either galactose or fructose by influencing the rate of reactions involving fructose 6-phosphate in a manner similar to that already reported for glucagon and 3':5'-cyclic AMP. Each agent acts to inhibit flux through phosphofructokinase (EC 2.7.1.11) and enhance flux through fructose diphosphatase (EC 3.1.3.11), resulting in the re-direction of carbon from lactate and pyruvate formation to glucose synthesis. In addition to 3':5'-cyclic GMP, dibutyryl 3':5'-cyclic GMP, 8-bromo 3':5'-cyclic GMP, 8-benzyl-thio 3':5'-cyclic GMP and 8-(4-chlorophenyl)thio 3':5'-cyclic GMP stimulate glucose formation and inhibit lactate and pyruvate formation from galactose. Guanosine monophosphate and 2':3'-cyclic GMP are inactive. As the stimulatory effect of epinephrine is inhibited by phenoxybenzamine and not by propranolol, and is not simulated by isoproterenol, it is concluded that catecholamine activity is expressed through the alpha-receptor. Increased extracellular glucose concentration (>10 mM) decreases the stimulatory effect of epinephrine, 3':5'-cyclic GMP, and partially that of 3':5'-cyclic AMP but does not alter the efficacy of glucagon.

  3. Multi- and single-fibre mesenteric and renal sympathetic responses to chemical stimulation of intestinal receptors in cats.

    PubMed Central

    Stein, R D; Weaver, L C

    1988-01-01

    1. In cats anaesthetized with alpha-chloralose and artificially respired, stimulation of intestinal receptors with bradykinin caused greater reflex excitation of mesenteric than of renal efferent multifibre nerve activity and significant pressor responses. 2. Activity of all nerve bundles used in this study was inhibited by stimulation of pressoreceptors. Increases in systemic arterial pressure caused inhibition of activity of renal nerves which was significantly greater than that of mesenteric nerves. 3. Spinal transection caused significant decreases in tonic renal nerve activity without altering the ongoing discharge rate of mesenteric nerves. Stimulation of intestinal receptors in spinal cats still caused significant increases is discharge of mesenteric and renal nerves, indicating that this reflex contains a spinal component. 4. Recordings of activity of individual fibres within mesenteric (21) and renal (23) nerves provided information regarding the basis for the multifibre responses to stimulation of intestinal receptors. The same proportion of fibres from both nerves was excited, but the increase in activity of mesenteric fibres was significantly greater than that of renal fibres. 5. Mesenteric fibres could be classified into two groups, based on their sensitivity to pressoreceptor influences. Fibres that exhibited pressoreceptor-independent discharge had the greatest responses to stimulation of intestinal receptors. 6. Following spinal transection the majority of mesenteric fibres continued to fire, whereas most renal fibres became quiescent. 7. The non-uniform pattern of neuronal excitation to chemical stimulation of intestinal receptors was manifest after spinal transection, demonstrating that exclusively spinal pathways can mediate this differential response pattern. 8. These results support the hypothesis that viscero-sympathetic reflexes may be organized to cause preferential excitation of neural activity directed to the organ from which the reflex

  4. Effect of blockade of postsynaptic H1 or H2 receptors or activation of presynaptic H3 receptors on catecholamine-induced stimulation of ACTH and prolactin secretion.

    PubMed

    Willems, E; Knigge, U; Jorgensen, H; Kjaer, A; Warberg, J

    2000-06-01

    The effect of inhibition of the neuronal histaminergic system by blockade of postsynaptic H1 or H2 receptors or activation of presynaptic H3 autoreceptors on the ACTH and prolactin responses to the catecholamines epinephrine and norepinephrine was investigated in conscious male rats. Intracerebroventricular infusion of epinephrine and norepinephrine stimulated ACTH and prolactin secretion. Prior intracerebroventricular infusion of the H1 receptor antagonist, mepyramine, or the H2 receptor antagonist, cimetidine, had no effect on the ACTH response to epinephrine or norepinephrine, while these responses were inhibited by pretreatment with the H3 receptor agonist, imetit. The prolactin response to norepinephrine was significantly inhibited by pretreatment with mepyramine, cimetidine or imetit whereas the three histaminergic compounds had no effect on the prolactin response to epinephrine. The findings suggest that the histaminergic system exerts a mediating or permissive action on the norepinephrine-induced stimulation of prolactin secretion, whereas an intact histaminergic system may not be required for catecholamines to stimulate ACTH secretion. The inhibitory effect of imetit on catecholamine-induced release of ACTH may be due to an activation of H3 receptors located presynaptically on non-histaminergic neurons, e.g. aminergic neurons. The study further indicates an important role of histamine in the neuroendocrine regulation of prolactin secretion.

  5. G protein-coupled receptors stimulation and the control of cell migration.

    PubMed

    Cotton, Mathieu; Claing, Audrey

    2009-07-01

    Cell migration is a fundamental biological process involved in normal physiology. Altered motile phenotypes are however often associated with the development and progression of diseases such as cancer and atherosclerosis. Remodeling of the actin cytoskeleton is required for cell shape changes and is controlled by a broad variety of cellular proteins. Interestingly, several extracellular stimuli can promote actin reorganization and result in enhanced cell migration. Namely, G protein-coupled receptors (GPCRs), which are activated by factors ranging from small amines, to hormones, and chemokines, initiate signalling cascades resulting in cell shape changes, formation of a migrating front (leading edge) and altered adhesion. GPCRs are heptahelical membrane proteins, which classically transmit signal via the activation of heterotrimeric G proteins. Sustained stimulation leads to the activation of G protein-coupled receptor kinases (GRKs) and the recruitment of arrestin proteins, which engage alternative signalling pathways. In this review, we will discuss the role of GPCR mediated signal transduction and review their importance in the regulation of actin remodeling leading to cell migration.

  6. Co-receptor and co-stimulation blockade for mixed chimerism and tolerance without myelosuppressive conditioning

    PubMed Central

    Graca, Luis; Daley, Stephen; Fairchild, Paul J; Cobbold, Stephen P; Waldmann, Herman

    2006-01-01

    Background A major challenge in the application of marrow transplantation as a route to immunological tolerance of a transplanted organ is to achieve hematopoietic stem cell (HSC) engraftment with minimal myelosuppressive treatments. Results We here describe a combined antibody protocol which can achieve long-term engraftment with clinically relevant doses of MHC-mismatched bone marrow, without the need for myelosuppressive drugs. Although not universally applicable in all strains, we achieved reliable engraftment in permissive strains with a two-stage strategy: involving first, treatment with anti-CD8 and anti-CD4 in advance of transplantation; and second, treatment with antibodies targeting CD4, CD8 and CD40L (CD154) at the time of marrow transplantation. Long-term mixed chimerism through co-receptor and co-stimulation blockade facilitated tolerance to donor-type skin grafts, without any evidence of donor-antigen driven regulatory T cells. Conclusion We conclude that antibodies targeting co-receptor and co-stimulatory molecules synergise to enable mixed hematopoietic chimerism and central tolerance, showing that neither cytoreductive conditioning nor 'megadoses' of donor bone marrow are required for donor HSC to engraft in permissive strains. PMID:16638128

  7. Activation of macrophages stimulated by the bengkoang fiber extract through toll-like receptor 4.

    PubMed

    Kumalasari, Ika Dyah; Nishi, Kosuke; Putra, Agus Budiawan Naro; Sugahara, Takuya

    2014-07-25

    Bengkoang (Pachyrhizus erosus (L.) Urban) is an edible root tuber containing fairly large amounts of carbohydrates and crude fibers. Our previous studies showed that the bengkoang fiber extract (BFE) stimulates activation of macrophages, leading to induction of phagocytotic activity and cytokine production. In the present study we investigated the mechanism underlying activation of murine macrophages by BFE. BFE increased production of TNF-α, IL-6, and nitric oxide by J774.1 cells. In addition BFE also facilitated the gene expression levels of inducible nitric oxide synthase. We examined the effect of a TLR4 inhibitor on cytokine production to investigate the membrane receptor of macrophage activation by BFE. Treatment of J774.1 cells with the TLR4 inhibitor significantly inhibited production of IL-6 and TNF-α, suggesting that TLR4 is the target membrane receptor for BFE. The main signal molecules located downstream of TLR4 such as JNK, p38, ERK, and NF-κB were activated by BFE treatment. The immunostimulatory effect of BFE was cancelled by the pectinase treatment, suggesting that the active ingredient in BFE is pectin-like molecules. Overall results suggested that BFE activates J774.1 cells via the MAPK and NF-κB signaling pathways.

  8. Angiopoietin-like proteins stimulate HSPC development through interaction with notch receptor signaling

    PubMed Central

    Lin, Michelle I; Price, Emily N; Boatman, Sonja; Hagedorn, Elliott J; Trompouki, Eirini; Satishchandran, Sruthi; Carspecken, Charles W; Uong, Audrey; DiBiase, Anthony; Yang, Song; Canver, Matthew C; Dahlberg, Ann; Lu, Zhigang; Zhang, Cheng Cheng; Orkin, Stuart H; Bernstein, Irwin D; Aster, Jon C; White, Richard M; Zon, Leonard I

    2015-01-01

    Angiopoietin-like proteins (angptls) are capable of ex vivo expansion of mouse and human hematopoietic stem and progenitor cells (HSPCs). Despite this intriguing ability, their mechanism is unknown. In this study, we show that angptl2 overexpression is sufficient to expand definitive HSPCs in zebrafish embryos. Angptl1/2 are required for definitive hematopoiesis and vascular specification of the hemogenic endothelium. The loss-of-function phenotype is reminiscent of the notch mutant mindbomb (mib), and a strong genetic interaction occurs between angptls and notch. Overexpressing angptl2 rescues mib while overexpressing notch rescues angptl1/2 morphants. Gene expression studies in ANGPTL2-stimulated CD34+ cells showed a strong MYC activation signature and myc overexpression in angptl1/2 morphants or mib restored HSPCs formation. ANGPTL2 can increase NOTCH activation in cultured cells and ANGPTL receptor interacted with NOTCH to regulate NOTCH cleavage. Together our data provide insight to the angptl-mediated notch activation through receptor interaction and subsequent activation of myc targets. DOI: http://dx.doi.org/10.7554/eLife.05544.001 PMID:25714926

  9. A novel calcium-sensing receptor antagonist transiently stimulates parathyroid hormone secretion in vivo.

    PubMed

    Arey, Brian J; Seethala, Ramakrishna; Ma, Zhengping; Fura, Aberra; Morin, Jennifer; Swartz, Joann; Vyas, Viral; Yang, Wu; Dickson, John K; Feyen, Jean H M

    2005-04-01

    Circulating calcium (Ca(2+)) is a primary regulator of bone homeostasis through its action on PTH secretion. Extracellular Ca(2+) modulates PTH secretion through a cell surface G protein-coupled receptor, the calcium-sensing receptor (CaR). The expression of the CaR suggests a critical role in cellular regulation by calcium in various organs, including parathyroid gland, bone, and kidney. Despite an obvious pharmacological utility for CaR antagonists in the treatment of disease, only a limited number of such classes of compounds exist. We have identified a novel class of small molecules with specific activity at the CaR. This class of compounds is represented by compound 1. It possesses potent antagonist activity at the human CaR with IC(50) values of 64 nm and 230 nm in inhibiting intracellular Ca(2+) flux and inositol phosphate generation in vitro, respectively. When administered to male rats in vivo, compound 1 robustly increased serum PTH levels. The stimulation of PTH secretion was rapid and transient when administered either iv or orally. The pharmacokinetic profile of compound 1 after oral administration revealed that maximal plasma levels of compound were reached within 1 h and the half-life of the compound to be approximately 2 h in rats. These data describe a representative compound of a novel chemical class than previously described allosteric modulators that offer a new avenue for the development of improved treatments of osteoporosis.

  10. Angiopoietin-like proteins stimulate HSPC development through interaction with notch receptor signaling.

    PubMed

    Lin, Michelle I; Price, Emily N; Boatman, Sonja; Hagedorn, Elliott J; Trompouki, Eirini; Satishchandran, Sruthi; Carspecken, Charles W; Uong, Audrey; DiBiase, Anthony; Yang, Song; Canver, Matthew C; Dahlberg, Ann; Lu, Zhigang; Zhang, Cheng Cheng; Orkin, Stuart H; Bernstein, Irwin D; Aster, Jon C; White, Richard M; Zon, Leonard I

    2015-02-25

    Angiopoietin-like proteins (angptls) are capable of ex vivo expansion of mouse and human hematopoietic stem and progenitor cells (HSPCs). Despite this intriguing ability, their mechanism is unknown. In this study, we show that angptl2 overexpression is sufficient to expand definitive HSPCs in zebrafish embryos. Angptl1/2 are required for definitive hematopoiesis and vascular specification of the hemogenic endothelium. The loss-of-function phenotype is reminiscent of the notch mutant mindbomb (mib), and a strong genetic interaction occurs between angptls and notch. Overexpressing angptl2 rescues mib while overexpressing notch rescues angptl1/2 morphants. Gene expression studies in ANGPTL2-stimulated CD34(+) cells showed a strong MYC activation signature and myc overexpression in angptl1/2 morphants or mib restored HSPCs formation. ANGPTL2 can increase NOTCH activation in cultured cells and ANGPTL receptor interacted with NOTCH to regulate NOTCH cleavage. Together our data provide insight to the angptl-mediated notch activation through receptor interaction and subsequent activation of myc targets.

  11. Chronic cannabinoid receptor stimulation selectively prevents motor impairments in a mouse model of Huntington's disease.

    PubMed

    Pietropaolo, Susanna; Bellocchio, Luigi; Ruiz-Calvo, Andrea; Cabanas, Magali; Du, Zhuowei; Guzmán, Manuel; Garret, Maurice; Cho, Yoon H

    2015-02-01

    Huntington's disease (HD) is a devastating neurodegenerative disease characterized by a progressive decline in motor abilities, as well as in cognitive and social behaviors. Most of these behavioral deficits are recapitulated in the R6/1 transgenic mouse, which can therefore be used as an experimental model to identify the neurobiological substrates of HD pathology and to design novel therapeutic approaches. The endocannabinoid system (ECS) is a relevant candidate to participate in the etiopathology of HD as it is a key modulator of brain function, especially in areas primarily affected by HD dysfunction such as the striatum. Thus, some studies have demonstrated an association between HD progression and alterations in the expression of several ECS elements, thereby suggesting that improving ECS function may constitute a useful strategy to eliminate or at least delay the appearance of HD symptoms. Here this hypothesis was specifically tested by evaluating whether the administration of a well-characterized cannabinoid receptor agonist (WIN 55,212), either acutely or chronically, improves the HD-like symptoms in R6/1 mice. While acute treatment did not change the behavioral phenotype of transgenic animals, chronic administration was able to prevent the appearance of motor deficits, to increase the number of striatal huntingtin inclusions and to prevent the loss of striatal medium-sized spiny neurons, without affecting the social or cognitive alterations. These findings suggest that prolonged administration of cannabinoid receptor agonists could be an appropriate strategy for selectively improving motor symptoms and stimulating neuroprotective processes in HD patients.

  12. Effects of peripheral and spinal κ-opioid receptor stimulation on the exercise pressor reflex in decerebrate rats

    PubMed Central

    Stone, Audrey J.; Yamauchi, Katsuya; Kaufman, Marc P.

    2014-01-01

    The exercise pressor reflex is greater in rats with ligated femoral arteries than it is in rats with freely perfused femoral arteries. The exaggerated reflex in rats with ligated arteries is attenuated by stimulation of μ-opioid and δ-opioid receptors on the peripheral endings of thin-fiber muscle afferents. The effect of stimulation of κ-opioid receptors on the exercise pressor reflex is unknown. We tested the hypothesis that stimulation of κ-opioid receptors attenuates the exercise pressor reflex in rats with ligated, but not freely perfused, femoral arteries. The pressor responses to static contraction were compared before and after femoral arterial or intrathecal injection of the κ-opioid receptor agonist U62066 (1, 10, and 100 μg). Femoral arterial injection of U62066 did not attenuate the pressor responses to contraction in either group of rats. Likewise, intrathecal injection of U62066 did not attenuate the pressor response to contraction in rats with freely perfused femoral arteries. In contrast, intrathecal injection of 10 and 100 μg of U62066 attenuated the pressor response to contraction in rats with ligated femoral arteries, an effect that was blocked by prior intrathecal injection of the κ-opioid receptor antagonist nor-binaltorphimine. In rats with ligated femoral arteries, the pressor response to stimulation of peripheral chemoreceptors by sodium cyanide was not changed by intrathecal U62066 injections, indicating that these injections had no direct effect on the sympathetic outflow. We conclude that stimulation of spinal, but not peripheral, κ-opioid receptors attenuates the exaggerated exercise pressor reflex in rats with ligated femoral arteries. PMID:24920732

  13. Stimulation of ANP secretion by 2-Cl-IB-MECA through A(3) receptor and CaMKII.

    PubMed

    Yuan, Kuichang; Bai, Guang Yi; Park, Woo Hyun; Kim, Sung Zoo; Kim, Suhn Hee

    2008-12-01

    Adenosine is a potent mediator of myocardial protection against hypertrophy via A(1) or A(3) receptors that may be partly related to atrial natriuretic peptide (ANP) release. However, little is known about the possible involvement of the A(3) receptor on ANP release. We studied the effects of the A(3) receptor on atrial functions and its modification in hypertrophied atria. A selective A(3) receptor agonist, 2-chloro-N(6)-(3-iodobenzyl) adenosine-5'-N-methyluronamide (2-CI-IB-MECA), was perfused into isolated, beating rat atria with and without receptor modifiers. 2-CI-IB-MECA dose-dependently increased the ANP secretion, which was blocked by the A(3) receptor antagonist, but the increased atrial contractility and decreased cAMP levels induced by 30muM 2-CI-IB-MECA were not affected. The 100muM 2-(1-hexylnyl)-N-methyladenosine (HEMADO) and N(6)-(3-iodobenzyl) adenosine-5'-N-methyluronamide (IB-MECA), A(3) receptor agonist, also stimulated the ANP secretion without positive inotropy. The potency for the stimulation of ANP secretion was 2-CI-IB-MECA>IB-MECA=HEMADO. The inhibition of the ryanodine receptor or calcium/calmodulin-dependent kinase II (CaMKII) attenuated 2-CI-IB-MECA-induced ANP release, positive inotropy, and translocation of extracellular fluid. However, the inhibition of L-type Ca(2+) channels, sarcoplasmic reticulum Ca(2+)-reuptake, phospholipase C or inositol 1,4,5-triphosphate receptors did not affect these parameters. 2-CI-IB-MECA decreased cAMP level, which was blocked only with an inhibitor of CaMKII or adenylyl cyclase. These results suggest that 2-CI-IB-MECA increases the ANP secretion mainly via A(3) receptor activation and positive inotropy by intracellular Ca(2+) regulation via the ryanodine receptor and CaMKII.

  14. Iron Mediates N-Methyl-d-aspartate Receptor-dependent Stimulation of Calcium-induced Pathways and Hippocampal Synaptic Plasticity*

    PubMed Central

    Muñoz, Pablo; Humeres, Alexis; Elgueta, Claudio; Kirkwood, Alfredo; Hidalgo, Cecilia; Núñez, Marco T.

    2011-01-01

    Iron deficiency hinders hippocampus-dependent learning processes and impairs cognitive performance, but current knowledge on the molecular mechanisms underlying the unique role of iron in neuronal function is sparse. Here, we investigated the participation of iron on calcium signal generation and ERK1/2 stimulation induced by the glutamate agonist N-methyl-d-aspartate (NMDA), and the effects of iron addition/chelation on hippocampal basal synaptic transmission and long-term potentiation (LTP). Addition of NMDA to primary hippocampal cultures elicited persistent calcium signals that required functional NMDA receptors and were independent of calcium influx through L-type calcium channels or α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors; NMDA also promoted ERK1/2 phosphorylation and nuclear translocation. Iron chelation with desferrioxamine or inhibition of ryanodine receptor (RyR)-mediated calcium release with ryanodine-reduced calcium signal duration and prevented NMDA-induced ERK1/2 activation. Iron addition to hippocampal neurons readily increased the intracellular labile iron pool and stimulated reactive oxygen species production; the antioxidant N-acetylcysteine or the hydroxyl radical trapper MCI-186 prevented these responses. Iron addition to primary hippocampal cultures kept in calcium-free medium elicited calcium signals and stimulated ERK1/2 phosphorylation; RyR inhibition abolished these effects. Iron chelation decreased basal synaptic transmission in hippocampal slices, inhibited iron-induced synaptic stimulation, and impaired sustained LTP in hippocampal CA1 neurons induced by strong stimulation. In contrast, iron addition facilitated sustained LTP induction after suboptimal tetanic stimulation. Together, these results suggest that hippocampal neurons require iron to generate RyR-mediated calcium signals after NMDA receptor stimulation, which in turn promotes ERK1/2 activation, an essential step of sustained LTP. PMID:21296883

  15. Sweet taste receptors in rat small intestine stimulate glucose absorption through apical GLUT2

    PubMed Central

    Mace, Oliver J; Affleck, Julie; Patel, Nick; Kellett, George L

    2007-01-01

    Natural sugars and artificial sweeteners are sensed by receptors in taste buds. T2R bitter and T1R sweet taste receptors are coupled through G-proteins, α-gustducin and transducin, to activate phospholipase C β2 and increase intracellular calcium concentration. Intestinal brush cells or solitary chemosensory cells (SCCs) have a structure similar to lingual taste cells and strongly express α-gustducin. It has therefore been suggested over the last decade that brush cells may participate in sugar sensing by a mechanism analogous to that in taste buds. We provide here functional evidence for an intestinal sensing system based on lingual taste receptors. Western blotting and immunocytochemistry revealed that all T1R members are expressed in rat jejunum at strategic locations including Paneth cells, SCCs or the apical membrane of enterocytes; T1Rs are colocalized with each other and with α-gustducin, transducin or phospholipase C β2 to different extents. Intestinal glucose absorption consists of two components: one is classical active Na+–glucose cotransport, the other is the diffusive apical GLUT2 pathway. Artificial sweeteners increase glucose absorption in the order acesulfame potassium ∼ sucralose > saccharin, in parallel with their ability to increase intracellular calcium concentration. Stimulation occurs within minutes by an increase in apical GLUT2, which correlates with reciprocal regulation of T1R2, T1R3 and α-gustducin versus T1R1, transducin and phospholipase C β2. Our observation that artificial sweeteners are nutritionally active, because they can signal to a functional taste reception system to increase sugar absorption during a meal, has wide implications for nutrient sensing and nutrition in the treatment of obesity and diabetes. PMID:17495045

  16. Stimulation of lactate receptor (HCAR1) affects cellular DNA repair capacity.

    PubMed

    Wagner, Waldemar; Kania, Katarzyna D; Ciszewski, Wojciech M

    2017-04-01

    Numerous G-protein coupled receptors have been reported to enhance cancer cell survival and resistance to clinically used chemotherapeutics. Recently, hydroxycarboxylic acid receptor 1 (HCAR1) was shown to drive lactate-dependent enhancement of cell survival and metastasis in pancreatic and breast cancers. Furthermore, our previous study confirmed the involvement of HCAR1 in lactate-related enhancement of DNA repair in cervical cancer cells. In the present study, we examined the possible mechanisms of HCAR1-mediated enhancement of DNA repair capacity. We observed that the HCAR1 agonist dihydroxybenzoic acid (DHBA) up-regulated BRCA1 (breast cancer type 1 susceptibility protein) and NBS1 (Nijmegen breakage syndrome 1) expression in HeLa cells. Moreover, HCAR1 silencing decreased mRNA and protein levels of BRCA1 by 30% and 20%, respectively. Immunocytochemical analyses of BRCA1, nibrin and DNA-PKcs indicated an increased accumulation of these proteins in cell nuclei after DHBA stimulation. Subsequently, these changes in the DNA repair protein levels translated into an enhanced DNA repair rate after doxorubicin treatment, as shown by γ-H2AX and comet assay experiments. In contrast, the down-regulation of HCAR1 decreased the efficiency of DNA repair. Finally, we observed the abrogation of DHBA-driven BRCA1 protein up-regulation and enhanced DNA repair following the preincubation of cells with the PKC inhibitor Gö6983. Taken together, our data indicate that lactate receptor/HCAR1 expression in cervical carcinoma cells may contribute to the modulation of cellular DNA repair mechanisms. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Repeated chlorpromazine administration increases a behavioural response of rats to 5-hydroxytryptamine receptor stimulation.

    PubMed Central

    Green, A R

    1977-01-01

    1 The hyperactivity syndrome produced in rats by administration of tranylcypromine (20 mg/kg i.p.) followed 30 min later by L-tryptophan (50 mg/kg i.p.) is generally considered to be due to increased 5-hydroxytryptamine (5-HT) functional activity. It is inhibited by chlorpromazine (30 mg/kg i.p.) injected 60 min before the tranylcypromine. However, chlorpromazine injection for 4 days either at a dose of 30 mg/kg once daily or 5 mg/kg twice daily results in an enhanced hyperactivity response to tranylcypromine and L-tryptophan administration 24 h after the final dose of chlorpromazine. 2 One injection of chlorpromazine (30 mg/kg) did not produce enhancement 24 h later and the inhibition of the tranylcypromine/L-tryptophan hyperactivity observed after acute chlorpromazine injection was seen if the rats were given tranylcypromine and L-tryptophan 1 h after the fourth chlorpromazine (30 mg/kg) dose. 3 Chlorpromazine (30 mg/kg) once daily or 5 mg/kg twice daily for 4 days resulted in rats displaying enhanced behavioral responses to the suggested 5-HT agonist 5-methoxy N,N-dimethyltryptamine (2 mg/kg) on day 5. 4 Chlorpromazine (30 mg/kg) once daily for 4 days produces a slight increase in brain 5-hydroxytryptamine (5-HT) concentration on day 5, but no difference in the rate of brain 5-HT synthesis or the rate of 5-HT accumulation after tranylcypromine and L-tryptophan administration. 5. There is some evidence that chlorpromazine blocks 5-HT receptors. It has also been observed that several other neuroleptic drugs do not produce enhanced 5-HT responses after repeated administration. It is suggested therefore that the enhanced behavioural response to 5-HT receptor stimulation following repeated chlorpromazine administration may be because this drug blocks 5-HT receptors. PMID:264797

  18. Prefrontal D2-receptor stimulation mediates flexible adaptation of economic preference hierarchies.

    PubMed

    van Eimeren, Thilo; Ko, Ji H; Pellechia, Giovanna; Cho, Sang S; Houle, Sylvain; Strafella, Antonio P

    2013-01-01

    Advantageous economic decision making requires flexible adaptation of gain-based and loss-based preference hierarchies. However, where the neuronal blueprints for economic preference hierarchies are kept and how they may be adapted remains largely unclear. Phasic cortical dopamine release likely mediates flexible adaptation of neuronal representations. In this PET study, cortical-binding potential (BP) for the D(2)-dopamine receptor ligand [(11)C]FLB 457 was examined in healthy participants during multiple sessions of a probabilistic four-choice financial decision-making task with two behavioral variants. In the changing-gains/constant-losses variant, the implicit gain-based preference hierarchy was unceasingly changing, whereas the implicit loss-based preference hierarchy was constant. In the constant-gains/changing-losses variant, it was the other way around. These variants served as paradigms, respectively, contrasting flexible adaptation versus maintenance of loss-based and gain-based preference hierarchies. We observed that in comparison with the constant-gains/changing-losses variant, the changing-gains/constant-losses variant was associated with a decreased D(2)-dopamine receptor-BP in the right lateral frontopolar cortex. In other words, lateral frontopolar D(2)-dopamine receptor stimulation was specifically increased during continuous adaptation of mental representations of gain-based preference hierarchies. This finding provides direct evidence for the existence of a neuronal blueprint of gain-based decision-making in the lateral frontopolar cortex and a crucial role of local dopamine in the flexible adaptation of mental concepts of future behavior. Copyright © 2011 Wiley Periodicals, Inc.

  19. L1CAM stimulates glioma cell motility and proliferation through the fibroblast growth factor receptor.

    PubMed

    Mohanan, Vishnu; Temburni, Murali K; Kappes, John C; Galileo, Deni S

    2013-04-01

    The L1CAM cell adhesion/recognition molecule (L1, CD171) and fibroblast growth factor receptor (FGFR) both are expressed by human high-grade glioma cells, but their potential actions in controlling cell behavior have not been linked. L1 actions in cancer cells have been attributed mainly to integrin receptors, and we demonstrated previously that L1-stimulated glioma cell migration correlates with integrin expression, increased focal adhesion kinase activation and focal complex turnover. Our analyses of datasets revealed FGFR is overexpressed in glioma regardless of grade, while ADAM10 metalloprotease expression increases with glioma grade. Here, we used dominant-negative and short hairpin RNA approaches to inhibit the activation of FGFR1 and expression of L1, respectively. An L1 peptide that inhibits L1-FGFR interaction and PD173074, a chemical inhibitor of FGFR1 activity, also were used to elucidate the involvement of L1-FGFR interactions on glioma cell behavior. Time-lapse cell motility studies and flow cytometry cell cycle analyses showed that L1 operates to increase glioma cell motility and proliferation through FGFR activation. Shutdown of both L1 expression and FGFR activity in glioma cells resulted in a complete termination of cell migration in vitro. These studies show for the first time that soluble L1 ectodomain (L1LE) acts on glioma cells through FGFRs, and that FGFRs are used by glioma cells for increasing motility as well as proliferation in response to activation by L1LE ligand. Thus, effective treatment of high-grade glioma may require simultaneous targeting of L1, FGFRs, and integrin receptors, which would reduce glioma cell motility as well as proliferation.

  20. Liver-Specific Loss of Lipolysis-Stimulated Lipoprotein Receptor Triggers Systemic Hyperlipidemia in Mice

    PubMed Central

    Narvekar, Prachiti; Berriel Diaz, Mauricio; Krones-Herzig, Anja; Hardeland, Ulrike; Strzoda, Daniela; Stöhr, Sigrid; Frohme, Marcus; Herzig, Stephan

    2009-01-01

    OBJECTIVE In mammals, proper storage and distribution of lipids in and between tissues is essential for the maintenance of energy homeostasis. In contrast, aberrantly high levels of triglycerides in the blood (“hypertriglyceridemia”) represent a hallmark of the metabolic syndrome and type 2 diabetes. As hypertriglyceridemia has been identified as an important risk factor for cardiovascular complications, in this study we aimed to identify molecular mechanisms in aberrant triglyceride elevation under these conditions. RESEARCH DESIGN AND METHODS To determine the importance of hepatic lipid handling for systemic dyslipidemia, we profiled the expression patterns of various hepatic lipid transporters and receptors under healthy and type 2 diabetic conditions. A differentially expressed lipoprotein receptor was functionally characterized by generating acute, liver-specific loss- and gain-of-function animal models. RESULTS We show that the hepatic expression of lipid transporter lipolysis-stimulated lipoprotein receptor (LSR) is specifically impaired in mouse models of obesity and type 2 diabetes and can be restored by leptin replacement. Experimental imitation of this pathophysiological situation by liver-specific knockdown of LSR promotes hypertriglyceridemia and elevated apolipoprotein (Apo)B and E serum levels in lean wild-type and ApoE knockout mice. In contrast, genetic restoration of LSR expression in obese animals to wild-type levels improves serum triglyceride levels and serum profiles in these mice. CONCLUSIONS The dysregulation of hepatic LSR under obese and diabetic conditions may provide a molecular rationale for systemic dyslipidemia in type 2 diabetes and the metabolic syndrome and represent a novel target for alternative treatment strategies in these patients. PMID:19188430

  1. Vasopressin/V2 receptor stimulates renin synthesis in the collecting duct

    PubMed Central

    Cifuentes-Araneda, Flavia; Ibaceta-Gonzalez, Cristobal; Gonzalez-Vergara, Alex; Zamora, Leonardo; Henriquez, Ricardo; Rosales, Carla B.; Gabriel Navar, L.; Prieto, Minolfa C.

    2015-01-01

    Renin is synthesized in the principal cells of the collecting duct (CD), and its production is increased via cAMP in angiotensin (ANG) II-dependent hypertension, despite suppression of juxtaglomerular (JG) renin. Vasopressin, one of the effector hormones of the renin-angiotensin system (RAS) via the type 2-receptor (V2R), activates the cAMP/PKA/cAMP response element-binding protein (CREB) pathway and aquaporin-2 expression in principal cells of the CD. Accordingly, we hypothesized that activation of V2R increases renin synthesis via PKA/CREB, independently of ANG II type 1 (AT1) receptor activation in CD cells. Desmopressin (DDAVP; 10−6 M), a selective V2R agonist, increased renin mRNA (∼3-fold), prorenin (∼1.5-fold), and renin (∼2-fold) in cell lysates and cell culture media in the M-1 CD cell line. Cotreatment with DDAVP+H89 (PKA inhibitor) or CREB short hairpin (sh) RNA prevented this response. H89 also blunted DDAVP-induced CREB phosphorylation and nuclear localization. In 48-h water-deprived (WD) mice, prorenin-renin protein levels were increased in the renal inner medulla (∼1.4- and 1.8-fold). In WD mice treated with an ACE inhibitor plus AT1 receptor blockade, renin mRNA and prorenin protein levels were still higher than controls, while renin protein content was not changed. In M-1 cells, ANG II or DDAVP increased prorenin-renin protein levels; however, there were no further increases by combined treatment. These results indicate that in the CD the activation of the V2R stimulates renin synthesis via the PKA/CREB pathway independently of RAS, suggesting a critical role for vasopressin in the regulation of renin in the CD. PMID:26608789

  2. Mice with Deletion of Neuromedin B Receptor Exhibit Decreased Oral Glucose-Stimulated Insulin Release.

    PubMed

    Paula, G S M; Souza, L L; Bressane, N O S; Maravalhas, R; Wilieman, M; Bento-Bernardes, T; Silva, K R; Mendonca, L S; Oliveira, K J; Pazos-Moura, C C

    2016-12-01

    Neuromedin B (NB) and gastrin-releasing peptide (GRP) are bombesin-like peptides, found in the gastrointestinal tube and pancreas, among other tissues. Consistent data proposed that GRP stimulates insulin secretion, acting directly in pancreatic cells or in the release of gastrointestinal hormones that are incretins. However, the role of NB remains unclear. We examined the glucose homeostasis in mice with deletion of NB receptor (NBR-KO). Female NBR-KO exhibited similar fasting basal glucose with lower insulinemia (48.4%) and lower homeostasis model assessment of insulin resistance index (50.5%) than wild type (WT). Additionally, they were more tolerant to oral glucose, demonstrated by a decrease in the area under the glucose curve (18%). In addition, 15 min after an oral glucose load, female and male NBR-KO showed lower insulin serum levels (45.6 and 26.8%, respectively) than WT, even though blood glucose rose to similar levels in both groups. Single injection of NB, one hour before the oral glucose administration, tended to induce higher serum insulin in WT (28.9%, p=0.3), however the same did not occur in NBR-KO. They showed no changes in fasting insulin content in pancreatic islets by immunohistochemistry, however, the fasting serum levels of glucagon-like peptide, a potent incretin, exhibited a strong trend to reduction (40%, p=0.07). Collectively, mice with deletion of NB receptor have lower insulinemia, especially in response to oral glucose, and females also exhibited a better glucose tolerance, suggesting the involvement of NB and its receptor in regulation of insulin secretion induced by incretins, and also, in insulin sensitivity. © Georg Thieme Verlag KG Stuttgart · New York.

  3. Homeobox A7 stimulates breast cancer cell proliferation by up-regulating estrogen receptor-alpha

    SciTech Connect

    Zhang, Yu; Cheng, Jung-Chien; Huang, He-Feng; Leung, Peter C.K.

    2013-11-01

    Highlights: •HOXA7 regulates MCF7 cell proliferation. •HOXA7 up-regulates ERα expression. •HOXA7 mediates estrogen-induced MCF7 cell proliferation. -- Abstract: Breast cancer is the most common hormone-dependent malignancy in women. Homeobox (HOX) transcription factors regulate many cellular functions, including cell migration, proliferation and differentiation. The aberrant expression of HOX genes has been reported to be associated with human reproductive cancers. Estradiol (E2) and its nuclear receptors, estrogen receptor (ER)-alpha and ER-beta, are known to play critical roles in the regulation of breast cancer cell growth. However, an understanding of the potential relationship between HOXA7 and ER in breast cancer cells is limited. In this study, our results demonstrate that knockdown of HOXA7 in MCF7 cells significantly decreased cell proliferation and ERα expression. In addition, HOXA7 knockdown attenuated E2-induced cell proliferation as well as progesterone receptor (PR) expression. The stimulatory effects of E2 on cell proliferation and PR expression were abolished by co-treatment with ICI 182780, a selective ERα antagonist. In contrast, overexpression of HOXA7 significantly stimulated cell proliferation and ERα expression. Moreover, E2-induced cell proliferation, as well as PR expression, was enhanced by the overexpression of HOXA7. Neither knockdown nor overexpression of HOXA7 affected the ER-beta levels. Our results demonstrate a novel mechanistic role for HOXA7 in modulating breast cancer cell proliferation via regulation of ERα expression. This finding contributes to our understanding of the role HOXA7 plays in regulating the proliferation of ER-positive cancer cells.

  4. Activation of transmembrane bile acid receptor TGR5 stimulates insulin secretion in pancreatic {beta} cells

    SciTech Connect

    Kumar, Divya P.; Rajagopal, Senthilkumar; Mahavadi, Sunila; Mirshahi, Faridoddin; Grider, John R.; Murthy, Karnam S.; Sanyal, Arun J.

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer G protein coupled receptor TGR5 is expressed in mouse and human islets. Black-Right-Pointing-Pointer TGR5 is coupled to activation of Gs and Ca{sup 2+} release via cAMP/Epac/PLC-{epsilon} pathway. Black-Right-Pointing-Pointer Activation of TGR5 by bile salts and selective ligands causes insulin secretion. Black-Right-Pointing-Pointer TGR5 could be a potential therapeutic target to treat diabetes. -- Abstract: Bile acids act as signaling molecules and stimulate the G protein coupled receptor, TGR5, in addition to nuclear farnesoid X receptor to regulate lipid, glucose and energy metabolism. Bile acid induced activation of TGR5 in the enteroendocrine cells promotes glucagon like peptide-1 (GLP-1) release, which has insulinotropic effect in the pancreatic {beta} cells. In the present study, we have identified the expression of TGR5 in pancreatic {beta} cell line MIN6 and also in mouse and human pancreatic islets. TGR5 selective ligands, oleanolic acid (OA) and INT-777 selectively activated G{alpha}{sub s} and caused an increase in intracellular cAMP and Ca{sup 2+}. OA and INT-777 also increased phosphoinositide (PI) hydrolysis and the increase was blocked by NF449 (a selective G{alpha}{sub s} inhibitor) or (U73122) (PI hydrolysis inhibitor). OA, INT-777 and lithocholic acid increased insulin release in MIN6 and human islets and the increase was inhibited by treatment with NF449, (U73122) or BAPTA-AM (chelator of calcium), but not with myristoylated PKI (PKA inhibitor), suggesting that the release is dependent on G{sub s}/cAMP/Ca{sup 2+} pathway. 8-pCPT-2 Prime -O-Me-cAMP, a cAMP analog, which activates Epac, but not PKA also stimulated PI hydrolysis. In conclusion, our study demonstrates that the TGR5 expressed in the pancreatic {beta} cells regulates insulin secretion and highlights the importance of ongoing therapeutic strategies targeting TGR5 in the control of glucose homeostasis.

  5. Expression and function of the human granulocyte-macrophage colony-stimulating factor receptor alpha subunit.

    PubMed

    Jubinsky, P T; Laurie, A S; Nathan, D G; Yetz-Aldepe, J; Sieff, C A

    1994-12-15

    To determine the expression and function of the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor alpha chain (GMR alpha) during hematopoiesis and on leukemic cells, monoclonal antibodies were raised by immunizing mice with cells expressing high levels of human GMR alpha. A pool of five antibodies isolated from three different mice was used to characterize GMR alpha. This antibody pool (anti-GMR alpha) immunoprecipitated a protein with the expected molecular weight of GMR alpha from COS cells transiently transfected with the GMR alpha gene. In factor-dependent cells, GMR alpha existed as a phosphoprotein. However, its phosphorylation was not stimulated by the presence of GM-CSF. Anti-GMR alpha inhibited the GM-CSF-dependent growth of cell lines and normal bone marrow cells and inhibited the binding of iodinated GM-CSF to its receptor. Cell surface expression of GMR alpha was examined using anti-GMR alpha and flow cytometry. GMR alpha was readily detectable on both blood monocytes and neutrophils. In adherence-depleted normal bone marrow, two separate populations expressed GMR alpha. The most positive cells were predominantly macrophages, whereas the cells that expressed less GMR alpha were largely myelocytes and metamyelocytes. A small population of lin-CD34+ or CD34+CD38- cells also expressed GMR alpha, but they were not capable of significant growth in colony-forming assays. In contrast, the majority of lin-CD34+ and CD34+CD38- cells were GMR alpha-, yet they produced large numbers of myeloid and erythroid colonies in the same assay. Malignant cells from patients with leukemia were also tested for GMR alpha expression. All of the myeloid leukemias and only rare lymphoid leukemias surveyed tested positive for GMR alpha. These results show that anti-GMR alpha is useful for the functional characterization of the GMR alpha and for the detection of myeloid leukemia and that GMR alpha is expressed on certain lineages throughout hematopoietic

  6. Transcranial random noise stimulation-induced plasticity is NMDA-receptor independent but sodium-channel blocker and benzodiazepines sensitive

    PubMed Central

    Chaieb, Leila; Antal, Andrea; Paulus, Walter

    2015-01-01

    Background: Application of transcranial random noise stimulation (tRNS) between 0.1 and 640 Hz of the primary motor cortex (M1) for 10 min induces a persistent excitability increase lasting for at least 60 min. However, the mechanism of tRNS-induced cortical excitability alterations is not yet fully understood. Objective: The main aim of this study was to get first efficacy data with regard to the possible neuronal effect of tRNS. Methods: Single-pulse transcranial magnetic stimulation (TMS) was used to measure levels of cortical excitability before and after combined application of tRNS at an intensity of 1 mA for 10 min stimulation duration and a pharmacological agent (or sham) on eight healthy male participants. Results: The sodium channel blocker carbamazepine showed a tendency toward inhibiting MEPs 5–60 min poststimulation. The GABAA agonist lorazepam suppressed tRNS-induced cortical excitability increases at 0–20 and 60 min time points. The partial NMDA receptor agonist D-cycloserine, the NMDA receptor antagonist dextromethorphan and the D2/D3 receptor agonist ropinirole had no significant effects on the excitability increases seen with tRNS. Conclusions: In contrast to transcranial direct current stimulation (tDCS), aftereffects of tRNS are seem to be not NMDA receptor dependent and can be suppressed by benzodiazepines suggesting that tDCS and tRNS depend upon different mechanisms. PMID:25914617

  7. The Synthesis and Evaluation of Dihydroquinazolin-4-ones and Quinazolin-4-ones as Thyroid Stimulating Hormone Receptor Agonists

    PubMed Central

    Englund, Erika E.; Neumann, Susanne; Eliseeva, Elena; McCoy, Joshua G.; Titus, Steven; Zheng, Wei; Southall, Noel; Shin, Paul; Leister, William; Thomas, Craig J.; Inglese, James; Austin, Christopher P.; Gershengorn, Marvin C.

    2011-01-01

    We herein describe the rapid synthesis of a diverse set of dihydroquinazolin-4-ones and quinazolin-4-ones, their biological evaluation as thyroid stimulating hormone receptor (TSHR) agonists, and SAR analysis. Among the compounds screened, 8b was 60-fold more potent than the hit compound 1a, which was identified from a high throughput screen of over 73,000 compounds. PMID:22408719

  8. Phorbol esters inhibit alpha/sub 1/-adrenergic receptor stimulated phosphoinositide hydrolysis and contraction in rat aorta

    SciTech Connect

    Not Available

    1986-03-01

    The mechanisms of pharmacomechanical coupling in vascular tissue are at the present time unclear. The authors and others have proposed that receptor-induced activation of phosphoinositide (PI) hydrolysis may be involved. To investigate this possibility they studied the actions of two biologically active phorbol esters: phorbol dibutyrate (PDB) and phorbol myristate diacetate (PMA) on receptor-stimulated PI hydrolysis in rat aortic rings. They found both PDB (IC/sub 5//sup 0/ approx. 5nM) and PMA (IC/sub 50/ approx. 30 nM) but not 4-..cap alpha..-phorbol (IC32%/sub 0/ > 10,000 nM) inhibited norepinephrine-stimulated PI hydrolysis. In the presence of the calcium channel antagonist nitrendipine, PDB potently inhibited both the phasic and tonic components of norepinephrine-induced vascular contraction. In the presence of 10/sup -7/M nitrendipine, PDB had an IC/sub 50/ for contraction of approximately 10nM. The results thus suggest a functional coupling between ..cap alpha../sub 1/-adrenergic receptor-stimulated PI hydrolysis and vascular contraction. The findings further imply a mode of feed-back regulation in vascular tissue involving phorbol ester and receptor-stimulated PI hydrolysis.

  9. Stimulation of serotonin-1A receptors in mammals to alleviate motion sickness and emesis induced by chemical agents

    NASA Technical Reports Server (NTRS)

    Lucot, James B. (Inventor); Crampton, George H. (Inventor)

    1990-01-01

    A method for the alleviation of both motion sickness and chemically-induced emesis is provided which includes the administration of a nontoxic, therapeutically effective amount of a composition which stimulates serotonin-1A receptors in a mammal in need of such treatment. The preferred compounds for use are buspirone and 8-hydroxy-2(di-n-propylamino)-tetralin (8-OH-DPAT).

  10. An interleukin-1 receptor antagonist blocks lipopolysaccharide-induced colony-stimulating factor production and early endotoxin tolerance.

    PubMed Central

    Henricson, B E; Neta, R; Vogel, S N

    1991-01-01

    In this report, administration of a recombinant interleukin-1 receptor antagonist protein to mice was found to inhibit induction of colony-stimulating factor as well as induction of early endotoxin tolerance by lipopolysaccharide. These findings provide direct evidence that interleukin-1 is an intermediate in these two lipopolysaccharide-induced phenomena. PMID:1825485

  11. Serum amyloid A stimulates macrophage foam cell formation via lectin-like oxidized low-density lipoprotein receptor 1 upregulation

    SciTech Connect

    Lee, Ha Young; Kim, Sang Doo; Baek, Suk-Hwan; Choi, Joon Hyuk; Cho, Kyung-Hyun; Zabel, Brian A.; Bae, Yoe-Sik

    2013-03-29

    Highlights: ► SAA induced macrophage foam cell formation. ► SAA stimulated upregulation of lectin-like oxidized low-density lipoprotein receptor 1 (LOX1). ► SAA-induced LOX1 expression and foam cell formation is mediated by JNK/NF-κB signaling. ► HDL-conjugated SAA also stimulates foam cell formation via LOX1 upregulation. ► The finding reveals a novel mechanism of action of SAA in the pathogenesis of atherosclerosis. -- Abstract: Elevated levels of serum amyloid A (SAA) is a risk factor for cardiovascular diseases, however, the role of SAA in the pathophysiology of atherosclerosis remains unclear. Here we show that SAA induced macrophage foam cell formation. SAA-stimulated foam cell formation was mediated by c-jun N-terminal kinase (JNK) signaling. Moreover, both SAA and SAA-conjugated high density lipoprotein stimulated the expression of the important scavenger receptor lectin-like oxidized low-density lipoprotein receptor 1 (LOX1) via nuclear factor-κB (NF-κB). A LOX1 antagonist carrageenan significantly blocked SAA-induced foam cell formation, indicating that SAA promotes foam cell formation via LOX1 expression. Our findings therefore suggest that SAA stimulates foam cell formation via LOX1 induction, and thus likely contributes to atherogenesis.

  12. Electric stimulation of human fibroblasts causes an increase in Ca2+ influx and the exposure of additional insulin receptors.

    PubMed

    Bourguignon, G J; Jy, W; Bourguignon, L Y

    1989-08-01

    Previously we reported that treating human fibroblasts in cell culture with high-voltage, pulsed galvanic stimulation (HVPGS) can significantly increase cellular protein and DNA synthesis (Bourguignon and Bourguignon: FASEB J., 1:398-402, 1987). In this study we have identified two of the early cellular events which occur following exposure to HVPGS: 1) an increase in Ca2+ uptake from the external medium and 2) an increase in the number of insulin receptors on the fibroblast cell surface. The increase in Ca2+ uptake begins within the first minute of electric stimulation while increased insulin binding is not detected until the second minute of stimulation. The HVPGS-induced increase in insulin binding can be inhibited by bepridil, a specific Ca2+ channel blocker, suggesting that the Ca2+ influx is required for the exposure of additional insulin receptors on the cell surface. Furthermore, we have determined that the addition of insulin to electrically stimulated cultures results in 1) an immediate, second increase in Ca2+ uptake and 2) significant increases in both protein and DNA synthesis compared to cells which were not stimulated. All three of these insulin-dependent effects are also inhibited by bepridil. Based on these results, we propose that HVPGS initially triggers the opening of voltage-sensitive calcium channels in the fibroblast plasma membrane. The increased level of intracellular Ca2+ then induces the exposure of additional insulin receptors, the fibroblasts will significantly increase both protein and DNA synthesis.

  13. Subcellular redistribution of m2 muscarinic acetylcholine receptors in striatal interneurons in vivo after acute cholinergic stimulation.

    PubMed

    Bernard, V; Laribi, O; Levey, A I; Bloch, B

    1998-12-01

    The purpose of our work was to investigate how the cholinergic environment influences the targeting and the intracellular trafficking of the muscarinic receptor m2 (m2R) in vivo. To address this question, we have used immunohistochemical approaches at light and electron microscopic levels to detect the m2R in control rats and rats treated with muscarinic receptor agonists. In control animals, m2Rs were located mostly at postsynaptic sites at the plasma membrane of perikarya and dendrites of cholinergic and NPY-somatostatin interneurons as autoreceptors and heteroreceptors, respectively. Presynaptic receptors were also detected in boutons. The m2Rs were usually detected at extrasynaptic sites, but they could be found rarely in association with symmetrical synapses, suggesting that the cholinergic transmission mediated by m2R occurs via synaptic and nonsynaptic mechanisms. The stimulation of muscarinic receptors with oxotremorine provoked a dramatic alteration of m2R compartmentalization, including endocytosis with a decrease of the density of m2R at the membrane (-63%) and an increase of those associated with endosomes (+86%) in perikarya. The very strong increase of m2R associated with multivesicular bodies (+732%) suggests that oxotremorine activated degradation. The slight increase in the Golgi apparatus (+26%) suggests that the m2R stimulation had an effect on the maturation of m2R. The substance P receptor located at the membrane of the same neurons was unaffected by oxotremorine. Our data demonstrate that cholinergic stimulation dramatically influences the subcellular distribution of m2R in striatal interneurons in vivo. These events may have key roles in controlling abundance and availability of muscarinic receptors via regulation of receptor endocytosis, degradation, and/or neosynthesis. Further, the control of muscarinic receptor trafficking may influence the activity of striatal interneurons, including neurotransmitter release and/or electric activity.

  14. Dopamine D1 receptor stimulation modulates the formation and retrieval of novel object recognition memory: Role of the prelimbic cortex

    PubMed Central

    Pezze, Marie A.; Marshall, Hayley J.; Fone, Kevin C.F.; Cassaday, Helen J.

    2015-01-01

    Previous studies have shown that dopamine D1 receptor antagonists impair novel object recognition memory but the effects of dopamine D1 receptor stimulation remain to be determined. This study investigated the effects of the selective dopamine D1 receptor agonist SKF81297 on acquisition and retrieval in the novel object recognition task in male Wistar rats. SKF81297 (0.4 and 0.8 mg/kg s.c.) given 15 min before the sampling phase impaired novel object recognition evaluated 10 min or 24 h later. The same treatments also reduced novel object recognition memory tested 24 h after the sampling phase and when given 15 min before the choice session. These data indicate that D1 receptor stimulation modulates both the encoding and retrieval of object recognition memory. Microinfusion of SKF81297 (0.025 or 0.05 μg/side) into the prelimbic sub-region of the medial prefrontal cortex (mPFC) in this case 10 min before the sampling phase also impaired novel object recognition memory, suggesting that the mPFC is one important site mediating the effects of D1 receptor stimulation on visual recognition memory. PMID:26277743

  15. Dopamine D1 receptor stimulation modulates the formation and retrieval of novel object recognition memory: Role of the prelimbic cortex.

    PubMed

    Pezze, Marie A; Marshall, Hayley J; Fone, Kevin C F; Cassaday, Helen J

    2015-11-01

    Previous studies have shown that dopamine D1 receptor antagonists impair novel object recognition memory but the effects of dopamine D1 receptor stimulation remain to be determined. This study investigated the effects of the selective dopamine D1 receptor agonist SKF81297 on acquisition and retrieval in the novel object recognition task in male Wistar rats. SKF81297 (0.4 and 0.8 mg/kg s.c.) given 15 min before the sampling phase impaired novel object recognition evaluated 10 min or 24 h later. The same treatments also reduced novel object recognition memory tested 24 h after the sampling phase and when given 15 min before the choice session. These data indicate that D1 receptor stimulation modulates both the encoding and retrieval of object recognition memory. Microinfusion of SKF81297 (0.025 or 0.05 μg/side) into the prelimbic sub-region of the medial prefrontal cortex (mPFC) in this case 10 min before the sampling phase also impaired novel object recognition memory, suggesting that the mPFC is one important site mediating the effects of D1 receptor stimulation on visual recognition memory.

  16. Further evidence for inhibition of episodic luteinizing hormone release in ovariectomized rats by stimulation of dopamine receptors.

    PubMed

    Drouva, S V; Gallo, R V

    1977-03-01

    Stimulation of dopamine receptors by apomorphine inhibits episodic LH release in ovariectomized rats. The present study was designed to examine further the role of dopamine in this process. Unrestrained, unanesthetized rats with indwelling right atrial cannulae were bled continuously (30 or 50 microliters of whole blood/5 min for 3-6 h) and whole blood samples analyzed for LH by radioimmunoassay. Animals were treated with various compounds reported to stimulate or block dopamine receptors. ET 495, a long acting dopamine receptor stimulating agent, caused a marked inhibition of episodic LH release (2 1/2-4 h). Control injections of distilled water had no effect. d-Butaclamol, a blocker of dopamine receptors, did not itself alter episodic LH release but prevented the inhibitory effects seen following apomorphine or ET 495. I-butaclamol, a biologically inactive form of butaclamol, had no effect. Measurement of plasma corticosterone levels in these same animals indicated increased values following apomorphine or ET 495 alone (when LH release was inhibited), as well as after apomorphine or ET 495 administration to d-butaclamol-pretreated rats (when LH levels did not change). These data support our previous hypothesis that in ovariectomized adult rats, activation of dopamine receptors is capable of inhibiting episodic LH release, but that dopamine may not play an inhibitory role under normal physiological conditions in the modulation of LH secretion. In addition, the inhibitory action of apomorphine and ET 495 does not appear to be exerted via a stress-induced release of adrenal corticosterone.

  17. Serum amyloid A stimulates macrophage foam cell formation via lectin-like oxidized low-density lipoprotein receptor 1 upregulation

    PubMed Central

    Lee, Ha Young; Kim, Sang Doo; Baek, Suk-Hwan; Choi, Joon Hyuk; Cho, Kyung-Hyun; Zabel, Brian A.; Bae, Yoe-Sik

    2013-01-01

    Elevated levels of serum amyloid A (SAA) is a risk factor for cardiovascular diseases, however, the role of SAA in the pathophysiology of atherosclerosis remains unclear. Here we show that SAA induced macrophage foam cell formation. SAA-stimulated foam cell formation was mediated by c-jun N-terminal kinase (JNK) signaling. Moreover, both SAA and SAA-conjugated high density lipoprotein stimulated the expression of the important scavenger receptor lectin-like oxidized low-density lipoprotein receptor 1 (LOX1) via nuclear factor-κB (NF-κB). A LOX1 antagonist carrageenan significantly blocked SAA-induced foam cell formation, indicating that SAA promotes foam cell formation via LOX1 expression. Our findings therefore suggest that SAA stimulates foam cell formation via LOX1 induction, and thus likely contributes to atherogenesis. PMID:23454129

  18. Endothelin receptor B protects granulocyte macrophage colony-stimulating factor mRNA from degradation.

    PubMed

    Jungck, David; Knobloch, Jürgen; Körber, Sandra; Lin, Yingfeng; Konradi, Jürgen; Yanik, Sarah; Stoelben, Erich; Koch, Andrea

    2015-06-01

    Evidence is lacking on the differential effects of the two therapeutic concepts of endothelin receptor antagonists (ERAs): the blockade of only the endothelin receptor A (ETAR; selective antagonism) versus both ETAR and endothelin receptor B (ETBR; dual blockade). Ambrisentan, a selective ERA, and bosentan, a dual blocker, are both available for therapy. We hypothesized that there are differences in the potential of ERAs to ameliorate inflammatory processes in human airway smooth muscle cells (HASMCs) and aimed to unravel underlying mechanisms. We used HASMC culture, enzyme-linked immunosorbent assay, and quantitative reverse-transcription polymerase chain reaction. Tumor necrosis factor α (TNFα) induced transcription and expression of chemokine (C-X-C motif) ligand 2 (CXCL2), chemokine (C-X-C motif) ligand 3 (CXCL3), granulocyte macrophage colony-stimulating factor (GM-CSF), and matrix metalloproteinase 12 (MMP12) in HASMCs. In concentration-response experiments, bosentan led to a significantly greater reduction of GM-CSF and MMP12 protein release than ambrisentan, whereas there was no significant difference in their effect on GM-CSF and MMP12 mRNA. Both ERAs reduced CXCL3 protein and mRNA equally but had no effect on CXCL2. Blocking mitogen-activated protein kinases revealed that both ETAR and ETBR signal through p38 mitogen-activated protein kinase, but ETBR also signals through extracellular signal-regulated kinase (ERK) 1/2 to induce GM-CSF expression. In the presence of the transcription inhibitor actinomycin D, bosentan, but not ambrisentan, reduced GM-CSF but not MMP12 or CXCL3 mRNA. In conclusion, blockade of each endothelin receptor subtype reduces GM-CSF transcription, but blocking ETBR additionally protects GM-CSF mRNA from degradation via ERK-1/2. Accordingly, blocking both ETAR and ETBR leads to a stronger reduction of TNFα-induced GM-CSF protein expression. This mechanism might be specific to GM-CSF. Our data stress the anti-inflammatory potential

  19. Beta-Adrenergic Receptors and Isoproterenol-stimulated Potassium Transport in Erythrocytes from Normal and Hypothyroid Turkeys

    PubMed Central

    Furukawa, Haruyasu; Loeb, John N.; Bilezikian, John P.

    1980-01-01

    We have previously reported that in hypothyroid turkeys the number of beta-adrenergic receptors in intact erythrocytes is reduced by ∼50% without any changes in the affinity of the receptor for the agonist, isoproterenol. In view of the physiological action of the catecholamines to stimulate bidirectional ion fluxes in these cells, we have now examined the possibility that the decrease in beta receptor number might be associated with concomitant changes in catecholamine-dependent potassium ion transport. Hypothyroid turkey erythrocytes display decreased sensitivity to isoproterenol-stimulated potassium influx. Half-maximal stimulation of potassium influx occurs at 9.2±1.7 nM in hypothyroid cells as opposed to only 3.8±0.4 nM in normal cells (P < 0.005). A maximal stimulatory concentration of isoproterenol (100 nM) leads to the same increment in ion flux in erythrocytes from hypothyroid and normal turkeys. Analysis of the quantitative relationship between isoproterenol concentration, receptor occupancy, and associated effects upon potassium influx shows that at low levels of isoproterenol, where occupancy is linear with agonist concentration, occupation of a given number of beta receptors leads to a stimulation of potassium transport that is identical in erythrocytes from normal and hypothyroid turkeys. Thus, decreased sensitivity to catecholamine-stimulated potassium transport in hypothyroidism can be attributed to the decrease in receptor number and the resulting two- to threefold higher isoproterenol concentration required for occupancy of the same number of beta receptors. Once a single receptor is occupied, however, the more distal components of the sequence of events mediating the physiological response to beta-adrenergic agonists in the hypothyroid cell function as they do under normal circumstances. It would appear, therefore, that the decrease in sensitivity to isoproterenol-dependent ion flux in the hypothyroid turkey erythrocyte can be accounted for

  20. Enhanced response to mouse thyroid-stimulating hormone (TSH) receptor immunization in TSH receptor-knockout mice.

    PubMed

    Nakahara, Mami; Mitsutake, Norisato; Sakamoto, Hikaru; Chen, Chun-Rong; Rapoport, Basil; McLachlan, Sandra M; Nagayama, Yuji

    2010-08-01

    Graves-like hyperthyroidism is induced in BALB/c mice by immunization with adenovirus expressing the human TSH receptor (TSHR) A-subunit (amino acids 1-289). However, because of nonidentity between the human and mouse TSHR ( approximately 87% amino acid homology), we compared the responses of mice immunized with adenoviruses expressing either the mouse or the human TSHR A-subunit. Wild-type (wt) BALB/c mice immunized with the mouse A-subunit developed neither TSHR antibodies (measured by flow cytometry) nor thyroid lymphocytic infiltration. However, wt C57BL/6 mice developed sparse intrathyroidal lymphocyte infiltration without antibody production. Depletion of naturally occurring regulatory CD4(+)CD25(+) T cells had little effect. These results indicate the inability to break tolerance to the mouse TSHR in wt mice. In contrast, TSHR knockout (KO) BALB/c mice generated mouse TSHR antibodies in response to mouse A-subunit immunization and augmented human TSHR antibody response to human A-subunit immunization. Thyroid-stimulating antibody titers measured in a functional bioassay were comparable in human A-subunit immunized wt mice and in TSHR KO mice immunized with either the mouse or human A-subunit. In conclusion, immune response to the mouse TSHR is readily induced in TSHR KO but not in wt mice. Only in the former does immunization with adenovirus expressing the mouse A-subunit generate antibodies capable of activating the mouse TSHR. TSHR KO mice are, therefore, of value for future studies dissecting the autoimmune response to the mouse TSHR.

  1. Influence of intramuscular heat stimulation on modulation of nociception: complex role of central opioid receptors in descending facilitation and inhibition.

    PubMed

    You, Hao-Jun; Lei, Jing; Ye, Gang; Fan, Xiao-Li; Li, Qiang

    2014-10-01

    It has been reported that the threshold to activate 'silent' or inactive descending facilitation of nociception is lower than that of descending inhibition. Thus, the development of pain therapy to effectively drive descending inhibition alone, without the confounding influences of facilitation is a challenge. To address this issue we investigated the effects of intramuscular stimulation with a heating-needle on spinal nociception, assessed by measuring nociceptive paw withdrawal reflex in rats. Additionally, involvement of the thalamic 'nociceptive discriminators' (thalamic mediodorsal (MD) and ventromedial (VM) nuclei), and opioid-mediated mechanisms were further explored. Descending facilitation and inhibition were elicited by 46°C noxious heating-needle stimulation, and were regulated by thalamic MD and VM nuclei, respectively. In contrast, innocuous heating-needle stimulation at a temperature of 43°C elicited descending inhibition modulated by the thalamic VM nucleus alone. Microinjection of μ/δ/κ-opioid receptor antagonists β-funaltrexamine hydrochloride/naltrindole/nor-binaltorphimine, into the VM nucleus attenuated the 46°C intramuscular heating-needle stimulation-evoked descending inhibition, whereas treatment of the MD nucleus with β-funaltrexamine hydrochloride significantly decreased the descending facilitation. By contrast, descending inhibition evoked by 43°C heating-needle stimulation was only depressed by naltrindole, as opposed to μ- and κ-opioid receptor antagonists, which failed to influence descending inhibition. The present study reveals distinct roles of μ-opioid receptors in the function of thalamic MD and VM nuclei,which exert facilitatory and inhibitory actions on nociception. Furthermore, innocuous, but not noxious, intramuscular heating-needle stimulation targeting δ-opioid receptors is suggested to be a promising avenue for the effective inhibition of pain.

  2. Stimulants of Toll-like receptor (TLR)-2 and TLR-4 are abundant in certain minimally-processed vegetables.

    PubMed

    Erridge, Clett

    2011-06-01

    Stimulants of the innate immune receptors Toll-like receptor (TLR)-2 and TLR4 have been shown to promote insulin resistance and atherosclerosis in animal models of these diseases. As minimally processed vegetables (MPV) can contain a relatively large bacterial load compared to other foodstuffs, we aimed to quantify the abundance of stimulants of TLR2 and TLR4 in MPV using a transfection-based bioassay calibrated with Escherichia coli LPS and the synthetic lipopeptide Pam(3)CSK(4). Of 5 classes of MPV and 3 classes of related vegetable products considered to be likely to contain a high microbial load, diced onion and bean sprouts contained the highest levels of stimulants of TLR2 (up to 18.5 μg Pam(3)CSK(4)-equivalents per g) and TLR4 (up to 11.4 μg LPS-equivalents per g). By contrast, the majority of fresh whole vegetables examined reproducibly contained minimal or undetectable levels of TLR2- or TLR4-stimulants. The accumulation of TLR-stimulants in MPVs correlated well with growth of enterobacterial spoilage organisms. In conclusion, the modern trend towards eating minimally processed vegetables rather than whole foods is likely to be associated with increased oral exposure to stimulants of TLR2 and TLR4. Copyright © 2011 Elsevier Ltd. All rights reserved.

  3. Effects of the cannabinoid receptor agonist CP 55,940 and the cannabinoid receptor antagonist SR 141716 on intracranial self-stimulation in Lewis rats.

    PubMed

    Arnold, J C; Hunt, G E; McGregor, I S

    2001-11-21

    Lewis rats were trained to self-stimulate the medial forebrain bundle (MFB) using a rate-frequency paradigm. They were then tested for the effects of the cannabinoid receptor agonist CP 55,940, the selective cannabinoid receptor antagonist SR 141716 and the dopamine D1 receptor antagonist SCH 23390. CP 55,940 (0, 10, 25 and 50 microg/kg i.p.) had no effect on MFB self-stimulation behaviour as assessed by the M50, the stimulation frequency at which half-maximal response rates were obtained. With SR 141716, only a very high dose (20 mg/kg i.p.) caused a significant inhibition of the rewarding efficacy of the stimulation. This was seen as an increase in the M50. All other doses of SR 141716 (0, 1, 3, 10 mg/kg i.p.) were ineffective in modulating the M50. By comparison, a relatively low dose (0.06 mg/kg i.p.) of SCH 23390 caused a large increase in M50. These results indicate a relatively modest influence, if any at all, of exogenous or endogenous cannabinoids on reward-relevant neurotransmission.

  4. Stimulation of Hepatic Apolipoprotein A-I Production by Novel Thieno-Triazolodiazepines: Roles of the Classical Benzodiazepine Receptor, PAF Receptor, and Bromodomain Binding

    PubMed Central

    Kempen, Herman J; Bellus, Daniel; Fedorov, Oleg; Nicklisch, Silke; Filippakopoulos, Panagis; Picaud, Sarah; Knapp, Stefan

    2013-01-01

    Expression and secretion of apolipoprotein A-I (apoA-I) by cultured liver cells can be markedly stimulated by triazolodiazepines (TZDs). It has been shown previously that the thieno-TZD Ro 11-1464 increases plasma levels of apoA-I and in vivomacrophage reverse cholesterol transport in mice. However, these effects were only seen at high doses, at which the compound could act on central benzodiazepine (BZD) receptors or platelet activating factor (PAF) receptors, interfering with its potential utility. In this work, we describe 2 new thieno-TZDs MDCO-3770 and MDCO-3783, both derived from Ro 11-1464. These compounds display the same high efficacy on apoA-I production, metabolic stability, and lack of cytotoxicity in cultured hepatocytes as Ro 11-1464, but they do not bind to the central BZD receptor and PAF receptor. The quinazoline RVX-208 was less efficacious in stimulating apoA-I production and displayed signs of cytotoxicity. Certain TZDs stimulating apoA-I production are now known to be inhibitors of bromodomain (BRD) extra-terminal (BET) proteins BRDT, BRD2, BRD3, and BRD4, and this inhibition was inferred as a main molecular mechanism for their effect on apoA-I expression. We show here that the thieno-TZD (+)-JQ1, a potent BET inhibitor, strongly stimulated apoA-I production in Hep-G2 cells, but that its enantiomer (−)-JQ1, which has no BET inhibitor activity, also showed considerable effect on apoA-I production. MDCO-3770 and MDCO-3783 also inhibited BRD3 and BRD4 in vitro, with potency somewhat below that of (+)-JQ1. We conclude that the effect of thieno-TZDs on apoA-I expression is not due to inhibition of the BZD or PAF receptors and is not completely explained by transcriptional repression by BET proteins. PMID:25278768

  5. Stimulation of Hepatic Apolipoprotein A-I Production by Novel Thieno-Triazolodiazepines: Roles of the Classical Benzodiazepine Receptor, PAF Receptor, and Bromodomain Binding.

    PubMed

    Kempen, Herman J; Bellus, Daniel; Fedorov, Oleg; Nicklisch, Silke; Filippakopoulos, Panagis; Picaud, Sarah; Knapp, Stefan

    2013-01-01

    Expression and secretion of apolipoprotein A-I (apoA-I) by cultured liver cells can be markedly stimulated by triazolodiazepines (TZDs). It has been shown previously that the thieno-TZD Ro 11-1464 increases plasma levels of apoA-I and in vivomacrophage reverse cholesterol transport in mice. However, these effects were only seen at high doses, at which the compound could act on central benzodiazepine (BZD) receptors or platelet activating factor (PAF) receptors, interfering with its potential utility. In this work, we describe 2 new thieno-TZDs MDCO-3770 and MDCO-3783, both derived from Ro 11-1464. These compounds display the same high efficacy on apoA-I production, metabolic stability, and lack of cytotoxicity in cultured hepatocytes as Ro 11-1464, but they do not bind to the central BZD receptor and PAF receptor. The quinazoline RVX-208 was less efficacious in stimulating apoA-I production and displayed signs of cytotoxicity. Certain TZDs stimulating apoA-I production are now known to be inhibitors of bromodomain (BRD) extra-terminal (BET) proteins BRDT, BRD2, BRD3, and BRD4, and this inhibition was inferred as a main molecular mechanism for their effect on apoA-I expression. We show here that the thieno-TZD (+)-JQ1, a potent BET inhibitor, strongly stimulated apoA-I production in Hep-G2 cells, but that its enantiomer (-)-JQ1, which has no BET inhibitor activity, also showed considerable effect on apoA-I production. MDCO-3770 and MDCO-3783 also inhibited BRD3 and BRD4 in vitro, with potency somewhat below that of (+)-JQ1. We conclude that the effect of thieno-TZDs on apoA-I expression is not due to inhibition of the BZD or PAF receptors and is not completely explained by transcriptional repression by BET proteins.

  6. An Antagonistic Vascular Endothelial Growth Factor (VEGF) Variant Inhibits VEGF-Stimulated Receptor Autophosphorylation and Proliferation of Human Endothelial Cells

    NASA Astrophysics Data System (ADS)

    Siemeister, Gerhard; Schirner, Michael; Reusch, Petra; Barleon, Bernhard; Marme, Dieter; Martiny-Baron, Georg

    1998-04-01

    Vascular endothelial growth factor (VEGF) is a potent mitogen with a unique specificity for endothelial cells and a key mediator of aberrant endothelial cell proliferation and vascular permeability in a variety of human pathological situations, such as tumor angiogenesis, diabetic retinopathy, rheumatoid arthritis, or psoriasis. VEGF is a symmetric homodimeric molecule with two receptor binding interfaces lying on each pole of the molecule. Herein we report on the construction and recombinant expression of an asymmetric heterodimeric VEGF variant with an intact receptor binding interface at one pole and a mutant receptor binding interface at the second pole of the dimer. This VEGF variant binds to VEGF receptors but fails to induce receptor activation. In competition experiments, the heterodimeric VEGF variant antagonizes VEGF-stimulated receptor autophosphorylation and proliferation of endothelial cells. A 15-fold excess of the heterodimer was sufficient to inhibit VEGF-stimulated endothelial cell proliferation by 50%, and a 100-fold excess resulted in an almost complete inhibition. By using a rational approach that is based on the structure of VEGF, we have shown the feasibility to construct a VEGF variant that acts as an VEGF antagonist.

  7. Co-expression of the Follicle Stimulating Hormone Receptor and Stem Cell Markers: A Novel Approach to Target Ovarian Cancer Stem Cells

    DTIC Science & Technology

    2012-09-01

    Award Number: 11 1 0623 TITLE: Co-expression of the Follicle Stimulating Hormone Receptor and Stem...Annual 3. DATES COVERED 1 Sep 2011 – 31 Aug 2012 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Coexpression of the Follicle Stimulating Hormone ...Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT The purpose of this project is to determine whether the Follicle-stimulating Hormone Receptor (FSHR) is co

  8. Roflumilast improves corticosteroid resistance COPD bronchial epithelial cells stimulated with toll like receptor 3 agonist.

    PubMed

    Milara, Javier; Morell, Anselm; Ballester, Bea; Sanz, Celia; Freire, Jose; Qian, Xiaozhong; Alonso-Garcia, Maggie; Morcillo, Esteban; Cortijo, Julio

    2015-02-05

    Chronic obstructive pulmonary disease (COPD) is characterised by chronic pulmonary inflammation punctuated by periods of viral exacerbations. Recent evidence suggests that the combination of roflumilast with corticosteroids may improve the compromised anti-inflammatory properties of corticosteroids in COPD. We analyzed differential and combination anti-inflammatory effects of dexamethasone and roflumilast N-oxide in human bronchial epithelial cells (HBECs) stimulated with viral toll like receptor (TLR) agonists. Lung tissue and HBECs were isolated from healthy (n = 15), smokers (n = 12) and smokers with COPD (15). TLR3 expression was measured in lung tissue and in HBECs. IL-8 secretion was measured in cell cultures after TLR3 stimulation with poly I:C 10 μg/mL. We found that TLR3 expression was increased by 1.95 fold (protein) and 2.5 fold (mRNA) in lung tissues from smokers with COPD and inversely correlated with lung function. The TLR3 agonist poly I:C 10 μg/mL increased the IL-8 release in HBECs that was poorly inhibited by dexamethasone in smokers (24.5%) and smokers with COPD (21.6%). In contrast, roflumilast showed similar inhibitory effects on IL-8 release in healthy (58.8%), smokers (56.6%) and smokers with COPD (50.5%). The combination of roflumilast N-oxide and dexamethasone showed additive inhibitory effects. Mechanistically, roflumilast N-oxide when combined with dexamethasone increased the expression of MKP1, and enhanced the inhibitory effects on phospho-p38, AP1 and NFκB activities which may explain the additive anti-inflammatory effects. Altogether, our data provide in vitro evidence for a possible clinical utility to add roflumilast on top of inhaled corticosteroid in COPD.

  9. Stimulating TSH receptor autoantibodies immunoassay: analytical evaluation and clinical performance in Graves' disease.

    PubMed

    Autilio, C; Morelli, R; Locantore, P; Pontecorvi, A; Zuppi, C; Carrozza, C

    2017-01-01

    Background Thyroid-stimulating hormone (TSH) receptor (TSHR) autoantibodies (TRAbs) are a heterogeneous group of antibodies (Abs) with different functionalities. Among all TRAbs, only the stimulating ones (S-TRAbs) are considered as the pathogenetic marker of Graves' disease (GD). To date, the methods available for TRAbs testing are based on immunoassays (IMAs) which detect total serum TRAbs or bioassays which are not suitable in clinical practice, even though they discern Abs functionality. The aim of our work was to evaluate the analytical and clinical performance of a very recent IMA (Immulite TSI method), supposed to test only the serum concentration of S-TRAbs, in comparison with a current method for total TRAbs (Roche/Elecsys IMA). Methods We evaluated serum samples of 145 subjects: 46 with untreated (GD), 36 with chronic autoimmune thyroiditis, 3 with atrophic thyroiditis, 10 with multinodular non-toxic goiter and 50 healthy subjects. Results The method showed an optimal analytical sensitivity and high precision levels (LoB: 0.04 UI/L, LoD:0.07 UI/L, LoQ:0.14 UI/L, intra-assay CV: 4.2-5.9%, inter-assay: 4.5-7.2%). By receiver operating characteristics curve analysis, we obtained a value of 0.57 (sensitivity: 98.0%, specificity: 99.9%) as the best cut-off to distinguish GD, apart from four cases. Passing Bablok regression and Bland Altman analysis pointed out a good correlation and agreement with Roche method (R(2 )= 0.98, slope = 1.03, bias = -2.70). Conclusions The new method presents very promising analytical characteristics and could be adopted in clinical practice for GD diagnosis. Moreover, the test allows to accurately detect very low values of analyte with a further clinical utility in detecting earlier possible relapses.

  10. Systemic Toll-Like Receptor Stimulation Suppresses Experimental Allergic Asthma and Autoimmune Diabetes in NOD Mice

    PubMed Central

    Pham Van, Linh; Bardel, Emilie; Gomez Alcala, Alejandro; Jeannin, Pascale; Akira, Shizuo; Bach, Jean-François; Thieblemont, Nathalie

    2010-01-01

    Background Infections may be associated with exacerbation of allergic and autoimmune diseases. Paradoxically, epidemiological and experimental data have shown that some microorganisms can also prevent these pathologies. This observation is at the origin of the hygiene hypothesis according to which the decline of infections in western countries is at the origin of the increased incidence of both Th1-mediated autoimmune diseases and Th2-mediated allergic diseases over the last decades. We have tested whether Toll-like receptor (TLR) stimulation can recapitulate the protective effect of infectious agents on allergy and autoimmunity. Methods and Findings Here, we performed a systematic study of the disease-modifying effects of a set of natural or synthetic TLR agonists using two experimental models, ovalbumin (OVA)-induced asthma and spontaneous autoimmune diabetes, presenting the same genetic background of the non obese diabetic mouse (NOD) that is highly susceptible to both pathologies. In the same models, we also investigated the effect of probiotics. Additionally, we examined the effect of the genetic invalidation of MyD88 on the development of allergic asthma and spontaneous diabetes. We demonstrate that multiple TLR agonists prevent from both allergy and autoimmunity when administered parenterally. Probiotics which stimulate TLRs also protect from these two diseases. The physiological relevance of these findings is further suggested by the major acceleration of OVA-induced asthma in MyD88 invalidated mice. Our results strongly indicate that the TLR-mediated effects involve immunoregulatory cytokines such as interleukin (IL)-10 and transforming growth factor (TGF)-β and different subsets of regulatory T cells, notably CD4+CD25+FoxP3+ T cells for TLR4 agonists and NKT cells for TLR3 agonists. Conclusions/Significance These observations demonstrate that systemic administration of TLR ligands can suppress both allergic and autoimmune responses. They provide a

  11. Dehydroepiandrosterone administration improves memory deficits following transient brain ischemia through sigma-1 receptor stimulation.

    PubMed

    Yabuki, Yasushi; Shinoda, Yasuharu; Izumi, Hisanao; Ikuno, Tatuya; Shioda, Norifumi; Fukunaga, Kohji

    2015-10-05

    Dehydroepiandrosterone (DHEA) is the most abundant neurosteroid synthesized de novo in the central nervous system. Oral DHEA administration elicits neuroprotection and cognitive improvement, but mechanisms underlying these functions in cerebral ischemia have remained unclear. Since DHEA is the endogenous ligand for the sigma-1 receptor (σ1R), we determined whether oral DHEA administration prevents neuronal cell death and improves cognition via σ1R stimulation in brain ischemia using a 20-min bilateral common carotid artery occlusion (BCCAO) mouse model. Twenty-four hours after BCCAO ischemia, mice were administered DHEA (15 or 30mg/kg p.o.) daily for 11 consecutive days. Memory deficits following brain ischemia were improved by DHEA administration dose-dependently. Accordingly, DHEA administration significantly prevented neuronal cell death in the hippocampal CA1 region in BCCAO mice. Interestingly, DHEA administration rescued decreases in Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) autophosphorylation and phosphorylation of extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) in the CA1 region. Moreover, DHEA administration significantly ameliorated decreases in adenosine 5'-triphosphate (ATP) levels and decreased σ1R expression levels in CA1 following BCCAO ischemia. Finally, co-treatment of mice with the σ1R antagonist NE-100 (1mg/kg, p.o.) blocked DHEA effects on memory improvement and neuroprotection in ischemic mice. Taken together, DHEA prevents neuronal cell death and activates CaMKII via σ1R stimulation, thereby improving cognitive deficits following brain ischemia. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Cellular taurine release triggered by stimulation of the Fas(CD95) receptor in Jurkat lymphocytes.

    PubMed

    Lang, F; Madlung, J; Uhlemann, A C; Risler, T; Gulbins, E

    1998-08-01

    One of the hallmarks of apoptosis is cell shrinkage which appears to be important for cell death. The mechanisms mediating cell volume decrease have, however, not been addressed. Mechanisms employed by swollen cells to decrease their cell volume include activation of ion transport pathways, such as ion channels and KCl cotransport, and release of cellular osmolytes, such as taurine, sorbitol, betaine and inositol. The present study has been performed to test for release of taurine. To this end Jurkat human T-lymphocytes were loaded with [3H]taurine and apoptotic cell death induced by triggering the Fas(CD95) receptor with monoclonal crosslinking antibody. Triggering the Fas(CD95) receptor led to a release of 60+/-5% of cellular taurine within 90 min. The release did not occur prior to 45 min. The release coincided with cell shrinkage as evidenced from forward scatter in FACS analysis and preceeded DNA fragmentation according to propidium iodide staining. The delay of taurine release was not influenced by exchange of medium and thus was not due to extracellular accumulation of a stimulator. The Fas(CD95)-induced taurine release, cell shrinkage and DNA fragmentation were blunted by lowering of ambient temperature to 23 degreesC. Following pretreatment of cells with Fas(CD95) antibody at 23 degreesC rewarming led to rapid taurine release, cell shrinkage and DNA fragmentation, indicating that the temperature-sensitive step is distal to the mechanisms accounting for the delay. Osmotic cell swelling led to an immediate release of taurine. In conclusion, Fas(CD95) triggering leads to delayed taurine release through a temperature-sensitive mechanism.

  13. Expression of transferrin receptors on mitogen-stimulated human peripheral blood lymphocytes: relation to cellular activation and related metabolic events.

    PubMed Central

    Galbraith, R M; Galbraith, G M

    1981-01-01

    Mitogen-activated normal human peripheral blood lymphocytes bind transferrin to specific membrane receptors. In this study, lymphocytes stimulated with phytohaemagglutinin for 0-66 hr were examined to determine the relation of this phenomenon to cellular activation and related metabolic events. Transferrin receptors were first detected at 20-24 hr. This event was consistently preceded by RNA and protein turnover which commenced during the first 6 hr of culture, whereas initiation of DNA synthesis was detected concurrently with the appearance of receptors or slightly later (24-30 hr). Exposure of cells to inhibitors of RNA and protein synthesis early during culture (at 0 or 24 hr) prevented the expression of transferrin receptors, but also caused generalized metabolic failure, and abrogated cellular activation. In contrast, later addition of these agents at 48 hr did not interfere significantly with the process of activation, but did suppress the terminal increase in receptor-bearing cells observed during the final 18 hr in control cultures lacking inhibitor. After deliberate thermal stripping of receptors from activated cells, the reappearance of membrance binding sites which normally occurred within 30 min, was also blocked by cycloheximide, puromycin and actinomycin D. However, similar inhibition of DNA which was induced by hydroxyurea had much less effect upon both the initial appearance of receptors and their reappearance after ligand-induced depletion. These results demonstrate that the appearance of transferrin receptors upon human lymphocytes is dependent upon cellular activation and requires synthesis of protein and RNA. PMID:6172372

  14. Stimulation of glutamate receptors in the ventral tegmental area is necessary for serotonin-2 receptor-induced increases in mesocortical dopamine release

    PubMed Central

    Pehek, E.A.; Hernan, A.E.

    2017-01-01

    Modulation of dopamine (DA) released by serotonin-2 (5-HT2) receptors has been implicated in the mechanism of action of antipsychotic drugs. The mesocortical DA system has been implicated particularly in the cognitive deficits observed in schizophrenia. Agonism at 5-HT2A receptors in the prefrontal cortex is associated with increases in cortical DA release. Evidence indicates that 5-HT2A receptors in the cortex regulate mesocortical DA release through stimulation of a “long-loop” feedback system from the PFC to the VTA and back. However, a causal role for VTA glutamate in the 5-HT2-induced increases in PFC DA has not been established. The present study does so by measuring 5-HT2 agonist-induced DA release in the cortex after infusions of glutamate antagonists into the VTA. Infusions of a combination of a NMDA (AP-5: 2-amino-5-phosphopentanoic acid) and an AMPA/kainate (CNQX: 6-cyano-7-nitroquinoxaline-2,3-dione) receptor antagonist into the VTA blocked the increases in cortical DA produced by administration of the 5-HT2 agonist DOI [(±)-2,5-Dimethoxy-4-iodoamphetamine] (2.5 mg/kg s.c.). These results demonstrate that stimulation of glutamate receptors in the VTA is necessary for 5-HT2 agonist-induced increases in cortical DA. PMID:25637799

  15. IL-21 Receptor Antagonist Inhibits Differentiation of B Cells toward Plasmablasts upon Alloantigen Stimulation

    PubMed Central

    de Leur, Kitty; Dor, Frank J. M. F.; Dieterich, Marjolein; van der Laan, Luc J. W.; Hendriks, Rudi W.; Baan, Carla C.

    2017-01-01

    Interaction between T follicular helper (Tfh) cells and B cells is complex and involves various pathways, including the production of IL-21 by the Tfh cells. Secretion of IL-21 results in B cell differentiation toward immunoglobulin-producing plasmablasts. In patients after kidney transplantation, the formation of alloantibodies produced by donor antigen-activated B cells are a major cause of organ failure. In this allogeneic response, the role of IL-21-producing Tfh cells that regulate B cell differentiation is unknown. Here, we tested, in an alloantigen-driven setting, whether Tfh cell help signals control B cell differentiation with its dependency on IL-21. Pre-transplantation patient PBMCs were sorted into pure CD4posCXCR5pos Tfh cells and CD19posCD27pos memory B cells and stimulated with donor antigen in the presence or absence of an IL-21 receptor (IL-21R) antagonist (αIL-21R). Donor antigen stimulation initiated expression of the activation markers inducible co-stimulator (ICOS) and programmed death 1 (PD-1) on Tfh cells and a shift toward a mixed Tfh2 and Tfh17 phenotype. The memory B cells underwent class switch recombination and differentiated toward IgM- and IgG-producing plasmablasts. In the presence of αIL-21R, a dose-dependent inhibition of STAT3 phosphorylation was measured in both T and B cells. Blockade of the IL-21R did not have an effect on PD-1 and ICOS expression on Tfh cells but significantly inhibited B cell differentiation. The proportion of plasmablasts decreased by 78% in the presence of αIL-21R. Moreover, secreted IgM and IgG2 levels were significantly lower in the presence of αIL-21R. In conclusion, our results demonstrate that IL-21 produced by alloantigen-activated Tfh cells controls B cell differentiation toward antibody producing plasmablasts. The IL-21R might, therefore, be a useful target in organ transplantation to prevent antigen-driven immune responses leading to graft failure. PMID:28373876

  16. Mineralocorticoid receptor in the NTS stimulates saline intake during fourth ventricular infusions of aldosterone

    PubMed Central

    Koneru, Bhuvaneswari; Bathina, Chandra Sekhar; Cherry, Brandon H.

    2013-01-01

    The purpose of this study was to determine whether neurons within the nucleus tractus solitarius (NTS) that express the mineralocorticoid receptor (MR) play a role in aldosterone stimulation of salt intake. Adult Wistar-Kyoto (WKY) rats received microinjections into the NTS of a short-hairpin RNA (shRNA) for the MR, to site specifically reduce levels of the MR by RNA interference (shRNA; n = 9) or scrambled RNA as a control (scRNA; n = 8). After injection of the viral construct, aldosterone-filled osmotic minipumps were implanted subcutaneously and connected to a cannula extending into the fourth ventricle to infuse aldosterone at a rate of 25 ng/h. Before and after surgeries, rats had ad libitum access to normal sodium (0.26%) rat chow and two graduated drinking bottles filled with either distilled water or 0.3 M NaCl. Before the surgeries, basal saline intake was 1.6 ± 0.6 ml in the scRNA group and 1.56 ± 0.6 ml in the shRNA group. Twenty-four days postsurgery, saline intake was elevated to a greater extent in the scRNA group (5.9 ± 1.07 ml) than in the shRNA group (2.41 ± 0.6 ml). Post mortem immunohistochemistry revealed a significant reduction in the number of NTS neurons exhibiting immunoreactivity for MR in shRNA-injected rats (23 ± 1 cells/section) versus scRNA-injected rats (33 ± 2 cells/section; P = 0.008). shRNA did not alter the level of 11-β-hydroxysteroid dehydrogenase type II (HSD2) protein in the NTS as judged by the number of HSD2 immunoreactive neurons. These results suggest that fourth ventricular infusions of aldosterone stimulate saline intake, and that this stimulation is at least in part mediated by hindbrain NTS neurons that express MR. PMID:24259463

  17. Serotonin stimulates lateral habenula via activation of the post-synaptic serotonin 2/3 receptors and transient receptor potential channels.

    PubMed

    Zuo, Wanhong; Zhang, Yong; Xie, Guiqin; Gregor, Danielle; Bekker, Alex; Ye, Jiang-Hong

    2016-02-01

    There is growing interest on the role of the lateral habenula (LHb) in depression, because it closely and bilaterally connects with the serotoninergic raphe nuclei. The LHb sends glutamate efferents to the raphe nuclei, while it receives serotoninergic afferents, and expresses a high density of serotonin (5-HT) receptors. Recent studies suggest that 5-HT receptors exist both in the presynaptic and postsynaptic sites of LHb neurons, and activation of these receptors may have different effects on the activity of LHb neurons. The current study focused on the effect of 5-HT on the postsynaptic membrane. We found that 5-HT initiated a depolarizing inward current (I((5-HTi))) and accelerated spontaneous firing in ∼80% of LHb neurons in rat brain slices. I((5-HTi)) was also induced by the 5-HT uptake blocker citalopram, indicating activity of endogenous 5-HT. I((5-HTi)) was diminished by 5-HT(2/3) receptor antagonists (ritanserin, SB-200646 or ondansetron), and activated by the selective 5-HT(2/3) agonists 1-(3-Chlorophenyl) piperazine hydrochloride or 1-(3-Chlorophenyl) biguanide hydrochloride. Furthermore, I((5-HTi)) was attenuated by 2-Aminoethyl diphenylborinate, a blocker of transient receptor potential channels, and an IP3 receptor inhibitor, indicating the involvement of transient receptor potential channels. These results demonstrate that the reciprocal connection between the LHb and the 5-HT system highlights a key role for 5-HT stimulation of LHb neurons that may be important in the pathogenesis of depression.

  18. Serotonin stimulates lateral habenula via activation of the post-synaptic serotonin 2/3 receptors and transient receptor potential channels

    PubMed Central

    Zuo, Wanhong; Zhang, Yong; Xie, Guiqin; Gregor, Danielle; Bekker, Alex; Ye, Jiang-Hong

    2015-01-01

    There is growing interest on the role of the lateral habenula (LHb) in depression, because it closely and bilaterally connects with the serotoninergic raphe nuclei. The LHb sends glutamate efferents to the raphe nuclei, while it receives serotoninergic afferents, and expresses a high density of serotonin (5-HT) receptors. Recent studies suggest that 5-HT receptors exist both in the presynaptic and postsynaptic sites of LHb neurons, and activation of these receptors may have different effects on the activity of LHb neurons. The current study focused on the effect of 5-HT on the postsynaptic membrane. We found that 5-HT initiated a depolarizing inward current (I(5-HTi)) and accelerated spontaneous firing in ~80% of LHb neurons in rat brain slices. I(5-HTi) was also induced by the 5-HT uptake blocker citalopram, indicating activity of endogenous 5-HT. I(5-HTi) was diminished by 5-HT2/3 receptor antagonists (ritanserin, SB-200646 or ondansetron), and activated by the selective 5-HT2/3 agonists 1-(3- Chlorophenyl) piperazine hydrochloride or 1-(3-Chlorophenyl) biguanide hydrochloride. Furthermore, I(5-HTi) was attenuated by 2-Aminoethyl diphenylborinate, a blocker of transient receptor potential channels, and an IP3 receptor inhibitor, indicating the involvement of transient receptor potential channels. These results demonstrate that the reciprocal connection between the LHb and the 5-HT system highlights a key role for 5-HT stimulation of LHb neurons that may be important in the pathogenesis of depression. PMID:26471419

  19. Initiation of DNA synthesis by human thrombin: relationships between receptor binding, enzymic activity, and stimulation of 86Rb+ influx.

    PubMed

    Stiernberg, J; Carney, D H; Fenton, J W; LaBelle, E F

    1984-09-01

    Stimulation of amiloride-sensitive sodium (Na+) influx and the subsequent activation of NA+, K+-ATPase by serum or growth factors have been implicated as early events leading to initiation of cell proliferation. We recently demonstrated that amiloride inhibits thrombin-initiated DNA synthesis not by inhibiting an early event occurring during the first 8 hr, but rather by inhibiting some later event 8 to 12 hr after thrombin addition. To further probe the relationship between stimulation of ion influx and initiation of cell proliferation, human alpha-thrombin was converted to gamma-thrombin, nitro-alpha-thrombin, and diisopropylphospho (DIP)-alpha-thrombin. These derivatives retain either the capacity to bind cell surface alpha-thrombin receptors or thrombin esterase activity, but they do not initiate DNA synthesis. At low concentrations of alpha-thrombin or the various thrombin derivatives, only alpha-thrombin stimulates 86Rb+ influx, suggesting a correlation between stimulation of influx and the ability of these derivatives to initiate DNA synthesis. Concentrations of a DIP-alpha-thrombin that saturate the alpha-thrombin receptors (up to 2 micrograms/ml) do not stimulate either the early or late influx of 86Rb+, indicating that DIP-alpha-thrombin binding alone is not sufficient to stimulate ion fluxes. High concentrations of either gamma-thrombin or nitro-alpha-thrombin, however, stimulate both early and late 86RB+ uptake but do not initiate DNA synthesis. These results demonstrate that events leading to both the early and late stimulation of 86Rb+ influx by themselves are not sufficient to initiate cell proliferation. Thus, initiation may require a combination of events that can be independently regulated by different transmembrane signals.

  20. Tpl2 and ERK transduce antiproliferative T cell receptor signals and inhibit transformation of chronically stimulated T cells.

    PubMed

    Tsatsanis, Christos; Vaporidi, Katerina; Zacharioudaki, Vassiliki; Androulidaki, Ariadne; Sykulev, Yuri; Margioris, Andrew N; Tsichlis, Philip N

    2008-02-26

    The protein kinase encoded by the Tpl2 protooncogene plays an obligatory role in the transduction of Toll-like receptor and death receptor signals in macrophages, B cells, mouse embryo fibroblasts, and epithelial cells in culture and promotes inflammatory responses in animals. To address its role in T cell activation, we crossed the T cell receptor (TCR) transgene 2C, which recognizes class I MHC presented peptides, into the Tpl2(-/-) genetic background. Surprisingly, the TCR2C(tg/tg)/Tpl2(-/-) mice developed T cell lymphomas with a latency of 4-6 months. The tumor cells were consistently TCR2C(+)CD8(+)CD4(-), suggesting that they were derived either from chronically stimulated mature T cells or from immature single positive (ISP) cells. Further studies showed that the population of CD8(+) ISP cells was not expanded in the thymus of TCR2C(tg/tg)/Tpl2(-/-) mice, making the latter hypothesis unlikely. Mature peripheral T cells of Tpl2(-/-) mice were defective in ERK activation and exhibited enhanced proliferation after TCR stimulation. The same cells were defective in the induction of CTLA4, a negative regulator of the T cell response, which is induced by TCR signals via ERK. These findings suggest that Tpl2 functions normally in a feedback loop that switches off the T cell response to TCR stimulation. As a result, Tpl2, a potent oncogene, functions as a tumor suppressor gene in chronically stimulated T cells.

  1. Differential sensitivity of intranuclear and systemic oxytocin release to central noradrenergic receptor stimulation during mid- and late gestation in rats.

    PubMed

    Lipschitz, David L; Crowley, William R; Bealer, Steven L

    2004-09-01

    A number of changes occur in the oxytocin (OT) system during gestation, such as increases in hypothalamic OT mRNA, increased neural lobe and systemic OT, and morphological and electrophysiological changes in OT-containing magnocellular neurons, suggestive of altered neuronal sensitivity, which may be mediated by ovarian steroids. Because central norepinephrine (NE) and histamine (HA) are potent stimulators of OT release during parturition and lactation, the present study investigated the effects of central noradrenergic and histaminergic receptor activation on systemic (NE, HA) and intranuclear (NE) OT release in pregnant rats and in ovariectomized rats treated with ovarian steroids. Plasma OT levels in late gestation were significantly higher compared with all other groups, and neither adrenergic nor histaminergic receptor blockade decreased these elevated levels. Furthermore, the alpha-adrenergic agonist phenylephrine, but not histamine, stimulated systemic OT release to a significantly greater extent in late gestation than in midpregnant, ovariectomized, or steroid-treated females. Although basal extracellular OT levels in the paraventricular nucleus, as measured with microdialysis, were unchanged during pregnancy or steroid treatment, noradrenergic receptor stimulation of intranuclear OT release was significantly elevated in midgestation females compared with all other groups. These studies indicate that sensitivity of intranuclear and systemic OT release to noradrenergic receptor activation differentially varies during the course of gestation.

  2. GABAA receptor-mediated stimulation of non-adrenergic non-cholinergic neurones in the dog ileocolonic junction.

    PubMed

    Boeckxstaens, G E; Pelckmans, P A; Rampart, M; Ruytjens, I F; Verbeuren, T J; Herman, A G; Van Maercke, Y M

    1990-10-01

    1. The inhibitory effects of gamma-aminobutyric acid (GABA), the GABAA receptor agonist homotaurine and the GABAB receptor agonist (+/-)-baclofen were investigated on circular muscle strips of the dog terminal ileum and ileocolonic junction. 2. In the presence of atropine, GABA and homotaurine induced concentration-dependent relaxations, similar to the non-adrenergic non-cholinergic (NANC)-mediated relaxations evoked by electrical stimulation or by acetylcholine. The ileocolonic junction was more sensitive to GABA and homotaurine than the ileum. (+/-)-Baclofen had no effect. Cross desensitization only occurred between GABA and homotaurine. 3. The GABAA receptor antagonist bicuculline shifted the concentration-response curves to GABA and homotaurine to the right. The maximal relaxation to GABA remained unaffected. 4. GABA-induced relaxations were not inhibited by timolol, guanethidine, domperidone, hexamethonium and desensitization to ATP, but were abolished by tetrodotoxin. 5. Bicuculline, and pretreatment with GABA or (+/-)-baclofen had no effect on the NANC-evoked relaxations to electrical stimulation and acetylcholine. 6. In conclusion, GABA stimulates GABAA receptors located on inhibitory NANC neurones in the dog ileocolonic junction. Our results suggest that it is unlikely that GABA is the final inhibitory NANC neurotransmitter.

  3. Effects of Peripherally Restricted κ Opioid Receptor Agonists on Pain-Related Stimulation and Depression of Behavior in Rats

    PubMed Central

    O'Connell, Robert; Morrissey, Ember; Cheng, Kejun; Rice, Kenner C.

    2012-01-01

    κ Opioid receptor agonists that do not readily cross the blood-brain barrier are peripherally restricted and distribute poorly to the central nervous system after systemic administration. Peripherally restricted κ agonists have promise as candidate analgesics, because they may produce antinociception mediated by peripheral κ receptors more potently than they produce undesirable sedative and psychotomimetic effects mediated by central κ receptors. The present study used assays of pain-related stimulation and depression of behavior in rats to compare effects of 1) two peripherally restricted κ agonists [the tetrapeptide d-Phe-d-Phe-d-Ile-d-Arg-NH2 (ffir) and the nonpeptidic compound ((R,S)-N-[2-(N-methyl-3,4-dichlorophenylacetamido)-2-(3-carboxyphenyl)-ethyl]pyrrolidine hydrochloride (ICI204448)], 2) a centrally penetrating κ agonist (salvinorin A), and 3) several reference drugs, including a nonsteroidal anti-inflammatory drug (NSAID; ketoprofen). Intraperitoneal injection of dilute lactic acid served as a noxious stimulus to stimulate a stretching response and depress intracranial self-stimulation (ICSS) maintained by the delivery of electrical brain stimulation to the medial forebrain bundle. Acid-stimulated stretching was blocked by ketoprofen, the peripherally restricted κ agonists, and salvinorin A. However, acid-induced depression of ICSS was blocked only by ketoprofen. The peripherally restricted κ agonists had little effect, and salvinorin A exacerbated acid-induced depression of ICSS. These results suggest that peripherally restricted κ agonists may be safer than centrally penetrating κ agonists but less efficacious than NSAIDS or μ opioid receptor agonists to block pain-related depression of behavior; however, the peripheral selectivity of ffir and ICI204448 is limited, and future studies with κ agonists capable of greater peripheral selectivity are warranted. PMID:22128346

  4. Roles of mitogen activated protein kinases and EGF receptor in arsenite-stimulated matrix metalloproteinase-9 production

    SciTech Connect

    Cooper, Karen L.; Myers, Terrance Alix; Rosenberg, Martina; Chavez, Miquella; Hudson, Laurie G. . E-mail: lghudson@unm.edu

    2004-11-01

    The dermatotoxicity of arsenic is well established and epidemiological studies identify an increased incidence of keratinocytic tumors (basal cell and squamous cell carcinoma) associated with arsenic exposure. Little is known about the underlying mechanisms of arsenic-mediated skin carcinogenesis, but activation of mitogen-activated protein (MAP) kinases and subsequent regulation of downstream target genes may contribute to tumor promotion and progression. In this study, we investigated activation of the extracellular signal regulated kinase (ERK) and the stress-associated kinase p38 by arsenite in HaCat cells, a spontaneously immortalized human keratinocyte cell line. Arsenite concentrations {>=}100 {mu}M stimulate rapid activation of p38 and ERK MAP kinases. However, upon extended exposure (24 h), persistent stimulation of p38 and ERK MAP kinases was detected at low micromolar concentrations of arsenite. Although ERK and p38 were activated with similar time and concentration dependence, the mechanism of activation differed for these two MAP kinases. ERK activation by arsenite was fully dependent on the catalytic activity of the epidermal growth factor (EGF) receptor and partially dependent on Src-family kinase activity. In contrast, p38 activation was independent of EGF receptor or Src-family kinase activity. Arsenite-stimulated MAP kinase signal transduction resulted in increased production of matrix metalloproteinase (MMP)-9, an AP-1 regulated gene product. MMP-9 induction by arsenite was prevented when EGF receptor or MAP kinase signaling was inhibited. These studies indicate that EGF receptor activation is a component of arsenite-mediated signal transduction and gene expression in keratinocytes and that low micromolar concentrations of arsenite stimulate key signaling pathways upon extended exposure. Stimulation of MAP kinase cascades by arsenic and subsequent regulation of genes including c-fos, c-jun, and the matrix degrading proteases may play an important

  5. Circulating thyroid stimulating hormone receptor messenger RNA and differentiated thyroid cancer: A diagnostic meta-analysis

    PubMed Central

    Kong, Chao-Yue; Li, Zhan-Ming; Wang, Li-Shun

    2017-01-01

    Thyroid stimulating hormone receptor messenger RNA (TSHR-mRNA) is over-expressed in thyroid cancer patients, which indicates that TSHR-mRNA is a potential biomarker of thyroid cancer. However, system evaluation for TSHR-mRNA as a diagnostic biomarker of thyroid cancer is deficient. The performance of TSHR-mRNA for thyroid cancer diagnosis was evaluated in this study. Three common international databases as well as a Chinese database were applied for literature researching. Quality assessment of the included literatures was conducted by the QUADAS-2 tool. Totally, 1027 patients from nine studies eligible for the meta-analysis were included in this study. Global sensitivity and specificity for the positivity of TSHR-mRNA in the thyroid cancer diagnosis is 72% and 82%. The value of AUC for this test performance was 0.84. Our meta-analysis suggests that TSHR-mRNA might be a potential biomarker to complete present diagnostic methods for early and precision diagnosis of thyroid cancer. Notably, this findings need validation thorough large-scale clinical studies. PMID:28036261

  6. Endotoxin down-modulates granulocyte colony-stimulating factor receptor (CD114) on human neutrophils.

    PubMed

    Hollenstein, U; Homoncik, M; Stohlawetz, P J; Marsik, C; Sieder, A; Eichler, H G; Jilma, B

    2000-07-01

    During infection, the development of nonresponsiveness to granulocyte colony-stimulating factor (G-CSF) may be influenced by the down-modulation of G-CSF receptor (G-CSFR) by cytokines. This down-modulation was studied during experimental human endotoxemia. Healthy volunteers received either 2 ng/kg endotoxin (lipopolysaccharide [LPS], n=20) or placebo (n=10) in a randomized, controlled trial. Endotoxin infusion increased the mean fluorescence intensity of the neutrophil activation marker CD11b >300% after 1 h (P<.001 vs. placebo). LPS infusion down-modulated G-CSFR expression in as early as 60 min (-17%; P=.001 vs. placebo). Down-modulation was almost maximal at 90 min and persisted for 6 h (-50% from baseline; P<.0001 vs. placebo). Plasma levels of G-CSF started to increase only after G-CSFR down-modulation had occurred and peaked 37-fold above baseline at 4 h (P<.0001 vs. placebo). In conclusion, LPS down-modulates G-CSFR expression in humans, which may render neutrophils less responsive to the effects of G-CSF and, thereby, compromise host defense mechanisms.

  7. A negative allosteric modulator demonstrates biased antagonism of the follicle stimulating hormone receptor

    PubMed Central

    Dias, James A.; Bonnet, Béatrice; Weaver, Barbara A.; Watts, Julie; Kluetzman, Kerri; Thomas, Richard M.; Poli, Sonia; Mutel, Vincent; Campo, Brice

    2015-01-01

    High quality gamete production in males and females requires the pituitary gonadotropin follicle stimulating hormone (FSH). In this report a novel chemical class of small molecule inhibitors of FSH receptor (FSHR) is described. ADX61623, a negative allosteric modulator (NAM), increased the affinity of interaction between 125I-hFSH and human FSHR (hFSHR) five fold. This form of FSHR occupied simultaneously by FSH and ADX61623 was inactive for cAMP and progesterone production in primary cultures of rat granulosa cells. In contrast, ADX61623 did not block estrogen production. This demonstrates for the first time, biased antagonism at the FSHR. To determine if ADX61623 blocked FSH induction of follicle development in vivo, a bioassay to measure follicular development and oocyte production in immature female rats was validated. ADX61623 was not completely effective in blocking FSH induced follicular development in vivo at doses up to 100 mg/kg as oocyte production and ovarian weight gain were only moderately reduced. These data illustrate that FSHR couples to multiple signaling pathways in vivo. Suppression of one pool of FSHR uncouples Gαs and cAMP production, and decreases progesterone production. Occupancy of another pool of FSHR sensitizes granulosa cells to FSH induced estradiol production. Therefore, ADX61623 is a useful tool to investigate further the mechanism of the FSHR signaling dichotomy. This may lead to a greater understanding of the signaling infrastructure which enables estrogen biosynthesis and may prove useful in treating estrogen dependent disease. PMID:21184806

  8. Rapid intracellular release of calcium in human platelets by stimulation of 5-HT2-receptors.

    PubMed Central

    Erne, P.; Pletscher, A.

    1985-01-01

    The concentration of intracellular free Ca2+ ( [Ca2+]i) in human blood platelets was measured by use of the fluorescent probe quin-2. 5-Hydroxytryptamine (5-HT) caused a rapid increase of [Ca2+]i in the presence or absence of Ca2+ in the medium. The [Ca2+]i-rise was less marked in the absence of Ca2+ and could be antagonized by 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate-hydrochloride (TMB-8), an inhibitor of calcium release from internal stores. 5-HT induced a shape change reaction in the presence or absence of extracellular Ca2+, but the pEC50 of 5-HT was slightly higher in the presence of the cation. Shape change reaction and [Ca2+]i-rise showed similar time courses. Various 5-HT-agonists caused a rise of [Ca2+]i, whereas 5-HT-antagonists, but not the 5-HT-uptake inhibitor desmethylimipramine and the alpha 2-adrenoceptor antagonist yohimbine, counteracted the 5-HT-induced rise of the cation in a stereospecific manner. The antagonists were more potent than the agonists. The orders of potencies of the drugs affecting [Ca2+]i and platelet shape were similar. It is concluded that stimulation of 5-HT2-receptors of platelets causes a rapid release of intracellular calcium which, by activation of the contractile system, mediates the shape change reaction. PMID:3156650

  9. Lipopolysaccharide stimulation upregulated Toll-like receptor 4 expression in chicken cerebellum.

    PubMed

    Ansari, Abdur Rahman; Wen, Le; Huang, Hai-Bo; Wang, Ji-Xiang; Huang, Xi-Yao; Peng, Ke-Mei; Liu, Hua-Zhen

    2015-08-15

    Toll-like receptors (TLRs) play crucial roles in innate and adaptive immune responses to invading pathogens. TLR4 is responsible for the recognition of bacterial lipopolysaccharide (LPS) in different parts of central nervous system of many vertebrates. To better understand the functions of TLR4 in cerebellum of chicken, present study was designed to identify the cell types that express TLR4 during postnatal stages as well as the changes in its expression in response to LPS challenge. For this purpose, cerebella were collected from chicken aged 1, 14 and 40 days (n=7 in each group) to analyze TLR4 distribution pattern. The cerebella from 14 chickens injected with LPS or sterilizing saline were also collected at Day 14 (n=7 in each group) to investigate changes in TLR4 expression. This expression was analyzed by immunohistochemistry using an anti-TLR4 antibody. TLR4 was constitutively expressed in the Purkinje cell layer, pia mater, neurons in medulla and blood vessels in the cerebellum and LPS stimulation significantly up-regulated TLR4 expression on Day 14 in the chicken cerebellum. This study provides evidence that neurons in chicken cerebellum can express TLR4 in vivo and suggests that these neurons may play an important role in initiating a defense reaction via activation of TLR4. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Stimulation of oral fibroblast chemokine receptors identifies CCR3 and CCR4 as potential wound healing targets.

    PubMed

    Buskermolen, Jeroen K; Roffel, Sanne; Gibbs, Susan

    2017-11-01

    The focus of this study was to determine which chemokine receptors are present on oral fibroblasts and whether these receptors influence proliferation, migration, and/or the release of wound healing mediators. This information may provide insight into the superior wound healing characteristics of the oral mucosa. The gingiva fibroblasts expressed 12 different chemokine receptors (CCR3, CCR4, CCR6, CCR9, CCR10, CXCR1, CXCR2, CXCR4, CXCR5, CXCR7, CX3CR1, and XCR1), as analyzed by flow cytometry. Fourteen corresponding chemokines (CCL5, CCL15, CCL20, CCL22, CCL25, CCL27, CCL28, CXCL1, CXCL8, CXCL11, CXCL12, CXCL13, CX3CL1, and XCL1) were used to study the activation of these receptors on gingiva fibroblasts. Twelve of these fourteen chemokines stimulated gingiva fibroblast migration (all except for CXCL8 and CXCL12). Five of the chemokines stimulated proliferation (CCL5/CCR3, CCL15/CCR3, CCL22/CCR4, CCL28/CCR3/CCR10, and XCL1/XCR1). Furthermore, CCL28/CCR3/CCR10 and CCL22/CCR4 stimulation increased IL-6 secretion and CCL28/CCR3/CCR10 together with CCL27/CCR10 upregulated HGF secretion. Moreover, TIMP-1 secretion was reduced by CCL15/CCR3. In conclusion, this in-vitro study identifies chemokine receptor-ligand pairs which may be used in future targeted wound healing strategies. In particular, we identified the chemokine receptors CCR3 and CCR4, and the mucosa specific chemokine CCL28, as having an predominant role in oral wound healing by increasing human gingiva fibroblast proliferation, migration, and the secretion of IL-6 and HGF and reducing the secretion of TIMP-1. © 2017 The Authors. Journal of Cellular Physiology Published by wiley periodicals, Inc.

  11. Sigma 1 receptor stimulation protects against oxidative damage through suppression of the ER stress responses in the human lens.

    PubMed

    Wang, Lixin; Eldred, Julie A; Sidaway, Peter; Sanderson, Julie; Smith, Andrew J O; Bowater, Richard P; Reddan, John R; Wormstone, I Michael

    2012-01-01

    Stimulation of sigma-1 receptors is reported to protect against oxidative stress. The present study uses cells and tissue from the human lens to elucidate the relationship between the sigma 1 receptor, ER stress and oxidative stress-induced damage. Exposure of the human lens cell line FHL124 to increasing concentrations of H(2)O(2) led to reduced cell viability and increased apoptosis. In response to 30 μM H(2)O(2), levels of the ER stress proteins BiP, ATF6 and pEIF2α were significantly increased within 4h of exposure. Expression of the sigma 1 receptor was markedly increased in response to H(2)O(2). Application of 10 and 30 μM (+)-pentazocine, a sigma 1 receptor agonist, significantly inhibited the H(2)O(2) induced cell death. (+)-Pentazocine also suppressed the oxidative stress induced reduction of pro-caspase 12 and suppressed the induction of the ER stress proteins BiP and EIF2α. When applied to cultured human lenses, (+)-pentazocine protected against apoptotic cell death, LDH release and against H(2)O(2) induced opacification. These data demonstrate that stimulation of the sigma 1 receptor provides significant protection from oxidative damage and is, therefore, a putative therapeutic approach to delay the onset of diseases that may be triggered by oxidative damage, including cataract formation.

  12. Adenosine receptors mediate synergistic stimulation of glucose uptake and transport by insulin and by contractions in rat skeletal muscle.

    PubMed Central

    Vergauwen, L; Hespel, P; Richter, E A

    1994-01-01

    The role of adenosine receptors in the regulation of muscle glucose uptake by insulin and contractions was studied in isolated rat hindquarters that were perfused with a standard medium containing no insulin or a submaximal concentration of 100 microU/ml. Adenosine receptor antagonism was induced by caffeine or 8-cyclopentyl-1,3-dipropylxantine (CPDPX). Glucose uptake and transport were measured before and during 30 min of electrically induced muscle contractions. Caffeine nor CPDPX affected glucose uptake in resting hindquarters. In contrast, the contraction-induced increase in muscle glucose uptake was inhibited by 30-50% by caffeine, as well as by CPDPX, resulting in a 20-25% decrease in the absolute rate of glucose uptake during contractions, compared with control values. This inhibition was independent of the rate of perfusate flow and only occurred in hindquarters perfused with insulin added to the medium. Thus, adenosine receptor antagonism inhibited glucose uptake during simultaneous exposure to insulin and contractions only. Accordingly, caffeine inhibited 3-O-methylglucose uptake during contractions only in oxidative muscle fibers that are characterized by a high sensitivity to insulin. In conclusion, the present data demonstrate A1 receptors to regulate insulin-mediated glucose transport in contracting skeletal muscle. The findings provide evidence that stimulation of sarcolemmic adenosine receptors during contractions is involved in the synergistic stimulation of muscle glucose transport by insulin and by contractions. PMID:8132783

  13. The Non-Benzodiazepine Anxiolytic Drug Etifoxine Causes a Rapid, Receptor-Independent Stimulation of Neurosteroid Biosynthesis

    PubMed Central

    do Rego, Jean Luc; Vaudry, David; Vaudry, Hubert

    2015-01-01

    Neurosteroids can modulate the activity of the GABAA receptors, and thus affect anxiety-like behaviors. The non-benzodiazepine anxiolytic compound etifoxine has been shown to increase neurosteroid concentrations in brain tissue but the mode of action of etifoxine on neurosteroid formation has not yet been elucidated. In the present study, we have thus investigated the effect and the mechanism of action of etifoxine on neurosteroid biosynthesis using the frog hypothalamus as an experimental model. Exposure of frog hypothalamic explants to graded concentrations of etifoxine produced a dose-dependent increase in the biosynthesis of 17-hydroxypregnenolone, dehydroepiandrosterone, progesterone and tetrahydroprogesterone, associated with a decrease in the production of dihydroprogesterone. Time-course experiments revealed that a 15-min incubation of hypothalamic explants with etifoxine was sufficient to induce a robust increase in neurosteroid synthesis, suggesting that etifoxine activates steroidogenic enzymes at a post-translational level. Etifoxine-evoked neurosteroid biosynthesis was not affected by the central-type benzodiazepine (CBR) receptor antagonist flumazenil, the translocator protein (TSPO) antagonist PK11195 or the GABAA receptor antagonist bicuculline. In addition, the stimulatory effects of etifoxine and the triakontatetraneuropeptide TTN, a TSPO agonist, were additive, indicating that these two compounds act through distinct mechanisms. Etifoxine also induced a rapid stimulation of neurosteroid biosynthesis from frog hypothalamus homogenates, a preparation in which membrane receptor signalling is disrupted. In conclusion, the present study demonstrates that etifoxine stimulates neurosteroid production through a membrane receptor-independent mechanism. PMID:25785994

  14. The role of nucleus accumbens shell GABA receptors on ventral tegmental area intracranial self-stimulation and a potential role for the 5-HT(2C) receptor.

    PubMed

    Hayes, Dave J; Hoang, John; Greenshaw, Andrew J

    2011-12-01

    Brain γ-aminobutyric acid (GABA) and 5-hydroxytryptamine (5-HT)(2C) receptors are implicated in the neuronal regulation of reward- and aversion-related behaviour. Within the mesocorticolimbic pathways of the brain, relationships between GABA containing neurons and 5-HT(2C) receptor activity may be important in this context. The primary aim of this study was to investigate the role of NAc shell GABA receptors on ventral tegmental area intracranial self-stimulation (ICSS) and to examine the systemic effects of GABAergic ligands in this context. The second aim was to investigate the relationship between GABA receptor- and 5-HT(2C) receptor-related ICSS behaviour, using systemic administration of the selective agonist WAY 161503. Locomotor activity was assessed to compare the potential motor effects of drugs; feeding behaviour and intra-NAc injections of amphetamine (1.0 µg/side) were used as positive controls. When administered systemically the GABA(A) receptor agonist muscimol and antagonist picrotoxin did not selectively change ICSS reward thresholds, although the 5-HT(2C) receptor agonist WAY 161503 (1.0 mg/kg) decreased reward measures. Intra-NAc shell administration of muscimol (225 ng/side) and picrotoxin (125 ng/side), respectively, decreased and increased measures of reward. Intra-NAc shell baclofen (0-225 ng/side; GABA(B) receptor agonist) did not affect any ICSS measures although it increased feeding. Combining picrotoxin and WAY 161503 attenuated the effects of each. These results suggest that a 5-HT(2C) and GABA(A) receptor-mediated neuronal relationship in the NAc shell may be relevant for the regulation of brain reward pathways.

  15. mu-Opioid receptor stimulation in the nucleus accumbens elevates fatty tastant intake by increasing palatability and suppressing satiety signals.

    PubMed

    Katsuura, Yoshihiro; Heckmann, Jennifer A; Taha, Sharif A

    2011-07-01

    Infusion of a μ-opioid receptor (MOR) agonist into the nucleus accumbens (NAcc) drives voracious food intake, an effect hypothesized to occur through increased tastant palatability. While intake of many palatable foods is elevated by MOR stimulation, this manipulation has a preferential effect on fatty food ingestion. Consumption of high-fat foods is increased by NAcc MOR stimulation even in rats that prefer a carbohydrate-rich alternative under baseline conditions. This suggests that NAcc MOR stimulation may not simply potentiate palatability signals and raises the possibility that mechanisms mediating fat intake may be distinct from those underlying intake of other tastants. The present study was conducted to investigate the physiological mechanisms underlying the effects of NAcc MOR stimulation on fatty food intake. In experiment 1, we analyzed lick microstructure in rats ingesting Intralipid to identify the changes underlying feeding induced by infusion of a MOR-specific agonist into the NAcc. MOR stimulation in the NAcc core, but not shell, increased burst duration and first-minute licks, while simultaneously increasing the rate and duration of Intralipid ingestion. These results suggest that MOR activation in the core increases Intralipid palatability and attenuates inhibitory postingestive feedback. In experiment 2, we measured the effects of MOR stimulation in the NAcc core on consumption of nonnutritive olestra. A MOR-specific agonist dose dependently increased olestra intake, demonstrating that caloric signaling is not required for hyperphagia induced by NAcc MOR stimulation. Feeding induced by drug infusion in both experiments 1 and 2 was blocked by a MOR antagonist. In experiment 3, we determined whether MOR activation in the NAcc core could attenuate satiety-related signaling caused by infusion of the melanocortin agonist MTII into the third ventricle. Suppression of intake caused by MTII was reversed by MOR stimulation. Together, our results suggest

  16. The differential effects of 5-HT(1A) receptor stimulation on dopamine receptor-mediated abnormal involuntary movements and rotations in the primed hemiparkinsonian rat.

    PubMed

    Dupre, Kristin B; Eskow, Karen L; Negron, Giselle; Bishop, Christopher

    2007-07-16

    Serotonin 1A receptor (5-HT(1A)R) agonists have emerged as valuable supplements to l-DOPA therapy, demonstrating that they can decrease side effects and enhance motor function in animal models of Parkinson's disease (PD) and human PD patients. The precise mechanism by which these receptors act remains unknown and there is limited information on how 5-HT(1A)R stimulation impacts striatal dopamine (DA) D1 receptor (D1R) and D2 receptor (D2R) function. The current study examined the effects of 5-HT(1A)R stimulation on DA receptor-mediated behaviors. Male Sprague-Dawley rats were rendered hemiparkinsonian by unilateral 6-OHDA lesions and primed with the D1R agonist SKF81297 (0.8 mg/kg, i.p.) in order to sensitize DA receptors. Using a randomized within subjects design, rats received a first injection of: Vehicle (dH(2)O) or the 5-HT(1A)R agonist +/-8-OH-DPAT (0.1 or 1.0 mg/kg, i.p.), followed by a second injection of: Vehicle (dimethyl sulfoxide), the D1R agonist SKF81297 (0.8 mg/kg, i.p.), the D2R agonist quinpirole (0.2 mg/kg, i.p.), or l-DOPA (12 mg/kg+benserazide, 15 mg/kg, i.p.). On test days, rats were monitored over a 2-h period immediately following the second injection for abnormal involuntary movements (AIMs), analogous to dyskinesia observed in PD patients, and contralateral rotations. The present findings indicate that 5-HT(1A)R stimulation reduces AIMs induced by D1R, D2R and l-DOPA administration while its effects on DA agonist-induced rotations were receptor-dependent, suggesting that direct 5-HT(1A)R and DA receptor interactions may contribute to the unique profile of 5-HT(1A)R agonists for the improvement of PD treatment.

  17. Activation of Distinct P2Y Receptor Subtypes Stimulates Insulin Secretion in MIN6 Mouse Pancreatic β Cells

    PubMed Central

    Balasubramanian, Ramachandran; de Azua, Inigo Ruiz; Wess, Jürgen; Jacobson, Kenneth A.

    2010-01-01

    Extracellular nucleotides and their receptor antagonists have therapeutic potential in disorders such as inflammation, brain disorders, and cardiovascular diseases. Pancreatic β cells express several purinergic receptors, and reported nucleotide effects on insulin secretion are contradictory. We studied the effect of P2Y receptors on insulin secretion and cell death in MIN6, mouse pancreatic β cells. Expression of P2Y1 and P2Y6 receptors was revealed by total mRNA analysis using RT-PCR. MIN6 cells were stimulated in the presence of 16.7 mM glucose with or without P2Y1 and P2Y6 agonists, 2-MeSADP and Up3U, respectively. Both the agonists increased insulin secretion with EC50 values of 44.6±7.0 nM and 30.7±12.7 nM respectively. The insulin secretion by P2Y1 and P2Y6 agonists was blocked by their selective antagonists MRS2179 and MRS2578, respectively. Binding of the selective P2Y1 receptor antagonist radioligand [125I]MRS2500 in MIN6 cell membranes was saturable (KD 4.74±0.47 nM), and known P2Y1 ligands competed with high affinities. Inflammation and glucose toxicity leads to pancreatic β cell death in diabetes. Flow cytometric analysis revealed that Up3U but not 2-MeSADP protected MIN6 cells against TNF-α induced apoptosis. Overall, the results demonstrate that selective stimulation of P2Y1 and P2Y6 receptors increases insulin secretion that accompanies intracellular calcium release, suggesting potential application of P2Y receptor ligands in the treatment of diabetes. PMID:20067775

  18. Stimulation of 5-HT(1B) receptors enhances cocaine reinforcement yet reduces cocaine-seeking behavior.

    PubMed

    Pentkowski, Nathan S; Acosta, Jazmin I; Browning, Jenny R; Hamilton, Elizabeth C; Neisewander, Janet L

    2009-09-01

    Paradoxically, stimulation of 5-HT(1B) receptors (5-HT(1B)Rs) enhances sensitivity to the reinforcing effects of cocaine but attenuates incentive motivation for cocaine as measured using the extinction/reinstatement model. We revisited this issue by examining the effects of a 5-HT(1B)R agonist, CP94253, on cocaine reinforcement and cocaine-primed reinstatement, predicting that CP94253 would enhance cocaine-seeking behavior reinstated by a low priming dose, similar to its effect on cocaine reinforcement. Rats were trained to self-administer cocaine (0.75 mg/kg, i.v.) paired with light and tone cues. For reinstatement experiments, they then underwent daily extinction training to reduce cocaine-seeking behavior (operant responses without cocaine reinforcement). Next, they were pre-treated with CP94253 (3-10 mg/kg, s.c.) and either tested for cocaine-primed (10 or 2.5 mg/kg, i.p.) or cue-elicited reinstatement of extinguished cocaine-seeking behavior. For reinforcement, effects of CP94253 (5.6 mg/kg) across a range of self-administered cocaine doses (0-1.5 mg/kg, i.v.) were examined. Cocaine dose-dependently reinstated cocaine-seeking behavior, but contrary to our prediction, CP94253 reduced reinstatement with both priming doses. Similarly, CP94253 reduced cue-elicited reinstatement. In contrast, CP94253 shifted the self-administration dose-effect curve leftward, consistent with enhanced cocaine reinforcement. When saline was substituted for cocaine, CP94253 reduced response rates (i.e. cocaine-seeking behavior). In subsequent control experiments, CP94253 decreased open-arm exploration in an elevated plus-maze suggesting an anxiogenic effect, but had no effect on locomotion or sucrose reinforcement. These results provide strong evidence that stimulation of 5-HT(1B)Rs produces opposite effects on cocaine reinforcement and cocaine-seeking behavior, and further suggest that 5-HT(1B)Rs may be a novel target for developing medications for cocaine dependence.

  19. Sympathetic nerves in the mediation of renal response to localized stimulation of atrial receptors in anaesthetized dogs.

    PubMed Central

    Karim, F; Majid, D S; Summerill, R A

    1989-01-01

    1. Dogs were anaesthetized with chloralose and artificially ventilated. Localized stimulation of left atrial receptors for 23-25 min was achieved by distension of three small balloons at the pulmonary vein-atrial junctions and one in atrial appendage. Renal blood flows were measured by electromagnetic flow probes, glomerular filtration rate by creatinine clearance, urinary sodium excretion by flame photometry and solute excretion by osmometry. The mean aortic pressure was held constant at 92.2 +/- 2.4 mmHg (mean +/- S.E.M., n = 27) by means of a pressure bottle connected to the aorta and beta-adrenergic receptor activity was blocked by continuous infusion of propranolol (17 micrograms kg-1 min-1, I.V.). 2. In twelve dogs stimulation of left atrial receptors resulted in significant increases of 11.8 +/- 2.4% (P less than 0.001) in renal blood flow; 32.5 +/- 7.2% (P less than 0.001) in glomerular filtration rate; 19.5 +/- 5.0% (P less than 0.005) in filtration fraction: 36.3 +/- 9.0% (P less than 0.001) in urine flow: 32.7 +/- 9.2% (P less than 0.005) in sodium excretion: 36.6 +/- 9.9% (P less than 0.005) in osmolar excretion and a decrease of 31.3 +/- 11.2% (P less than 0.025) in free water clearance. Left atrial pressure and heart rate did not change significantly. In eight of the dogs ligation of the renal nerves resulted in similar changes in all of the renal variables; subsequent stimulation of atrial receptors did not cause significant changes in the renal variables. 3. In five additional dogs, in which heart rate and aortic pressure were allowed to change, stimulation of left atrial receptors for the same period resulted in significant increases in heart rate (4.3 +/- 0.7%. P less than 0.001) and mean aortic pressure (2.0 +/- 0.6%, P less than 0.025). Under this condition both the intact right kidneys and the denervated left kidneys showed significant responses in urine flow, sodium excretion, osmolar excretion and free water clearance. 4. The results show

  20. Vagus nerve stimulation protects against ventricular fibrillation independent of muscarinic receptor activation.

    PubMed

    Brack, Kieran E; Coote, John H; Ng, G André

    2011-08-01

    The role of the vagus in the ventricle is controversial, although the vagus can protect against ventricular fibrillation (VF) via nitric oxide (NO). This study aims to determine whether the mechanisms involved are dependent on post-ganglionic release and muscarinic receptor activation. For this purpose, NO release and electrophysiological effects of vagus nerve stimulation (VNS) were evaluated in relation to acetylcholine and vasoactive intestinal peptide (VIP). In addition, the role of the coronary endothelium and afferent nerves was tested. Using the isolated innervated rabbit heart, we measured ventricular NO release using 4,5-diaminofluorescein (DAF-2) fluorescence and ventricular fibrillation threshold (VFT) during VNS after muscarinic, ganglionic, and VIP inhibition [atropine, hexamethonium, and VIP (6-28), respectively] and after Triton-X endothelial functional dysfunction. The vagal-mediated increases in NO and VFT were not significantly affected (P> 0.05) during (i) atropine perfusion [increase in NO: 196.8 ± 35.2 mV (control) vs. 156.1 ± 20.3 mV (atropine) and VFT 3.1 ± 0.5 mA (control) vs. 2.7 ± 0.4 mA (atropine)], (ii) VIP inhibition-increase in NO: 243.0 ± 42.4 mV (control) vs. 203.9 ± 28.5 mV [VIP(6-28)] and VFT 3.3 ± 0.3 mA (control) vs. 3.9 ± 0.6 mA [VIP(6-28)], or (iii) after endothelial functional dysfunction [increase in NO: 127.7 ± 31.7 mV (control) vs. 172.1 ± 31.5 mV (Triton-X) and VFT 2.6 ± 0.4 mA (control) vs. 2.5 ± 0.5 mA (Triton-X)]. However, the vagal effects were inhibited during ganglionic blockade [increase in NO: 175.1 ± 38.1 mV (control) vs. 0.6 ± 25.3 mV (hexamethonium) and VFT 3.3 ± 0.5 mA (control) vs. -0.3 ± 0.3 mA (hexamethonium)]. We show that the vagal anti-fibrillatory action in the rabbit ventricle occurs via post-ganglionic efferent nerve fibres, independent of muscarinic receptor activation, VIP, and the endothelium. Together with our previous publications, our data support the possibility of a novel

  1. The effect on the efferent vagal nerves to the heart of stimulating atrial receptors in the dog.

    PubMed

    Walters, G E; Mary, D A

    1986-10-01

    In chloralose-anaesthetized dogs, distension of small balloons at the pulmonary vein-atrial junctions to stimulate atrial receptors with myelinated vagal afferent nerves causes an increase in heart rate but does not influence the activity in efferent vagal cardiac nerves. However, distension of these small balloons also stimulates atrial receptors with non-myelinated vagal and sympathetic afferent nerves, which are thought to affect the heart rate and activity in efferent vagal cardiac nerves. In the present investigation, seven dogs anaesthetized with chloralose were studied by distension of small balloons at the pulmonary vein-atrial junctions and in the left atrial appendage, and by graded cooling of the vagal nerves in the neck; cooling to 9 degrees C was used to prevent the increase in activity in myelinated vagal afferent nerves to distension of the small balloons and cooling to 0 degree C was used to prevent responses to the distension in all vagal afferent nerves. Eleven vagal efferent nerve fibers were studied which responded to stimulation of carotid baroreceptors and chemoreceptors. Distension of the small balloons did not affect the activity in these eleven efferent vagal nerve fibres, with the vagi at 37 degrees C or during vagal cooling to 9 degrees C or to 0 degree C. The results indicate that upon distension of the small balloons, none of the three types of atrial receptor influence the activity in efferent vagal cardiac nerves. The results support the conclusion that stimulation of atrial receptors with myelinated vagal afferent nerves, responsible for the reflex increase in heart rate, does not influence the activity in efferent vagal cardiac nerves.

  2. Angiotensin II stimulates expression of the chemokine RANTES in rat glomerular endothelial cells. Role of the angiotensin type 2 receptor.

    PubMed Central

    Wolf, G; Ziyadeh, F N; Thaiss, F; Tomaszewski, J; Caron, R J; Wenzel, U; Zahner, G; Helmchen, U; Stahl, R A

    1997-01-01

    Glomerular influx of monocytes/macrophages (M/M) occurs in many immune- and non-immune-mediated renal diseases. The mechanisms targeting M/M into the glomerulus are incompletely understood, but may involve stimulated expression of chemokines. We investigated whether angiotensin II (ANG II) induces the chemokine RANTES in cultured glomerular endothelial cells of the rat and in vivo. ANG II stimulated mRNA and protein expression of RANTES in cultured glomerular endothelial cells. The ANG II-induced RANTES protein was chemotactic for human monocytes. Surprisingly, the ANG II-stimulated RANTES expression was transduced by AT2 receptors because the AT2 receptor antagonists PD 123177 and CGP-42112A, but not an AT1 receptor blocker, abolished the induced RANTES synthesis. Intraperitoneal infusion of ANG II (500 ng/h) into naive rats for 4 d significantly stimulated glomerular RANTES mRNA and protein expression compared with solvent-infused controls. Immunohistochemistry revealed induction of RANTES protein mainly in glomerular endothelial cells and small capillaries. Moreover, ANG II- infused animals exhibited an increase in glomerular ED-1- positive cells compared with controls. Oral treatment with PD 123177 (50 mg/liter drinking water) attenuated the glomerular M/M influx without normalizing the slightly elevated systolic blood pressure caused by ANG II infusion, suggesting that the effects on blood pressure and RANTES induction can be separated. We conclude that the vasoactive peptide ANG II may play an important role in glomerular chemotaxis of M/M through local induction of the chemokine RANTES. The observation that the ANG II- mediated induction of RANTES is transduced by AT2 receptors may influence the decision as to which substances might be used for the therapeutic interference with the activity of the renin-angiotensin system. PMID:9276721

  3. Retinoids Stimulate Periosteal Bone Resorption by Enhancing the Protein RANKL, a Response Inhibited by Monomeric Glucocorticoid Receptor*

    PubMed Central

    Conaway, H. Herschel; Pirhayati, Amir; Persson, Emma; Pettersson, Ulrika; Svensson, Olle; Lindholm, Catharina; Henning, Petra; Tuckermann, Jan; Lerner, Ulf H.

    2011-01-01

    Increased vitamin A (retinol) intake has been suggested to increase bone fragility. In the present study, we investigated effects of retinoids on bone resorption in cultured neonatal mouse calvarial bones and their interaction with glucocorticoids (GC). All-trans-retinoic acid (ATRA), retinol, retinalaldehyde, and 9-cis-retinoic acid stimulated release of 45Ca from calvarial bones. The resorptive effect of ATRA was characterized by mRNA expression of genes associated with osteoclast differentiation, enhanced osteoclast number, and bone matrix degradation. In addition, the RANKL/OPG ratio was increased by ATRA, release of 45Ca stimulated by ATRA was blocked by exogenous OPG, and mRNA expression of genes associated with bone formation was decreased by ATRA. All retinoid acid receptors (RARα/β/γ) were expressed in calvarial bones. Agonists with affinity to all receptor subtypes or specifically to RARα enhanced the release of 45Ca and mRNA expression of Rankl, whereas agonists with affinity to RARβ/γ or RARγ had no effects. Stimulation of Rankl mRNA by ATRA was competitively inhibited by the RARα antagonist GR110. Exposure of calvarial bones to GC inhibited the stimulatory effects of ATRA on 45Ca release and Rankl mRNA and protein expression. This inhibitory effect was reversed by the glucocorticoid receptor (GR) antagonist RU 486. Increased Rankl mRNA stimulated by ATRA was also blocked by GC in calvarial bones from mice with a GR mutation that blocks dimerization (GRdim mice). The data suggest that ATRA enhances periosteal bone resorption by increasing the RANKL/OPG ratio via RARα receptors, a response that can be inhibited by monomeric GR. PMID:21715325

  4. Localization and expression of follicle-stimulating hormone receptor gene in buffalo (Bubalus bubalis) pre-antral follicles.

    PubMed

    Sharma, G Taru; Dubey, P K; Kumar, G Sai

    2011-02-01

    Follicle-stimulating hormone (FSH) stimulates antral follicles to grow, but its role in earlier stages (pre-antral) of follicle development, if any, is obscure. Aim of this study was to study the expression of follicle-stimulating hormone receptor (FSHR) gene in different sizes of pre-antral follicles (PFs) (<150, 200, 250, 300, 350, 400 μm) and to find out an optimum dose of FSH for better growth, development and steroidogenesis of PFs in vitro. Buffalo ovaries were collected from a local abattoir, and PFs were isolated by mechanical method. A semi-quantitative RT-PCR amplification strategy was used for mRNA expression, while FSHR protein was localized by immunohistochemistry. Isolated pre-antral follicles (80-85 μm) were cultured in TCM-199 supplemented with 10% foetal bovine serum, 1% ITS and 30 ng/ml EGF served as control medium. Addition of three different doses of FSH (0.5, 1.0, 2.0 μg/ml) in control medium was considered as treatment groups. A single 2.184-kb receptor mRNA transcript was present in all sizes (<150-400 μm) of follicles. Follicle-stimulating hormone receptor was also localized immunohistochemically in granulosa cells of all sizes of follicles. Survival and growth rate of follicles significantly (p<0.05) increased following supplementation of FSH at a concentration of 1.0 μg/ml and the culture medium also showed a significantly (p<0.05) greater accumulation of oestradiol and progesterone. In conclusion, FSHR is expressed in all sizes of PFs and in vitro survival, growth and steroidogenesis of follicles are optimally stimulated by 1.0 μg/ml FSH. These findings demonstrate that FSH has an important role during the recruitment, growth and development of buffalo ovarian PFs.

  5. Growth differentiation factor-9 stimulates progesterone synthesis in granulosa cells via a prostaglandin E2/EP2 receptor pathway.

    PubMed

    Elvin, J A; Yan, C; Matzuk, M M

    2000-08-29

    Growth differentiation factor-9 (GDF-9), an oocyte-secreted member of the transforming growth factor beta superfamily, progesterone receptor, cyclooxygenase 2 (Cox2; Ptgs2), and the EP2 prostaglandin E(2) (PGE(2)) receptor (EP2; Ptgerep2) are required for fertility in female but not male mice. To define the interrelationship of these factors, we used a preovulatory granulosa cell culture system in which we added recombinant GDF-9, prostaglandins, prostaglandin receptor agonists, or cyclooxygenase inhibitors. GDF-9 stimulated Cox2 mRNA within 2 h, and PGE(2) within 6 h; however, progesterone was not increased until 12 h after addition of GDF-9. This suggested that Cox2 is a direct downstream target of GDF-9 but that progesterone synthesis required an intermediate. To determine whether prostaglandin synthesis was required for progesterone production, we analyzed the effects of PGE(2) and cyclooxygenase inhibitors on this process. PGE(2) can stimulate progesterone synthesis by itself, although less effectively than GDF-9 (3-fold vs. 6-fold increase over 24 h, respectively). Furthermore, indomethacin or NS-398, inhibitors of Cox2, block basal and GDF-9-stimulated progesterone synthesis. However, addition of PGE(2) to cultures containing both GDF-9 and NS-398 overrides the NS-398 block in progesterone synthesis. To further define the PGE(2)-dependent pathway, we show that butaprost, a specific EP2 agonist, stimulates progesterone synthesis and overrides the NS-398 block. In addition, GDF-9 stimulates EP2 mRNA synthesis by a prostaglandin- and progesterone-independent pathway. Thus, GDF-9 induces an EP2 signal transduction pathway which appears to be required for progesterone synthesis in cumulus granulosa cells. These studies further demonstrate the importance of oocyte-somatic cell interactions in female reproduction.

  6. Receptor-targeted, magneto-mechanical stimulation of osteogenic differentiation of human bone marrow-derived mesenchymal stem cells.

    PubMed

    Hu, Bin; El Haj, Alicia J; Dobson, Jon

    2013-09-23

    Mechanical cues are employed to promote stem cell differentiation and functional tissue formation in tissue engineering and regenerative medicine. We have developed a Magnetic Force Bioreactor (MFB) that delivers highly targeted local forces to cells at a pico-newton level, utilizing magnetic micro- and nano-particles to target cell surface receptors. In this study, we investigated the effects of magnetically targeting and actuating specific two mechanical-sensitive cell membrane receptors-platelet-derived growth factor receptor α (PDGFRα) and integrin ανβ3. It was found that a higher mineral-to-matrix ratio was obtained after three weeks of magneto-mechanical stimulation coupled with osteogenic medium culture by initially targeting PDGFRα compared with targeting integrin ανβ3 and non-treated controls. Moreover, different initiation sites caused a differentiated response profile when using a 2-day-lagged magneto-mechanical stimulation over culture periods of 7 and 12 days). However, both resulted in statistically higher osteogenic marker genes expression compared with immediate magneto-mechanical stimulation. These results provide insights into important parameters for designing appropriate protocols for ex vivo induced bone formation via magneto-mechanical actuation.

  7. Locus Coeruleus Stimulation Facilitates Long-Term Depression in the Dentate Gyrus That Requires Activation of β-Adrenergic Receptors

    PubMed Central

    Hansen, Niels; Manahan-Vaughan, Denise

    2015-01-01

    Synaptic plasticity comprises a cellular mechanism through which the hippocampus most likely enables memory formation. Neuromodulation, related to arousal, is a key aspect in information storage. The activation of locus coeruleus (LC) neurons by novel experience leads to noradrenaline release in the hippocampus at the level of the dentate gyrus (DG). We explored whether synaptic plasticity in the DG is influenced by activation of the LC via electrical stimulation. Coupling of test-pulses that evoked stable basal synaptic transmission in the DG with stimulation of the LC induced β-adrenoreceptor-dependent long-term depression (LTD) at perforant path–DG synapses in adult rats. Furthermore, persistent LTD (>24 h) induced by perforant path stimulation also required activation of β-adrenergic receptors: Whereas a β-adrenergic receptor antagonist (propranolol) prevented, an agonist (isoproterenol) strengthened the persistence of LTD for over 24 h. These findings support the hypothesis that persistent LTD in the DG is modulated by β-adrenergic receptors. Furthermore, LC activation potently facilitates DG LTD. This suggests in turn that synaptic plasticity in the DG is tightly regulated by activity in the noradrenergic system. This may reflect the role of the LC in selecting salient information for subsequent synaptic processing in the hippocampus. PMID:24464942

  8. Inhibitory 5-hydroxytryptamine receptors involved in pressor effects obtained by stimulation of sympathetic outflow from spinal cord in pithed rats.

    PubMed Central

    Morán, A; Velasco, C; Salvador, T; Martín, M L; San Román, L

    1994-01-01

    1. A study was made of the effects of 5-hydroxytryptamine (5-HT) on pressor response induced in vivo by electrical stimulation of the sympathetic outflow from the spinal cord of pithed rats. All animals had been pretreated with atropine. Intravenous infusion of 5-hydroxytryptamine at doses of 10 and 20 micrograms kg-1 min-1 reduced the pressor effects obtained by electrical stimulation at intervals of 10 min over the 1 h of infusion. 2. This inhibitory action of 5-HT was depressed by cyproheptadine and methiothepin but was not modified by ketanserin or MDL-72222. By contrast, the inhibitory action of 5-HT was lost in pithed rats that had been pretreated with exogenous noradrenaline. 3. The 5-HT1 receptor agonist 5-carboxamidotryptamine (5-CT) caused an inhibition of the pressor response, whereas the 5-HT3 receptor agonist, 1-phenylbiguanide, produced a variable but significant increase in the pressor response. The 5-HT2 receptor agonist, m-CPP, did not modify the pressor sympathetic response. 4. Our results suggest that 5-hydroxytryptamine interferes with sympathetic neurotransmission by inhibiting pressor effects as a result of stimulation of the complete sympathetic outflow, and that this inhibition is mainly through a presynaptic 5-HT1 mechanism. PMID:7889292

  9. Expression cloning of a human granulocyte colony-stimulating factor receptor: a structural mosaic of hematopoietin receptor, immunoglobulin, and fibronectin domains

    PubMed Central

    1990-01-01

    We report the isolation from a placental library, of two cDNAs that can encode high affinity receptors for granulocyte colony-stimulating factor (G-CSF) when expressed in COS-7 cells. The cDNAs are predicted to encode integral membrane proteins of 759 and 812 amino acids in length. The predicted extracellular and membrane spanning sequences of the two clones are identical, as are the first 96 amino acids of their respective cytoplasmic regions. Different COOH termini of 34 or 87 residues are predicted for the two cDNAs, due apparently to alternate splicing. The receptor with the longer cytoplasmic domain is the closest human homologue of the murine G-CSF receptor recently described by Fukunaga et al. (Fukunaga, R., E. Ishizaka-Ikeda, Y. Seto, and S. Nagata. 1990. Cell. 61:341). A hybridization probe derived from the placental G-CSF receptor cDNA detects a approximately 3-kb transcript in RNAs isolated from placenta and a number of lymphoid and myeloid cells. The extracellular region of the G-CSF receptors is composed of four distinct types of structural domains, previously recognized in other cell surface proteins. In addition to the two domains of the HP receptor family-defining region (Patthy, L. 1990. Cell. 61:13) it incorporates one NH2-terminal Ig-like domain, and three additional repeats of fibronectin type III-like domains. The presence of both an NH2-terminal Ig-like domain and multiple membrane-proximal FN3-like domains suggests that the G-CSF receptor may be derived from an ancestral NCAM-like molecule and that the G-CSF receptor may function in some adhesion or recognition events at the cell surface in addition to the binding of G-CSF. PMID:2147944

  10. Varenicline attenuates nicotine-enhanced brain-stimulation reward by activation of α4β2 nicotinic receptors in rats

    PubMed Central

    Spiller, Krista; Xi, Zheng-Xiong; Li, Xia; Ashby, Charles R.; Callahan, Patrick M.; Tehim, Ashok; Gardner, Eliot L.

    2009-01-01

    Varenicline, a partial α4β2 and full α7 nicotinic receptor agonist, has been shown to inhibit nicotine self-administration and nicotine-induced increases in extracellular dopamine in the nucleus accumbens. In the present study, we investigated whether varenicline inhibits nicotine-enhanced electrical brain-stimulation reward (BSR), and if so, which receptor subtypes are involved. Systemic administration of nicotine (0.25–1.0 mg/kg, i.p.) or varenicline (0.03–3 mg/kg, i.p.) produced biphasic effects, with low doses producing enhancement (e.g., decreased BSR threshold), and high doses inhibiting BSR. Pretreatment with low dose (0.03–1.0 mg/kg) varenicline dose-dependently attenuated nicotine (0.25 or 0.5 mg/kg)-enhanced BSR. The BSR-enhancing effect produced by varenicline was blocked by mecamylamine (a high affinity nicotinic receptor antagonist) or dihydro-β-erythroidine (a relatively selective nicotinic α4-containing receptor antagonist), but not methyllycaconitine (a selective α7 receptor antagonist), suggesting an effect mediated by activation of α4β2 receptors. This suggestion is supported by findings that the α4β2 receptor agonist SIB-1765F produced a dose-dependent enhancement of BSR, while pretreatment with SIB-1765F attenuated nicotine (0.5 mg/kg)-enhanced BSR. In contrast, the selective α7 receptor agonist ARR-17779, altered neither BSR itself nor nicotine-enhanced BSR, at any dose tested. These findings suggest that: 1) varenicline inhibits nicotine-enhanced BSR, supporting its use as a smoking cessation aid; and 2) varenicline-enhanced BSR by itself and varenicline's anti-nicotine effects are mediated by activation of α4β2, but not α7, receptors. PMID:19393252

  11. Activation of histamine H3 receptor decreased cytoplasmic Ca(2+) imaging during electrical stimulation in the skeletal myotubes.

    PubMed

    Chen, Yan; Paavola, Jere; Stegajev, Vasili; Stark, Holger; Chazot, Paul L; Wen, Jian Guo; Konttinen, Yrjö T

    2015-05-05

    Histamine is a neurotransmitter and chemical mediator in multiple physiological processes. Histamine H3 receptor is expressed in the nervous system, heart, and gastrointestinal tract; however, little is known about H3 receptor in skeletal muscle. The aim of this study was to investigate the role of H3 receptor in skeletal myotubes. The expression of H3 receptor and myosin heavy chain (MHC), a late myogenesis marker, was assessed by real-time PCR and immunostaining in C2C12 skeletal myogenesis and adult mid-urethral skeletal muscle tissues. H3 receptor mRNA showed a significant increase upon differentiation of C2C12 into myotubes: 1-, 26-, 91-, and 182-fold at days 0, 2, 4, and 6, respectively. H3 receptor immunostaining in differentiated C2C12 cells and adult skeletal muscles was positive and correlated with that of MHC. The functional role of H3receptor in differentiated myotubes was assessed using an H3 receptor agonist, (R)-a-methylhistamine ((R)-α-MeHA). Ca(2+) imaging, stimulated by electric pacing, was decreased by 55% after the treatment of mature C2C12 myotubes with 1μM (R)-α-MeHA for 10min and 20min, while treatment with 100nm (R)-α-MeHA for 5min caused 45% inhibition. These results suggested that H3 receptor may participate in the maintenance of the relaxed state and prevention of over-contraction in mature differentiated myotubes. The elucidation of the role of H3R in skeletal myogenesis and adult skeletal muscle may open a new direction in the treatment of skeletal muscle disorders, such as muscle weakness, atrophy, and myotonia in motion systems or peri-urethral skeletal muscle tissues.

  12. Novel Small Molecule Glucagon-Like Peptide-1 Receptor Agonist Stimulates Insulin Secretion in Rodents and From Human Islets

    PubMed Central

    Sloop, Kyle W.; Willard, Francis S.; Brenner, Martin B.; Ficorilli, James; Valasek, Kathleen; Showalter, Aaron D.; Farb, Thomas B.; Cao, Julia X.C.; Cox, Amy L.; Michael, M. Dodson; Gutierrez Sanfeliciano, Sonia Maria; Tebbe, Mark J.; Coghlan, Michael J.

    2010-01-01

    OBJECTIVE The clinical effectiveness of parenterally-administered glucagon-like peptide-1 (GLP-1) mimetics to improve glucose control in patients suffering from type 2 diabetes strongly supports discovery pursuits aimed at identifying and developing orally active, small molecule GLP-1 receptor agonists. The purpose of these studies was to identify and characterize novel nonpeptide agonists of the GLP-1 receptor. RESEARCH DESIGN AND METHODS Screening using cells expressing the GLP-1 receptor and insulin secretion assays with rodent and human islets were used to identify novel molecules. The intravenous glucose tolerance test (IVGTT) and hyperglycemic clamp characterized the insulinotropic effects of compounds in vivo. RESULTS Novel low molecular weight pyrimidine-based compounds that activate the GLP-1 receptor and stimulate glucose-dependent insulin secretion are described. These molecules induce GLP-1 receptor-mediated cAMP signaling in HEK293 cells expressing the GLP-1 receptor and increase insulin secretion from rodent islets in a dose-dependent manner. The compounds activate GLP-1 receptor signaling, both alone or in an additive fashion when combined with the endogenous GLP-1 peptide; however, these agonists do not compete with radiolabeled GLP-1 in receptor-binding assays. In vivo studies using the IVGTT and the hyperglycemic clamp in Sprague Dawley rats demonstrate increased insulin secretion in compound-treated animals. Further, perifusion assays with human islets isolated from a donor with type 2 diabetes show near-normalization of insulin secretion upon compound treatment. CONCLUSIONS These studies characterize the insulinotropic effects of an early-stage, small molecule GLP-1 receptor agonist and provide compelling evidence to support pharmaceutical optimization. PMID:20823098

  13. Transcriptional mechanisms that control expression of the macrophage colony-stimulating factor receptor locus.

    PubMed

    Rojo, Rocio; Pridans, Clare; Langlais, David; Hume, David A

    2017-08-15

    The proliferation, differentiation, and survival of cells of the macrophage lineage depends upon signals from the macrophage colony-stimulating factor (CSF) receptor (CSF1R). CSF1R is expressed by embryonic macrophages and induced early in adult hematopoiesis, upon commitment of multipotent progenitors to the myeloid lineage. Transcriptional activation of CSF1R requires interaction between members of the E26 transformation-specific family of transcription factors (Ets) (notably PU.1), C/EBP, RUNX, AP-1/ATF, interferon regulatory factor (IRF), STAT, KLF, REL, FUS/TLS (fused in sarcoma/ranslocated in liposarcoma) families, and conserved regulatory elements within the mouse and human CSF1R locus. One element, the Fms-intronic regulatory element (FIRE), within intron 2, is conserved functionally across all the amniotes. Lineage commitment in multipotent progenitors also requires down-regulation of specific transcription factors such as MYB, FLI1, basic leucine zipper transcriptional factor ATF-like (BATF3), GATA-1, and PAX5 that contribute to differentiation of alternative lineages and repress CSF1R transcription. Many of these transcription factors regulate each other, interact at the protein level, and are themselves downstream targets of CSF1R signaling. Control of CSF1R transcription involves feed-forward and feedback signaling in which CSF1R is both a target and a participant; and dysregulation of CSF1R expression and/or function is associated with numerous pathological conditions. In this review, we describe the regulatory network behind CSF1R expression during differentiation and development of cells of the mononuclear phagocyte system. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  14. NMDA receptor mediates chronic visceral pain induced by neonatal noxious somatic stimulation.

    PubMed

    Miranda, Adrian; Mickle, Aaron; Bruckert, Mitchell; Kannampalli, Pradeep; Banerjee, Banani; Sengupta, Jyoti N

    2014-12-05

    NMDA receptors (NMDAR) are important in the development and maintenance of central sensitization. Our objective was to investigate the role of spinal neurons and NMDAR in the maintenance of chronic visceral pain. Neonatal rats were injected with acidic saline adjusted to pH 4.0 in the gastrocnemius muscle every other day for 12 days. In adult rats, NR1 and NR2B subunits were examined in the lumbo-sacral (LS) spinal cord. A baseline, visceromotor response (VMR) to graded colorectal distension (CRD) was recorded before and after administration of the NMDA antagonist, CGS-19755. Extracellular recordings were performed from CRD-sensitive LS spinal neurons and pelvic nerve afferents (PNA) before and after CGS-19755. Rats that received pH 4.0 saline injections demonstrated a significant increase in the expression NR2B subunits and VMR response to CRD>20 mmHg. CGS-19755 (i.v. or i.t.) had no effect in naïve rats, but significantly decreased the response to CRD in pH 4.0 saline injected rats. CGS-19755 had no effect on the spontaneous firing of SL-A, but decreased that of SL-S. Similarly, CGS-19755 attenuates the responses of SL-S neurons to CRD, but had no effect on SL-A neurons or on the response characteristics of PNA fibers. Neonatal noxious somatic stimulation results in chronic visceral hyperalgesia and sensitizes a specific subpopulation of CRD-sensitive spinal neurons. The sensitization of these SL-S spinal neurons is attenuated by the NMDAR antagonist. The results of this study suggest that spinal NMDARs play an important role in the development of hyperalgesia early in life. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Effect of β2-adrenergic receptor stimulation on lung fluid in stable heart failure patients.

    PubMed

    Taylor, Bryan J; Snyder, Eric M; Richert, Maile L; Wheatley, Courtney M; Chase, Steven C; Olson, Lyle J; Johnson, Bruce D

    2017-04-01

    The purpose of this study was to determine: (1) whether stable heart failure patients with reduced ejection fraction (HFrEF) have elevated extravascular lung water (EVLW) when compared with healthy control subjects; and (2) the effect of acute β2-adrenergic receptor (β2AR) agonist inhalation on lung fluid balance. Twenty-two stable HFrEF patients and 18 age- and gender-matched healthy subjects were studied. Lung diffusing capacity for carbon monoxide (DLCO), alveolar-capillary membrane conductance (DmCO), pulmonary capillary blood volume (Vc) (via re-breathe) and lung tissue volume (Vtis) (via computed tomography) were assessed before and within 30 minutes after administration of nebulized albuterol. EVLW was derived as Vtis - Vc. Before administration of albuterol, Vtis and EVLW were higher in HFrEF vs control (998 ± 200 vs 884 ± 123 ml, p = 0.041; and 943 ± 202 vs 802 ± 133 ml, p = 0.015, respectively). Albuterol decreased Vtis and EVLW in HFrEF patients (-4.6 ± 7.8%, p = 0.010; -4.6 ± 8.8%, p = 0.018) and control subjects (-2.8 ± 4.9%, p = 0.029; -3.0 ± 5.7%, p = 0.045). There was an inverse relationship between pre-albuterol values and pre- to post-albuterol change for EVLW (r(2) = -0.264, p = 0.015) and DmCO (r(2) = -0.343, p = 0.004) in HFrEF only. Lung fluid is elevated in stable HFrEF patients relative to healthy subjects. Stimulation of β2ARs may cause fluid removal in HFrEF, especially in patients with greater evidence of increased lung water at baseline. Copyright © 2017 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.

  16. The role of lipolysis stimulated lipoprotein receptor in breast cancer and directing breast cancer cell behavior.

    PubMed

    Reaves, Denise K; Fagan-Solis, Katerina D; Dunphy, Karen; Oliver, Shannon D; Scott, David W; Fleming, Jodie M

    2014-01-01

    The claudin-low molecular subtype of breast cancer is of particular interest for clinically the majority of these tumors are poor prognosis, triple negative, invasive ductal carcinomas. Claudin-low tumors are characterized by cancer stem cell-like features and low expression of cell junction and adhesion proteins. Herein, we sought to define the role of lipolysis stimulated lipoprotein receptor (LSR) in breast cancer and cancer cell behavior as LSR was recently correlated with tumor-initiating features. We show that LSR was expressed in epithelium, endothelium, and stromal cells within the healthy breast tissue, as well as in tumor epithelium. In primary breast tumor bioposies, LSR expression was significantly correlated with invasive ductal carcinomas compared to invasive lobular carcinomas, as well as ERα positive tumors and breast cancer cell lines. LSR levels were significantly reduced in claudin-low breast cancer cell lines and functional studies illustrated that re-introduction of LSR into a claudin-low cell line suppressed the EMT phenotype and reduced individual cell migration. However, our data suggest that LSR may promote collective cell migration. Re-introduction of LSR in claudin-low breast cancer cell lines reestablished tight junction protein expression and correlated with transepithelial electrical resistance, thereby reverting claudin-low lines to other intrinsic molecular subtypes. Moreover, overexpression of LSR altered gene expression of pathways involved in transformation and tumorigenesis as well as enhanced proliferation and survival in anchorage independent conditions, highlighting that reestablishment of LSR signaling promotes aggressive/tumor initiating cell behaviors. Collectively, these data highlight a direct role for LSR in driving aggressive breast cancer behavior.

  17. Granulocyte colony-stimulating factor receptor expression on human transitional cell carcinoma of the bladder.

    PubMed Central

    Tachibana, M.; Miyakawa, A.; Uchida, A.; Murai, M.; Eguchi, K.; Nakamura, K.; Kubo, A.; Hata, J. I.

    1997-01-01

    Receptors for granulocyte colony-stimulating factor (G-CSFRs) have been confirmed on the cell surfaces of several non-haematopoietic cell types, including bladder cancer cells. This observation has naturally led to the hypothesis that the expression of G-CSFR on these cells may enhance their growth by G-CSF. In this study, the expression of G-CSFR was determined in both established human bladder cancer cell lines and primary bladder cancers. We studied five different human bladder cancer cell lines (KU-1, KU-7, T-24, NBT-2 and KK) and 26 newly diagnosed bladder tumours. G-CSFR mRNA expressions on cultured cell lines were determined using the reverse transcriptase polymerase chain reaction (RT-PCR) method. Furthermore, the G-CSFR binding experiments on the cultured cell lines were conducted using the Na(125)I-labelled G-CSF ligand-binding assay method. Moreover, the G-CSFR mRNA expressions on primary bladder tumour specimens were assessed using the in situ RT-PCR method. Three out of the five cultured cell lines (KU-1, NBT-2 and KK) exhibited G-CSFR mRNA signals when the RT-PCR method was used. The G-CSFR binding experiments showed an equilibrium dissociation constant (K[d]) of 490 pM for KU-1, 340 pM for NBT-2 and 103 pM for KK cells. With in situ RT-PCR, the tumour cells of 6 out of 26 primary bladder tumour specimens (23.1%) presented positive G-CSFR mRNA signals. Thus, in this study, G-CSFR expression was frequently observed on bladder cancer cells. Therefore, the clinical use of G-CSF for patients with bladder cancer should be selected with great care. Images Figure 1 Figure 3 Figure 4 PMID:9166942

  18. NMDA receptor mediates chronic visceral pain induced by neonatal noxious somatic stimulation

    PubMed Central

    Miranda, Adrian; Mickle, Aaron; Bruckert, Mitchell; Kannampalli, Pradeep; Banerjee, Banani; Sengupta, Jyoti N.

    2014-01-01

    NMDA receptors (NMDAR) are important in the development and maintenance of central sensitization. Our objective was to investigate the role of spinal neurons and NMDAR in the maintenance of chronic visceral pain. Neonatal rats were injected with acidic saline adjusted to pH4.0 in the gastrocnemius muscle every other day for 12 days. In adult rats, NR1 and NR2B subunits were examined in the lumbo-sacral (LS) spinal cord. A baseline, visceromotor response (VMR) to graded colorectal distension (CRD) was recorded before and after administration of the NMDA antagonist, CGS-19755. Extracellular recordings were performed from CRD-sensitive LS spinal neurons and pelvic nerve afferents (PNA) before and after CGS-19755. Rats that received pH 4.0 saline injections demonstrated a significant increase in the expression NR2B subunits and VMR response to CRD >20mmHg. CGS-19755 (i.v. or i.t.) had no effect in naïve rats, but significantly decreased the response to CRD in pH4.0 saline injected rats. CGS-19755 had no effect on the spontaneous firing of SL-A, but decreased that of SL-S. Similarly, CGS-19755 attenuates the responses of SL-S neurons to CRD, but had no effect on SL-A neurons or on the response characteristics of PNA fibers. Neonatal noxious somatic stimulation results in chronic visceral hyperalgesia and sensitizes a specific subpopulation of CRD-sensitive spinal neurons. The sensitization of these SL-S spinal neurons is attenuated by the NMDAR antagonist. The results of this study suggest that spinal NMDARs play an important role in the development of hyperalgesia early in life. PMID:25281204

  19. Glucagon-like peptide-1 receptor stimulation increases GFR and suppresses proximal reabsorption in the rat.

    PubMed

    Thomson, Scott C; Kashkouli, Ali; Singh, Prabhleen

    2013-01-15

    The incretin hormone glucagon-like peptide-1 (GLP-1) is released from the gut in response to fat or carbohydrate and contributes to negative feedback control of blood glucose by stimulating insulin secretion, inhibiting glucagon, and slowing gastric emptying. GLP-1 receptors (GLP-1R) are also expressed in the proximal tubule, and possibly elsewhere in the kidney. Presently, we examined the effect of a GLP-1R agonist on single-nephron glomerular filtration rate (GFR; SNGFR), proximal reabsorption (Jprox), tubuloglomerular feedback (TGF) responses, and urine flow rate in hydropenic male Wistar and Wistar-Froemter rats. Micropuncture and whole-kidney data were obtained before and during infusion of the GLP-1 agonist exenatide (1 nmol/h iv). SNGFR and Jprox were measured by late proximal collection at both extremes of TGF activation, which was achieved by perfusing Henle's loop at 0 or 50 nl/min. Primary changes in Jprox were revealed by analysis of covariance for Jprox with SNGFR as a covariate. Effects on TGF activation were determined in a separate set of experiments by comparing early distal and late proximal collections. Exenatide increased SNGFR by 33-50%, suppressed proximal tubular reabsorption by 20-40%, doubled early distal flow rate, and increased urine flow rate sixfold without altering the efficiency of glomerulotubular balance, TGF responsiveness, or the tonic influence of TGF. This implies that exenatide is both a proximal diuretic and a renal vasodilator. Since the natural agonist for the GLP-1R is regulated by intake of fat and carbohydrate, but not by salt or fluid, the control of salt excretion by the GLP-1R system departs from the usual negative-feedback paradigm for regulating salt balance.

  20. Graves disease in children: thyroid-stimulating hormone receptor antibodies as remission markers.

    PubMed

    Gastaldi, Roberto; Poggi, Elena; Mussa, Alessandro; Weber, Giovanna; Vigone, Maria Cristina; Salerno, Mariacarolina; Delvecchio, Maurizio; Peroni, Elena; Pistorio, Angela; Corrias, Andrea

    2014-05-01

    To evaluate clinical and biochemical features of 115 children (98 female, mean age 11.3 ± 3.5 years) with Graves disease to identify possible determinants of remission. We defined as positive outcome the improvement of clinical features and restoration of euthyroidism or induction of hypothyroidism after antithyroid drug (ATD) therapy and as negative outcome hyperthyroidism persistent over 2 years of ATD therapy or relapsed after ATD withdrawal. Thirty-eight children (33%) had remission after 2 years of ATD therapy. The absence of goiter at diagnosis was correlated with a better outcome. Median thyroid-stimulating hormone receptor antibody (TRAb) values at diagnosis were significantly lower in patients with a positive outcome (P = .031). We found a significant relationship between the time required for TRAb normalization and the patient outcome; TRAb normalization within 1 year from time of Graves disease diagnosis was significantly more common among patients with a positive outcome (P < .0001), and the mean time for TRAb normalization was significantly shorter in patients with a positive outcome (1.3 ± 0.8 years) compared with that observed in patients with a negative outcome (2.5 ± 2.7 years, P = .026). Although no clinical variable investigated is constantly associated with a definite outcome, the absence of goiter at the diagnosis may be associated with a better outcome. The most relevant predictor of Graves disease outcome was serum level; TRAb at time of Graves disease diagnosis less than 2.5 times the upper reference limit, TRAb normalization during ATD, and TRAb normalization timing each may predict positive outcomes. These results may have a role in the empiric clinical management of pediatric patients with Graves disease. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Prevalence and clinical relevance of thyroid stimulating hormone receptor-blocking antibodies in autoimmune thyroid disease.

    PubMed

    Diana, T; Krause, J; Olivo, P D; König, J; Kanitz, M; Decallonne, B; Kahaly, G J

    2017-09-01

    The prevalence and clinical relevance of thyroid stimulating hormone (TSH) receptor (TSHR) blocking antibodies (TBAb) in patients with autoimmune thyroid disease (AITD) was investigated. Serum TBAb were measured with a reporter gene bioassay using Chinese hamster ovary cells. Blocking activity was defined as percentage inhibition of luciferase expression relative to induction with bovine TSH alone (cut-off 40% inhibition). All samples were measured for TSHR stimulatory antibody (TSAb) and TSHR binding inhibiting immunoglobulins (TBII). A total of 1079 unselected, consecutive patients with AITD and 302 healthy controls were included. All unselected controls were negative for TBAb and TSAb. In contrast, the prevalence of TBAb-positive patients with Hashimoto's thyroiditis and Graves' disease was 67 of 722 (9·3%) and 15 of 357 (4·2%). Of the 82 TBAb-positive patients, thirty-nine (48%), 33 (40%) and 10 (12%) were hypothyroid, euthyroid and hyperthyroid, respectively. Ten patients were both TBAb- and TSAb-positive (four hypothyroid, two euthyroid and four hyperthyroid). Thyroid-associated orbitopathy was present in four of 82 (4·9%) TBAb-positive patients, with dual TSHR antibody positivity being observed in three. TBAb correlated positively with TBII (r = 0·67, P < 0·001) and negatively with TSAb (r = -0·86, P < 0·05). The percentage of TBII-positive patients was higher the higher the level of inhibition in the TBAb assay. Of the TBAb-positive samples with  > 70% inhibition, 87% were TBII-positive. Functional TSHR antibodies impact thyroid status. TBAb determination is helpful in the evaluation and management of patients with AITD. The TBAb assay is a relevant and important tool to identify potentially reversible hypothyroidism. © 2017 British Society for Immunology.

  2. Antimicrobial peptide scolopendrasin VII, derived from the centipede Scolopendra subspinipes mutilans, stimulates macrophage chemotaxis via formyl peptide receptor 1.

    PubMed

    Park, Yoo Jung; Lee, Ha Young; Jung, Young Su; Park, Joon Seong; Hwang, Jae Sam; Bae, Yoe-Sik

    2015-08-01

    In this study, we report that one of the antimicrobial peptides scolopendrasin VII, derived from Scolopendra subspinipes mutilans, stimulates actin polymerization and the subsequent chemotactic migration of macrophages through the activation of ERK and protein kinase B (Akt) activity. The scolopendrasin VII-induced chemotactic migration of macrophages is inhibited by the formyl peptide receptor 1 (FPR1) antagonist cyclosporine H. We also found that scolopendrasin VII stimulate the chemotactic migration of FPR1-transfected RBL-2H3 cells, but not that of vector-transfected cells; moreover, scolopendrasin VII directly binds to FPR1. Our findings therefore suggest that the antimicrobial peptide scolopendrasin VII, derived from Scolopendra subspinipes mutilans, stimulates macrophages, resulting in chemotactic migration via FPR1 signaling, and the peptide can be useful in the study of FPR1-related biological responses.

  3. Measurement of thyroid stimulating immunoglobulins using a novel thyroid stimulating hormone receptor-guanine nucleotide-binding protein, (GNAS) fusion bioassay.

    PubMed

    Pierce, M; Sandrock, R; Gillespie, G; Meikle, A W

    2012-11-01

    Hyperthyroidism, defined by overproduction of thyroid hormones, has a 2-3% prevalence in the population. The most common form of hyperthyroidism is Graves' disease. A diagnostic biomarker for Graves' disease is the presence of immunoglobulins which bind to, and stimulate, the thyroid stimulating hormone receptor (TSHR), a G-protein coupled receptor (GPCR). We hypothesized that the ectopically expressed TSHR gene in a thyroid stimulating immunoglobulin (TSI) assay could be engineered to increase the accumulation of the GPCR pathway second messenger, cyclic AMP (cAMP), the molecule measured in the assay as a marker for pathway activation. An ectopically expressing TSHR-mutant guanine nucleotide-binding protein, (GNAS) Chinese hamster ovary (CHO) cell clone was constructed using standard molecular biology techniques. After incubation of the new clone with sera containing various levels of TSI, GPCR pathway activation was then quantified by measuring cAMP accumulation in the clone. The clone, together with a NaCl-free cell assay buffer containing 5% polyethylene glycol (PEG)6000, was tested against 56 Graves' patients, 27 toxic thyroid nodule patients and 119 normal patients. Using receiver operating characteristic analysis, when comparing normal with Graves' sera, the assay yielded a sensitivity of 93%, a specificity of 99% and an efficiency of 98%. Total complex precision (within-run, across runs and across days), presented as a percentage coefficient of variation, was found to be 7·8, 8·7 and 7·6% for low, medium and high TSI responding serum, respectively. We conclude that the performance of the new TSI assay provides sensitive detection of TSI, allowing for accurate, early detection of Graves' disease.

  4. Altered beta-adrenergic receptor-stimulated cAMP formation in cultured skin fibroblasts from Alzheimer donors.

    PubMed

    Huang, H M; Gibson, G E

    1993-07-15

    An alteration in signal transduction systems in Alzheimer's disease would likely be of pathophysiological significance, because these steps are critical to normal brain function. Since dynamic processes are difficult to study in autopsied brain, the current studies utilized cultured skin fibroblasts. The beta-adrenergic-stimulated increase in cAMP was reduced approximately 80% in fibroblasts from Alzheimer's disease compared with age-matched controls. The deficit in Alzheimer fibroblasts in response to various adrenergic agonists paralleled their beta-adrenergic potency, and enhancement of cAMP accumulation by a non-adrenergic agonist, such as prostaglandin E1, was similar in Alzheimer and control fibroblasts. Diminished adenylate cyclase activity did not underlie these abnormalities, since direct stimulation of adenylate cyclase by forskolin elevated cAMP production equally in Alzheimer and control fibroblasts. Cholera toxin equally stimulated cAMP formation in Alzheimer and control fibroblasts. Moreover, cholera toxin partially reduced isoproterenol-induced cAMP deficit in Alzheimer fibroblasts. Pertussis toxin, on the other hand, did not alter the Alzheimer deficits. The results suggest either that the coupling of the GTP-binding protein(s) to the beta-adrenergic receptor is abnormal or that the sensitivity of receptor is altered with Alzheimer's disease. Further, any hypothesis about Alzheimer's disease must explain why a reduced beta-adrenergic-stimulated cAMP formation persists in tissue culture.

  5. Transitions in oral and intestinal microflora composition and innate immune receptor-dependent stimulation during mouse development.

    PubMed

    Hasegawa, Mizuho; Osaka, Toshifumi; Tawaratsumida, Kazuki; Yamazaki, Takashi; Tada, Hiroyuki; Chen, Grace Y; Tsuneda, Satoshi; Núñez, Gabriel; Inohara, Naohiro

    2010-02-01

    Commensal bacteria possess immunostimulatory activities that can modulate host responses to affect development and homeostasis in the intestine. However, how different populations of resident bacteria stimulate the immune system remains largely unknown. We characterized here the ability of intestinal and oral microflora to stimulate individual pattern recognition receptors (PRRs) in bone marrow-derived macrophages and mesothelial cells. The intestinal but not oral microflora elicited age- and cell type-specific immunostimulation. The immunostimulatory activity of the intestinal microflora varied among individual mice but was largely mediated via Toll-like receptor 4 (TLR4) during breast-feeding, whereas it became TLR4 independent after weaning. This transition was associated with a change from a microflora rich in TLR4-stimulatory proteobacteria to one dominated by Bacteroidales and/or Clostridiales that poorly stimulate TLR4. The major stimulatory activity of the intestinal microflora was still intact in NOD1-, NOD2-, TLR2-, TLR4-, TLR5-, TLR9-, TLR11-, ASC-, or RICK-deficient cells but still relied on the adaptor MyD88. These studies demonstrate a transition in the intestinal microflora accompanied by a dynamic change of its ability to stimulate different PRRs which control intestinal homeostasis.

  6. Insulin-stimulated Na/sup +/ transport in a model renal epithelium: protein synthesis dependence and receptor interactions

    SciTech Connect

    Blazer-Yost, B.L.; Cox, M.

    1987-05-01

    The urinary bladder of the toad, Bufo marinus, is a well characterized model of the mammalian distal nephron. Porcine insulin (approx. 0.5-5.0 ..mu..M) stimulates net mucosal to serosal Na/sup +/ flux within 10 minutes of hormone addition. The response is maintained for at least 5 hr and is completely abolished by low doses (10..mu..M) of the epithelial Na/sup +/ channel blocker amiloride. Insulin-stimulated Na/sup +/ transport does not require new protein synthesis since it is actinomycin-D (10..mu..g/ml) insensitive. Also in 3 separate experiments in which epithelial cell proteins were examined by /sup 35/S-methionine labeling, 2-dimensional polyacrylamide gel electrophoresis/autoradiography, no insulin induced proteins were observed. Equimolar concentrations of purified porcine proinsulin and insulin (0.64..mu..M) stimulate Na/sup +/ transport to the same extent. Thus, the putative toad insulin receptor may have different affinity characteristics than those demonstrated for insulin and proinsulin in mammalian tissues. Alternatively, the natriferic action of insulin in toad urinary bladders may be mediated by occupancy of another receptor. Preliminary experiments indicating that nanomolar concentrations of IGF/sub 1/ stimulate Na/sup +/ transport in this tissue support the latter contention.

  7. Activation of adenosine A(3) receptors supports hematopoiesis-stimulating effects of granulocyte colony-stimulating factor in sublethally irradiated mice.

    PubMed

    Hofer, Michal; Pospísil, Milan; Sefc, Ludek; Dusek, Ladislav; Vacek, Antonín; Holá, Jirina; Hoferová, Zuzana; Streitová, Denisa

    2010-08-01

    Research areas of 'post-exposure treatment' and 'cytokines and growth factors' have top priority among studies aimed at radiological nuclear threat countermeasures. The experiments were aimed at testing the ability of N(6)-(3-iodobenzyl)adenosine-5'-N-methyluronamide (IB-MECA), an adenosine A(3) receptor agonist, to modulate hematopoiesis in sublethally irradiated mice, when administered alone or in a combination with granulocyte colony-stimulating factor (G-CSF) in a two-day post-irradiation treatment regimen. A complete analysis of hematopoiesis including determination of numbers of bone marrow hematopoietic progenitor and precursor cells, as well as of numbers of peripheral blood cells, was performed. The outcomes of the treatment were assessed at days 3 to 22 after irradiation. IB-MECA alone has been found to induce a significant elevation of numbers of bone marrow granulocyte-macrophage progenitor cells (GM-CFC) and peripheral blood neutrophils. IB-MECA given concomitantly with G-CSF increased significantly bone marrow GM-CFC and erythroid progenitor cells (BFU-E) in comparison with the controls and with animals administered each of the drugs alone. The findings suggest the ability of IB-MECA to stimulate hematopoiesis and to support the hematopoiesis-stimulating effects of G-CSF in sublethally irradiated mice.

  8. Modulation of the vagal bradycardia evoked by stimulation of upper airway receptors by central 5-HT1 receptors in anaesthetized rabbits

    PubMed Central

    Dando, Simon B; Skinner, Matthew R; Jordan, David; Ramage, Andrew G

    1998-01-01

    The effects of central application of 5-HT1A and 5-HT1B/1D receptor ligands on the reflex bradycardia, apnoea, renal sympathoexcitation and pressor response evoked by stimulating upper airway receptors with smoke in atenolol-pretreated anaesthetized rabbits were studied.Intracisternal administration of the 5-HT1A receptor antagonists WAY-100635 (100 μg kg−1) and (−)pindolol (100 μg kg−1) significantly reduced the smoke-induced bradycardia, attenuated the pressor response and in the case of (−)pindolol, sympathetic nerve activity. The same dose of WAY-100635 i.v. was without effect.Buspirone (200 μg kg−1, i.c.) potentiated the reflex bradycardia. This action was prevented if the animals were pretreated with WAY-100635 (100 μg kg−1, i.v.)(+)8-OH-DPAT (25 μg kg−1, i.c.) attenuated the evoked bradycardia, pressor response, apnoea and renal sympathoexcitation. The attenuation of the apnoea and renal sympathoexcitation, but not the bradycardia or pressor response was prevented in animals pretreated with WAY-100635 (100 μg kg−1, i.v.). The attenuation of the reflex bradycardia and the reduction in the renal sympathoexcitation were reduced by pretreatment with the 5-HT1B/1D receptor antagonist GR127935 (100 μg kg−1, i.v.).In WAY-100635 (100 μg kg−1, i.v.) pretreated animals, sumatriptan (a 5-HT1B/1D receptor agonist) reduced the reflex bradycardia and the pressor response. The 5-HT1B/1D receptor antagonist GR127935 (20 μg kg−1, i.c. or 100 μg kg−1, i.v.) had no effect on the reflex responses.In conclusion, the present data are consistent with the hypothesis that activation of central 5-HT1A receptors potentiate whilst activation of 5-HT1B/1D receptors attenuate the reflex activation of cardiac preganglionic vagal motoneurones evoked by stimulation of upper airway receptors with smoke in rabbits. PMID:9786516

  9. Role of follicle-stimulating hormone receptor Ser680Asn polymorphism in the efficacy of follicle-stimulating hormone.

    PubMed

    de Castro, Francisco; Ruiz, Rocío; Montoro, Luis; Pérez-Hernández, Dámaso; Sánchez-Casas Padilla, Elisa; Real, Luis M; Ruiz, Agustín

    2003-09-01

    To evaluate the association between FSH efficacy and FSHR alleles. Retrospective study. University-based fertility unit and a private center for biomedical research. One hundred two women with ovarian function who were undergoing controlled ovarian stimulation (COS). Women were categorized as poor responders (< or =3 ovarian follicles at the end of the cycle) or normal responders (>3 follicles). Daily administration of exogenous FSH. Number of good or poor responders. The allele frequency and genotype distribution of the Ser680Asn marker differed significantly between groups. Cycle cancellations were increased (21%) among women who were homozygous for Ser680 compared with Ser/Asn and Asn/Asn patients, and 36% of poor-responders were homozygous for Ser680. The results support a role for FSHR gene in COS outcome. However, the weight of this factor is probably low. The Ser680 allele may act in concert with other environmental and genetic factors that contribute to FSH efficacy.

  10. Sweet Taste Receptor Activation in the Gut Is of Limited Importance for Glucose-Stimulated GLP-1 and GIP Secretion.

    PubMed

    Saltiel, Monika Y; Kuhre, Rune E; Christiansen, Charlotte B; Eliasen, Rasmus; Conde-Frieboes, Kilian W; Rosenkilde, Mette M; Holst, Jens J

    2017-04-22

    Glucose stimulates the secretion of the incretin hormones: glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP). It is debated whether the sweet taste receptor (STR) triggers this secretion. We investigated the role of STR activation for glucose-stimulated incretin secretion from an isolated perfused rat small intestine and whether selective STR activation by artificial sweeteners stimulates secretion. Intra-luminal administration of the STR agonists, acesulfame K (3.85% w/v), but not sucralose (1.25% w/v) and stevioside (2.5% w/v), stimulated GLP-1 secretion (acesulfame K: 31 ± 3 pmol/L vs. 21 ± 2 pmol/L, p < 0.05, n = 6). In contrast, intra-arterial administration of sucralose (10 mM) and stevioside (10 mM), but not acesulfame K, stimulated GLP-1 secretion (sucralose: 51 ± 6 pmol/L vs. 34 ± 4 pmol/L, p < 0.05; stevioside: 54 ± 6 pmol/L vs. 32 ± 2 pmol/L, p < 0.05, n = 6), while 0.1 mM and 1 mM sucralose did not affect the secretion. Luminal glucose (20% w/v) doubled GLP-1 and GIP secretion, but basolateral STR inhibition by gurmarin (2.5 µg/mL) or the inhibition of the transient receptor potential cation channel 5 (TRPM5) by triphenylphosphine oxide (TPPO) (100 µM) did not attenuate the responses. In conclusion, STR activation does not drive GIP/GLP-1 secretion itself, nor does it have a role for glucose-stimulated GLP-1 or GIP secretion.

  11. Insulin and Metabolic Stress Stimulate Multisite Serine/Threonine Phosphorylation of Insulin Receptor Substrate 1 and Inhibit Tyrosine Phosphorylation*

    PubMed Central

    Hançer, Nancy J.; Qiu, Wei; Cherella, Christine; Li, Yedan; Copps, Kyle D.; White, Morris F.

    2014-01-01

    IRS1 and IRS2 are key substrates of the insulin receptor tyrosine kinase. Mass spectrometry reveals more than 50 phosphorylated IRS1 serine and threonine residues (Ser(P)/Thr(P) residues) in IRS1 from insulin-stimulated cells or human tissues. We investigated a subset of IRS1 Ser(P)/Thr(P) residues using a newly developed panel of 25 phospho-specific monoclonal antibodies (αpS/TmAbIrs1). CHO cells overexpressing the human insulin receptor and rat IRS1 were stimulated with insulin in the absence or presence of inhibitors of the PI3K → Akt → mechanistic target of rapamycin (mTOR) → S6 kinase or MEK pathways. Nearly all IRS1 Ser(P)/Thr(P) residues were stimulated by insulin and significantly suppressed by PI3K inhibition; fewer were suppressed by Akt or mTOR inhibition, and none were suppressed by MEK inhibition. Insulin-stimulated Irs1 tyrosine phosphorylation (Tyr(P)Irs1) was enhanced by inhibition of the PI3K → Akt → mTOR pathway and correlated with decreased Ser(P)-302Irs1, Ser(P)-307Irs1, Ser(P)-318Irs1, Ser(P)-325Irs1, and Ser(P)-346Irs1. Metabolic stress modeled by anisomycin, thapsigargin, or tunicamycin increased many of the same Ser(P)/Thr(P) residues as insulin, some of which (Ser(P)-302Irs1, Ser(P)-307Irs1, and four others) correlated significantly with impaired insulin-stimulated Tyr(P)Irs1. Thus, IRS1 Ser(P)/Thr(P) is an integrated response to insulin stimulation and metabolic stress, which associates with reduced Tyr(P)Irs1 in CHOIR/IRS1 cells. PMID:24652289

  12. Insulin and metabolic stress stimulate multisite serine/threonine phosphorylation of insulin receptor substrate 1 and inhibit tyrosine phosphorylation.

    PubMed

    Hançer, Nancy J; Qiu, Wei; Cherella, Christine; Li, Yedan; Copps, Kyle D; White, Morris F

    2014-05-02

    IRS1 and IRS2 are key substrates of the insulin receptor tyrosine kinase. Mass spectrometry reveals more than 50 phosphorylated IRS1 serine and threonine residues (Ser(P)/Thr(P) residues) in IRS1 from insulin-stimulated cells or human tissues. We investigated a subset of IRS1 Ser(P)/Thr(P) residues using a newly developed panel of 25 phospho-specific monoclonal antibodies (αpS/TmAb(Irs1)). CHO cells overexpressing the human insulin receptor and rat IRS1 were stimulated with insulin in the absence or presence of inhibitors of the PI3K → Akt → mechanistic target of rapamycin (mTOR) → S6 kinase or MEK pathways. Nearly all IRS1 Ser(P)/Thr(P) residues were stimulated by insulin and significantly suppressed by PI3K inhibition; fewer were suppressed by Akt or mTOR inhibition, and none were suppressed by MEK inhibition. Insulin-stimulated Irs1 tyrosine phosphorylation (Tyr(P)(Irs1)) was enhanced by inhibition of the PI3K → Akt → mTOR pathway and correlated with decreased Ser(P)-302(Irs1), Ser(P)-307(Irs1), Ser(P)-318(Irs1), Ser(P)-325(Irs1), and Ser(P)-346(Irs1). Metabolic stress modeled by anisomycin, thapsigargin, or tunicamycin increased many of the same Ser(P)/Thr(P) residues as insulin, some of which (Ser(P)-302(Irs1), Ser(P)-307(Irs1), and four others) correlated significantly with impaired insulin-stimulated Tyr(P)(Irs1). Thus, IRS1 Ser(P)/Thr(P) is an integrated response to insulin stimulation and metabolic stress, which associates with reduced Tyr(P)(Irs1) in CHO(IR)/IRS1 cells.

  13. Sweet Taste Receptor Activation in the Gut Is of Limited Importance for Glucose-Stimulated GLP-1 and GIP Secretion

    PubMed Central

    Saltiel, Monika Y.; Kuhre, Rune E.; Christiansen, Charlotte B.; Eliasen, Rasmus; Conde-Frieboes, Kilian W.; Rosenkilde, Mette M.; Holst, Jens J.

    2017-01-01

    Glucose stimulates the secretion of the incretin hormones: glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP). It is debated whether the sweet taste receptor (STR) triggers this secretion. We investigated the role of STR activation for glucose-stimulated incretin secretion from an isolated perfused rat small intestine and whether selective STR activation by artificial sweeteners stimulates secretion. Intra-luminal administration of the STR agonists, acesulfame K (3.85% w/v), but not sucralose (1.25% w/v) and stevioside (2.5% w/v), stimulated GLP-1 secretion (acesulfame K: 31 ± 3 pmol/L vs. 21 ± 2 pmol/L, p < 0.05, n = 6). In contrast, intra-arterial administration of sucralose (10 mM) and stevioside (10 mM), but not acesulfame K, stimulated GLP-1 secretion (sucralose: 51 ± 6 pmol/L vs. 34 ± 4 pmol/L, p < 0.05; stevioside: 54 ± 6 pmol/L vs. 32 ± 2 pmol/L, p < 0.05, n = 6), while 0.1 mM and 1 mM sucralose did not affect the secretion. Luminal glucose (20% w/v) doubled GLP-1 and GIP secretion, but basolateral STR inhibition by gurmarin (2.5 µg/mL) or the inhibition of the transient receptor potential cation channel 5 (TRPM5) by triphenylphosphine oxide (TPPO) (100 µM) did not attenuate the responses. In conclusion, STR activation does not drive GIP/GLP-1 secretion itself, nor does it have a role for glucose-stimulated GLP-1 or GIP secretion. PMID:28441725

  14. Effects of toluene exposure on signal transduction: toluene reduced the signaling via stimulation of human muscarinic acetylcholine receptor m2 subtypes in CHO cells.

    PubMed

    Tsuga, Hirofumi; Haga, Tatsuya; Honma, Takeshi

    2002-07-01

    The organic solvent toluene is used widely in industry and is toxic to the central nervous system (CNS). To clarify the mechanisms of CNS toxicity following toluene exposure, especially with respect to the G protein-coupling of receptors, we determined the effects of toluene on the activation of Gi by stimulating human muscarinic acetylcholine receptor m2 subtypes (hm2 receptors) expressed in Chinese hamster ovary (CHO) cells. We first examined whether toluene affects the inhibition of adenylyl cyclase by Gi. The attenuation of forskolin-stimulated cAMP formation by the stimulation of hm2 receptors was reduced in a medium containing toluene. Next, we determined the effects of toluene on carbamylcholine-stimulated [35S]GTPgammaS binding using membrane fractions of CHO cell expressing hm2 receptors. Carbamylcholine-stimulated [35S]GTPgammaS binding activity was markedly reduced when assayed using reaction buffers containing toluene. However, carbamylcholine-stimulated [35S]GTPgammaS binding activity was essentially unchanged following pretreatment of the cells with a toluene-saturated medium prior to membrane isolation. Toluene pretreatment and the toluene itself did not alter the characteristics of the binding of carbamylcholine and [3H]N-methylscopolamine to hm2 receptors. On the contrary of the effect of toluene for [35S]GTPgammaS binding, the effect of toluene for attenuation of forskolin-stimulated cAMP formation by the stimulation of hm2 receptors was irreversible. These observations indicate that toluene acts as an inhibitor of the signal transduction via hm2 receptor stimulation in CHO cells, and at least two mechanisms exist in the inhibition mechanisms by toluene.

  15. mu-opioid receptor-stimulated synthesis of reactive oxygen species is mediated via phospholipase D2.

    PubMed

    Koch, Thomas; Seifert, Anja; Wu, Dai-Fei; Rankovic, Marija; Kraus, Jürgen; Börner, Christine; Brandenburg, Lars-Ove; Schröder, Helmut; Höllt, Volker

    2009-08-01

    We have recently shown that the activation of the rat mu-opioid receptor (MOPr, also termed MOR1) by the mu-agonist [D-Ala(2), Me Phe(4), Glyol(5)]enkephalin (DAMGO) leads to an increase in phospholipase D2 (PLD2) activity and an induction of receptor endocytosis, whereas the agonist morphine which does not induce opioid receptor endocytosis fails to activate PLD2. We report here that MOPr-mediated activation of PLD2 stimulates production of reactive oxygen molecules via NADH/NADPH oxidase. Oxidative stress was measured with the fluorescent probe dichlorodihydrofluorescein diacetate and the role of PLD2 was assessed by the PLD inhibitor D-erythro-sphingosine (sphinganine) and by PLD2-small interfering RNA transfection. To determine whether NADH/NADPH oxidase contributes to opioid-induced production of reactive oxygen species, mu-agonist-stimulated cells were pre-treated with the flavoprotein inhibitor, diphenylene iodonium, or the specific NADPH oxidase inhibitor, apocynin. Our results demonstrate that receptor-internalizing agonists (like DAMGO, beta-endorphin, methadone, piritramide, fentanyl, sufentanil, and etonitazene) strongly induce NADH/NADPH-mediated ROS synthesis via PLD-dependent signaling pathways, whereas agonists that do not induce MOPr endocytosis and PLD2 activation (like morphine, buprenorphine, hydromorphone, and oxycodone) failed to activate ROS synthesis in transfected human embryonic kidney 293 cells. These findings indicate that the agonist-selective PLD2 activation plays a key role in the regulation of NADH/NADPH-mediated ROS formation by opioids.

  16. Role of brainstem adenosine A1 receptors in the cardiovascular response to hypothalamic defence area stimulation in the anaesthetized rat.

    PubMed Central

    St Lambert, J. H.; Dashwood, M. R.; Spyer, K. M.

    1996-01-01

    1. The role of centrally located adenosine A1 receptors in the cardiovascular changes associated with the hypothalamic defence response has been investigated by in vitro autoradiography and the intraventricular application of an A1 receptor antagonist. 2. 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX), a highly selective adenosine A1 antagonist and its vehicle, ethanol, were administered directly into the posterior portion of the fourth ventricle of alpha-chloralose anaesthetized, paralysed and artificially ventilated rats. 3. DPCPX (0.01 to 0.3 mg kg-1) caused a dose-dependent decrease in the magnitude of the evoked pressor response (from -13 to -23 mmHg) elicited on hypothalamic defence area stimulation at a dose 10 fold lower than that required to produce an equivalent effect following systemic administration whilst ethanol, the vehicle, had no effect. 4. In vitro autoradiography revealed a heterogeneous distribution of adenosine A1 binding sites in the lower brainstem of rats. Image analysis showed the ventrolateral medulla to have the highest density of A1 receptors. Intermediate levels of binding were seen in caudal regions of the nucleus tractus solitarii and the hypoglossal nucleus. 5. These data imply that a proportion of the cardiovascular response to hypothalamic defence area stimulation are produced by the activation of adenosine A1 receptors localized close to the surface of, or adjacent to, the fourth ventricle in the immediate vicinity of the injection site. PMID:8789379

  17. Requisite Role of Basolateral Amygdala Glucocorticoid Receptor Stimulation in Drug Context-Induced Cocaine-Seeking Behavior

    PubMed Central

    Stringfield, Sierra J.; Higginbotham, Jessica A.

    2016-01-01

    Background: Exposure to cocaine-associated stimuli triggers a robust rise in circulating glucocorticoid levels. Glucocorticoid receptors are richly expressed in the basolateral amygdala, a brain region that controls the reinstatement of cocaine-seeking behavior upon exposure to a previously cocaine-paired environmental context. In the present study, we investigated whether glucocorticoid receptor stimulation in the basolateral amygdala is integral to drug context-induced motivation to seek cocaine in a rat model of drug relapse. Methods: Rats were trained to lever press for cocaine reinforcement in a distinct environmental context and were then given daily extinction training sessions in a different context. At test, the rats received bilateral glucocorticoid receptor antagonist (mifepristone; 3 or 10ng/hemisphere) or vehicle microinfusions into either the basolateral amygdala or the overlying posterior caudate-putamen (anatomical control region). Immediately thereafter, drug-seeking behavior (i.e., nonreinforced lever presses) was assessed in the previously cocaine-paired context and locomotor activity was assessed in a novel context. Results: Intra-basolateral amygdala, but not intra-posterior caudate-putamen, mifepristone dose-dependently attenuated drug context-induced cocaine-seeking behavior relative to vehicle, such that responding was similar to that observed in the extinction context. In contrast, mifepristone treatment did not alter locomotor activity. Conclusions: These findings suggest that basolateral amygdala glucocorticoid receptor stimulation is necessary for drug context-induced motivation to seek cocaine. PMID:27521756

  18. The effect of blockade of dopamine receptors on the inhibition of episodic luteinizing hormone release during electrical stimulation of the arcuate nucleus in ovariectomized rats.

    PubMed

    Gallo, R V

    1978-04-01

    This study examined the possible involvement of dopamine (DA) in mediating the inhibition of episodic LH release that occurs during electrical stimulation of the arcuate nucleus (ARH) in ovariectomized rats. Animals were treated before stimulation with pimozide (1.26--2.0 mg/kg) or d-butaclamol (1 mg/kg), blockers of DA receptors, or l-butaclamol. Apomorphine, which inhibits episodic LH release by activating DA receptors, was given near the end of the experiment to determine if these receptors were blocked. ARH stimulation suppressed pulsatile LH release in six rats when DA receptors were not blocked by pimozide (as well as two in which blockade was not tested). A transient increase occurred in one other animal. When DA receptors were blocked by pimozide, stimulation of the ARH inhibited episodic LH release in nine rats, suggesting that DA may have no role in mediating this inhibition. However, because increased LH release occurred in five additional animals, as well as in one with partial receptor blockade, the possibility remains that DA may perhaps have a minor role in this inhibitory response. Although ARH stimulation increased LH release after DA receptor blockade by d-butaclamol, this effect could not be ascribed to the DA antagonist property of this agent, because elevated blood LH levels also occurred during stimulation in rats treated with l-butaclamol, in which DA receptors were not blocked. d- and l-butaclamol may possess a non-stereospecific action on a non-dopaminergic event, thus reversing the response to ARH stimulation. Finally, whether DA receptors were blocked or not by pimozide, d-, or l-butaclamol, activation of the ventromedial hypothalamic and periventricular nucleus regions suppressed episodic LH release, but did not increase LH secretion. This suggests that the region through which stimulation can inhibit, but not increase, LH release may extend in the hypothalamus to these two areas.

  19. Muscarinic receptor-stimulated phosphatidylinositol turnover in the rat corpus striatum: role of muscarinic receptor subtypes and regulation

    SciTech Connect

    Monsma, F.J.

    1987-01-01

    The coupling between the M1 and M2 muscarinic receptor subtypes and phosphatidylinositol (Pl) hydrolysis has been examined in the corpus striatum and cerebral cortex of the rat brain. Receptor binding by the various muscarinic ligands was assessed using a preparation of intact brain cell aggregates, under similar conditions as the assay of Pl hydrolysis. In striatal cell aggregates, (/sup 3/H)-quinuclidinyl benzilate ((/sup 3/H)-QNB) bound to a single class of muscarinic sites with high affinity, inhibition of (/sup 3/H)-QNB binding by muscarinic receptor ligands which exhibit selectivity for subtypes of the muscarinic receptor revealed the presence of both the M1 and M2 subtypes in approximately equal numbers.

  20. Identification and characterization of receptors for granulocyte colony-stimulating factor on human placenta and trophoblastic cells

    SciTech Connect

    Uzumaki, Hiroya; Okabe, Tetsuro; Sasaki, Norio; Hagiwara, Koichi; Takaku, Fumimaro; Tobita, Masahito; Yasukawa, Kaoru ); Ito, Seiga ); Umezawa, Yoshimi )

    1989-12-01

    Since radioiodination of human granulocyte colony-stimulating factor (G-CSF) is difficult, the authors synthesized a mutein of human G-CSF that retains full biological activity and receptor-binding capacity for at least 2 weeks after radioiodination. Receptors for human G-CSF were characterized in the plasma membrane fraction from the human term placenta (human placental membranes) and trophoblastic cells by using the {sup 125}I-labeled mutein of human G-CSF (KW-2228). The specific binding of {sup 125}I-labeled KW-2228 to placental membranes was pH-dependent, with maximal specific binding at pH 7.8; it increased linearly with protein to 3.7 mg of protein per ml and was both time- and temperature-dependent, with maximal binding at 4{degree}C after a 24-hr incubation. When the authors examined the ability of hematopoietic growth factors to inhibit {sup 125}I-labeled KW-2228 binding, they found that KW-2228 and intact human G-CSF ihibited {sup 125}I-labeled KW-2228 binding, whereas erythropoietin or granulocyte-macrophage colony-stimulating factor did not. Scatchard analysis revealed a single receptor type. The human G-CSF receptors on human placental membranes were shown to consist of two molecular species that could be specifically cross-linked to {sup 125}I-labeled KW-2228. Human trophoblastic cells, T3M-3, also possessed a single receptor for G-CSF. They have identified the receptor for human G-CSF on human placental membranes and trophoblastic cells.

  1. In adult female hamsters hypothyroidism stimulates D1 receptor-mediated breathing without altering D1 receptor expression.

    PubMed

    Schlenker, Evelyn H; Del Rio, Rodrigo; Schultz, Harold D

    2015-11-01

    Hypothyroidism affects cardiopulmonary regulation and function of dopaminergic receptors. Here we evaluated effects of 5 months of hypothyroidism on dopamine D1 receptor modulation of breathing in female hamsters using a D1 receptor antagonist SCH 23390. Euthyroid hamsters (EH) served as controls. Results indicated that hypothyroid female hamsters (HH) exhibited decreased body weights and minute ventilation (VE) following hypoxia due to decreased frequency of breathing (F). Moreover, SCH 23390 administration in HH increased VE by increasing tidal volume during exposure to air, hypoxia and following hypoxia. Relative to vehicle, SCH 23390 treatment decreased body temperature and hypoxic VE responsiveness in both groups. In EH, SCH 23390 decreased F in air, hypoxia and post hypoxia, and VE during hypoxia trended to decrease (P=0.053). Finally, expression of D1 receptor protein was not different between the two groups in any region evaluated. Thus, hypothyroidism in older female hamsters affected D1 receptor modulation of ventilation differently relative to euthyroid animals, but not expression of D1 receptors.

  2. In Adult Female Hamsters Hypothyroidism Stimulates D1 Receptor-mediated Breathing without altering D1 Receptor Expression

    PubMed Central

    Schlenker, Evelyn H.; Rio, Rodrigo Del; Schultz, Harold D.

    2015-01-01

    Hypothyroidism affects cardiopulmonary regulation and function of dopaminergic receptors. Here we evaluated effects of 5 months of hypothyroidism on dopamine D1 receptor modulation of breathing in female hamsters using a D1 receptor antagonist SCH23390. Euthyroid hamsters (EH) served as controls. Results indicated that hypothyroid female hamsters (HH) exhibited decreased body weights and minute ventilation (VE) following hypoxia due to decreased frequency of breathing (F). Moreover, SCH 23390 administration in HH increased VE by increasing tidal volume during exposure to air, hypoxia and following hypoxia. Relative to vehicle, SCH 23390 treatment decreased body temperature and hypoxic VE responsiveness in both groups. In EH, SCH 23390 decreased F in air, hypoxia and post hypoxia, and VE during hypoxia trended to decrease (P=0.053). Finally, expression of D1 receptor protein was not different between the two groups in any region evaluated. Thus, hypothyroidism in older female hamsters affected D1 receptor modulation of ventilation differently relative to euthyroid animals, but not expression of D1 receptors. PMID:26232642

  3. Modern approaches to the design of memory and cognitive function stimulants based on AMPA receptor ligands

    NASA Astrophysics Data System (ADS)

    Grigoriev, V. V.; Proshin, A. N.; Kinzirsky, A. S.; Bachurin, Sergey O.

    2009-05-01

    Data on the structure and properties of compounds acting on AMPA receptors, the key subtype of ionotropic glutamate receptors of the mammalian central nervous system, are analyzed. Data on the role of these receptors in provision of memory and cognitive function formation and impairment processes are presented. The attention is focused on the modern views on the mechanisms of AMPA receptor desensitization and deactivation and action of substances affecting these processes. The structures of key positive modulators of AMPA receptors are given. The problems of application of these substances as therapeutic means for preventing and treating neurodegenerative and psychoneurological diseases are discussed. Bibliography — 121 references.

  4. Pivotal roles of alpha-melanocyte-stimulating hormone and the melanocortin 4 receptor in leptin stimulation of prolactin secretion in rats.

    PubMed

    Watanobe, Hajime; Schiöth, Helgi B; Izumi, Junkichi

    2003-04-01

    Leptin, the obese gene product, was reported to stimulate prolactin (PRL) secretion, but the neuroendocrine mechanism underlying this hormonal response is largely unknown. Thus, in this study we examined the involvement of several important PRL regulators in the leptin-induced PRL secretion in male rats. Compared with the values in normally fed rats, food deprivation for 3 days significantly decreased both PRL and leptin levels in the plasma. These changes were reverted to normal by a 3-day constant infusion of 75 microg/kg/day of leptin to the fasted rats, while 225 microg/kg/day of leptin further elevated both PRL and leptin levels. These four groups of animals were used for the following experiments. Results of dopamine and serotonin turnover studies in the brain and the pituitary indicated that neither of these biogenic amines plays a primary role in mediating leptin's effects on PRL. Repeated intracerebroventricular injections over 72 h of neutralizing antibodies against vasoactive intestinal peptide, PRL-releasing peptide, or beta-endorphin, did not significantly suppress the leptin actions. However, both the blockade of the melanocortin (MC) 4 receptor (R) and the immunoquenching of brain alpha-melanocyte-stimulating hormone (alpha-MSH) completely abolished the leptin-induced PRL release, and the stimulation of the MC4-R, but not the MC3-R, significantly elevated PRL levels in the fasted rats. These results suggest that alpha-MSH, a cleaved peptide from pro-opiomelanocortin of which synthesis is stimulated by leptin, may be the pivotal neuropeptide in the brain mediating the leptin's stimulatory influence on PRL secretion. It was also suggested that the MC4-R may be the primary subtype of the MC-Rs mediating this action of alpha-MSH.

  5. Ascites fluid containing monoclonal antibody (EGFR1) to the epidermal growth factor (EGF) receptor (EGFR) stimulates tyrosine phosphorylation in solubilized placental membranes

    SciTech Connect

    Michiel, D.F.; Hollenberg, M.D.

    1986-03-05

    Ascites fluid containing EGFR1 antibody precipitated EGFR prelabelled by phosphorylating human placental membranes with EGF and (..gamma..-/sup 32/P)ATP. The ascites fluid (at 1:100 dilution) was also found to stimulate phosphorylation of the solubilized placental membranes EGFR and the 35 kDa Ca/sup 2 +/-dependent substrate of the EGFR/kinase from placental membrane. In intact membranes, ascites fluid produced a small but noticeable stimulation of EGFR autophosphorylation. In solubilized membrane preparations, ascites fluid and EGF produced a comparable stimulation of phosphorylation both of the receptor and of pp35. The effect of EGF and ascites fluid appeared to be additive. Since EGFR1 does not inhibit the binding of EGF to the receptor, the data suggest that the antibody stimulates the receptor kinase activity by interacting with a site distinct from the EGF-binding site; the antibody effect is unmasked by receptor solubilization.

  6. Lactam formation increases receptor binding, adenylyl cyclase stimulation and bone growth stimulation by human parathyroid hormone (hPTH)(1-28)NH2.

    PubMed

    Whitfield, J F; Morley, P; Willick, G E; Isaacs, R J; MacLean, S; Ross, V; Barbier, J R; Divieti, P; Bringhurst, F R

    2000-05-01

    Human parathyroid hormone (1-28)NH2 [hPTH(1-28)NH2] is the smallest of the PTH fragments that can fully stimulate adenylyl cyclase in ROS 17/2 rat osteoblast-like osteosarcoma cells. This fragment has an IC50 of 110 nM for displacing 125I-[Nle8,18,Tyr34]bovine PTH(1-34)NH2 from HKRK B7 porcine kidney cells, which stably express 950,000 human type 1 PTH/PTH-related protein (PTHrP) receptors (PTH1Rs) per cell. It also has an EC50 of 23.9 nM for stimulating adenylyl cyclase in ROS 17/2 cells. Increasing the amphiphilicity of the alpha-helix in the residue 17-28 region by replacing Lys27 with Leu and stabilizing the helix by forming a lactam between Glu22 and Lys26 to produce the [Leu27]cyclo(Glu22-Lys26)hPTH(1-28)NH2 analog dramatically reduced the IC50 for displacing 125I-[Nle8,18,Tyr34]bPTH(1-34)NH2 from hPTH1Rs from 110 to 6 nM and dropped the EC50 for adenylyl cyclase stimulation in ROS 17/2 cells from 23.9 to 9.6 nM. These modifications also increased the osteogenic potency of hPTH(1-28)NH2. Thus, hPTH(1-28)NH2 did not significantly stimulate either femoral or vertebral trabecular bone growth in rats when injected daily at a dose of 5 nmol/100 g body weight for 6 weeks, beginning 2 weeks after ovariectomy (OVX), but it strongly stimulated the growth of trabeculae in the cancellous bone of the distal femurs and L5 vertebrae when injected at 25 nmol/100 g body weight. By contrast [Leu27]cyclo(Glu22-Lys26)hPTH(1-28)NH2 significantly stimulated trabecular bone growth when injected at 5 nmol/100 g of body weight. Thus, these modifications have brought the bone anabolic potency of hPTH(1-28)NH2 considerably closer to the potencies of the larger PTH peptides and analogs.

  7. Toll-Like Receptor 2 Stimulation of Osteoblasts Mediates Staphylococcus Aureus Induced Bone Resorption and Osteoclastogenesis through Enhanced RANKL

    PubMed Central

    Kassem, Ali; Lindholm, Catharina; Lerner, Ulf H

    2016-01-01

    Severe Staphylococcus aureus (S. aureus) infections pose an immense threat to population health and constitute a great burden for the health care worldwide. Inter alia, S. aureus septic arthritis is a disease with high mortality and morbidity caused by destruction of the infected joints and systemic bone loss, osteoporosis. Toll-Like receptors (TLRs) are innate immune cell receptors recognizing a variety of microbial molecules and structures. S. aureus recognition via TLR2 initiates a signaling cascade resulting in production of various cytokines, but the mechanisms by which S. aureus causes rapid and excessive bone loss are still unclear. We, therefore, investigated how S. aureus regulates periosteal/endosteal osteoclast formation and bone resorption. S. aureus stimulation of neonatal mouse parietal bone induced ex vivo bone resorption and osteoclastic gene expression. This effect was associated with increased mRNA and protein expression of receptor activator of NF-kB ligand (RANKL) without significant change in osteoprotegerin (OPG) expression. Bone resorption induced by S. aureus was abolished by OPG. S. aureus increased the expression of osteoclastogenic cytokines and prostaglandins in the parietal bones but the stimulatory effect of S. aureus on bone resorption and Tnfsf11 mRNA expression was independent of these cytokines and prostaglandins. Stimulation of isolated periosteal osteoblasts with S. aureus also resulted in increased expression of Tnfsf11 mRNA, an effect lost in osteoblasts from Tlr2 knockout mice. S. aureus stimulated osteoclastogenesis in isolated periosteal cells without affecting RANKL-stimulated resorption. In contrast, S. aureus inhibited RANKL-induced osteoclast formation in bone marrow macrophages. These data show that S. aureus enhances bone resorption and periosteal osteoclast formation by increasing osteoblast RANKL production through TLR2. Our study indicates the importance of using different in vitro approaches for studies of how S

  8. Stimulation of the B-cell receptor activates the JAK2/STAT3 signaling pathway in chronic lymphocytic leukemia cells

    PubMed Central

    Rozovski, Uri; Wu, Ji Yuan; Harris, David M.; Liu, Zhiming; Li, Ping; Hazan-Halevy, Inbal; Ferrajoli, Alessandra; Burger, Jan A.; O’Brien, Susan; Jain, Nitin; Verstovsek, Srdan; Wierda, William G.; Keating, Michael J.

    2014-01-01

    In chronic lymphocytic leukemia (CLL), stimulation of the B-cell receptor (BCR) triggers survival signals. Because in various cells activation of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway provides cells with survival advantage, we wondered whether BCR stimulation activates the JAK/STAT pathway in CLL cells. To stimulate the BCR we incubated CLL cells with anti-IgM antibodies. Anti-IgM antibodies induced transient tyrosine phosphorylation and nuclear localization of phosphorylated (p) STAT3. Immunoprecipitation studies revealed that anti-JAK2 antibodies coimmunoprecipitated pSTAT3 and pJAK2 in IgM-stimulated but not unstimulated CLL cells, suggesting that activation of the BCR induces activation of JAK2, which phosphorylates STAT3. Incubation of CLL cells with the JAK1/2 inhibitor ruxolitinib inhibited IgM-induced STAT3 phosphorylation and induced apoptosis of IgM-stimulated but not unstimulated CLL cells in a dose- and time-dependent manner. Whether ruxolitinib treatment would benefit patients with CLL remains to be determined. PMID:24778152

  9. Stimulation of the B-cell receptor activates the JAK2/STAT3 signaling pathway in chronic lymphocytic leukemia cells.

    PubMed

    Rozovski, Uri; Wu, Ji Yuan; Harris, David M; Liu, Zhiming; Li, Ping; Hazan-Halevy, Inbal; Ferrajoli, Alessandra; Burger, Jan A; O'Brien, Susan; Jain, Nitin; Verstovsek, Srdan; Wierda, William G; Keating, Michael J; Estrov, Zeev

    2014-06-12

    In chronic lymphocytic leukemia (CLL), stimulation of the B-cell receptor (BCR) triggers survival signals. Because in various cells activation of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway provides cells with survival advantage, we wondered whether BCR stimulation activates the JAK/STAT pathway in CLL cells. To stimulate the BCR we incubated CLL cells with anti-IgM antibodies. Anti-IgM antibodies induced transient tyrosine phosphorylation and nuclear localization of phosphorylated (p) STAT3. Immunoprecipitation studies revealed that anti-JAK2 antibodies coimmunoprecipitated pSTAT3 and pJAK2 in IgM-stimulated but not unstimulated CLL cells, suggesting that activation of the BCR induces activation of JAK2, which phosphorylates STAT3. Incubation of CLL cells with the JAK1/2 inhibitor ruxolitinib inhibited IgM-induced STAT3 phosphorylation and induced apoptosis of IgM-stimulated but not unstimulated CLL cells in a dose- and time-dependent manner. Whether ruxolitinib treatment would benefit patients with CLL remains to be determined.

  10. Stimulation of fat storage by prostacyclin and selective agonists of prostanoid IP receptor during the maturation phase of cultured adipocytes.

    PubMed

    Khan, Ferdous; Syeda, Pinky Karim; Nartey, Michael Nii N; Rahman, Mohammad Shahidur; Islam, Mohammad Safiqul; Nishimura, Kohji; Jisaka, Mitsuo; Shono, Fumia