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Sample records for opioid receptor binding

  1. Mu opioid receptor binding sites in human brain

    International Nuclear Information System (INIS)

    Pilapil, C.; Welner, S.; Magnan, J.; Zamir, N.; Quirion, R.

    1986-01-01

    Our experiments focused on the examination of the distribution of mu opioid receptor binding sites in normal human brain using the highly selective ligand [ 3 H]DAGO, in both membrane binding assay and in vitro receptor autoradiography. Mu opioid binding sites are very discretely distributed in human brain with high densities of sites found in the posterior amygdala, caudate, putamen, hypothalamus and certain cortical areas. Moreover the autoradiographic distribution of [ 3 H]DAGO binding sites clearly reveals the discrete lamination (layers I and III-IV) of mu sites in cortical areas

  2. Adrenergic Agonists Bind to Adrenergic-Receptor-Like Regions of the Mu Opioid Receptor, Enhancing Morphine and Methionine-Enkephalin Binding: A New Approach to "Biased Opioids"?

    Science.gov (United States)

    Root-Bernstein, Robert; Turke, Miah; Subhramanyam, Udaya K Tiruttani; Churchill, Beth; Labahn, Joerg

    2018-01-17

    Extensive evidence demonstrates functional interactions between the adrenergic and opioid systems in a diversity of tissues and organs. While some effects are due to receptor and second messenger cross-talk, recent research has revealed an extracellular, allosteric opioid binding site on adrenergic receptors that enhances adrenergic activity and its duration. The present research addresses whether opioid receptors may have an equivalent extracellular, allosteric adrenergic binding site that has similar enhancing effects on opioid binding. Comparison of adrenergic and opioid receptor sequences revealed that these receptors share very significant regions of similarity, particularly in some of the extracellular and transmembrane regions associated with adrenergic binding in the adrenergic receptors. Five of these shared regions from the mu opioid receptor (muOPR) were synthesized as peptides and tested for binding to adrenergic, opioid and control compounds using ultraviolet spectroscopy. Adrenergic compounds bound to several of these muOPR peptides with low micromolar affinity while acetylcholine, histamine and various adrenergic antagonists did not. Similar studies were then conducted with purified, intact muOPR with similar results. Combinations of epinephrine with methionine enkephalin or morphine increased the binding of both by about half a log unit. These results suggest that muOPR may be allosterically enhanced by adrenergic agonists.

  3. Binding-site analysis of opioid receptors using monoclonal anti-idiotypic antibodies

    International Nuclear Information System (INIS)

    Conroy, W.G.

    1988-01-01

    Structural relatedness between the variable region of anti-ligand antibodies and opioid binding sites allowed the generation of anti-idiotypic antibodies which recognized opioid receptors. The IgG 3 k antibodies which bound to opioid receptors were obtained when an anti-morphine antiserum was the idiotype. Both antibodies bound to opioid receptors, but only one of these blocked the binding of [ 3 H]naloxone. The antibody which did not inhibit the binding of [ 3 H]naloxone was itself displaced from the receptor by opioid ligands. The unique binding properties displayed by this antibody indicated that anti-idiotypic antibodies are not always a perfect image of the original ligand, and therefore may be more useful than typical ligands as probes for the receptor. An auto-anti-idiotypic technique was successfully used to obtain anti-opioid receptor antibodies. Another IgG 3 k antibody that blocked the binding of [ 3 H]naloxone to rat brain opioid receptors was obtained when a mouse was immunized with naloxone conjugated to bovine serum albumin. These data confirmed that an idiotype-anti-idiotype network which can generate an anti-receptor antibody normally functions when an opioid ligand is introduced into an animal in an immunogenic form

  4. Mu receptor binding of some commonly used opioids and their metabolites

    International Nuclear Information System (INIS)

    Chen, Zhaorong; Irvine, R.J.; Somogyi, A.A.; Bochner, F.

    1991-01-01

    The binding affinity to the μ receptor of some opioids chemically related to morphine and some of their metabolites was examined in rat brain homogenates with 3 H-DAMGO. The chemical group at position 6 of the molecule had little effect on binding. Decreasing the length of the alkyl group at position 3 decreased the K i values (morphine < codeine < ethylmorphine < pholcodine). Analgesics with high clinical potency containing a methoxyl group at position 3 had relatively weak receptor binding, while their O-demethylated metabolites had much stronger binding. Many opioids may exert their pharmacological actions predominantly through metabolites

  5. Mu receptor binding of some commonly used opioids and their metabolites

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zhaorong; Irvine, R.J. (Univ. of Adelaide (Australia)); Somogyi, A.A.; Bochner, F. (Univ. of Adelaide (Australia) Royal Adelaide Hospital (Australia))

    1991-01-01

    The binding affinity to the {mu} receptor of some opioids chemically related to morphine and some of their metabolites was examined in rat brain homogenates with {sup 3}H-DAMGO. The chemical group at position 6 of the molecule had little effect on binding. Decreasing the length of the alkyl group at position 3 decreased the K{sub i} values (morphine < codeine < ethylmorphine < pholcodine). Analgesics with high clinical potency containing a methoxyl group at position 3 had relatively weak receptor binding, while their O-demethylated metabolites had much stronger binding. Many opioids may exert their pharmacological actions predominantly through metabolites.

  6. Cholinergic, opioid and glycine receptor binding sites localized in human spinal cord by in vitro autoradiography

    International Nuclear Information System (INIS)

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

    1985-01-01

    Binding sites for the receptor ligands 3 H-quinuclidinylbenzilate, 3 H-alpha-bungarotoxin ( 3 H-alpha-Btx), 3 H-etorphine and 3 H-strychnine were localized autoradiographically at cervical, thoracic and lumbar levels of spinal cords from post-mortem human control subjects and subjects with amyotrophic lateral sclerosis (ALS). The highest densities of muscarinic binding sites were found in the motor neuron areas and in the substantia gelatinosa, while the grey matter binding was very low within Clarke's column. Both 3 H-alpha-Btx and opioid receptor binding sites were numerous within the substantia gelatinosa, while glycine receptor binding sites were more uniformly distribute within the spinal grey matter. In ALS cases, muscarinic receptor binding sites were markedly reduced in motor neuron areas and slightly reduced in the dorsal horn, while the other binding sites studied were relatively unchanged. (author)

  7. Receptor binding properties and antinociceptive effects of chimeric peptides consisting of a micro-opioid receptor agonist and an ORL1 receptor antagonist.

    Science.gov (United States)

    Kawano, Susumu; Ito, Risa; Nishiyama, Miharu; Kubo, Mai; Matsushima, Tomoko; Minamisawa, Motoko; Ambo, Akihiro; Sasaki, Yusuke

    2007-07-01

    Receptor binding properties and antinociceptive activities of chimeric peptides linked by spacers were investigated. The peptides consisted of the micro-opioid receptor ligand dermorphin (Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH(2)) or its analog YRFB (Tyr-D-Arg-Phe-betaAla-NH(2)) linked to the ORL1 receptor ligand Ac-Arg-Tyr-Tyr-Arg-Ile-Lys-NH(2) (Ac-RYYRIK-NH(2)). All chimeric peptides were found to possess high receptor binding affinities for both micro-opioid and ORL1 receptors in mouse brain membranes although their binding affinities for both receptors in spinal membranes were significantly lower. Among them, chimeric peptide 2, which consists of dermorphin and Ac-RYYRIK-NH(2) connected by a long spacer, had the highest binding affinity towards both receptors. In the tail-flick test following intrathecal (i.t.) administration to mice, all chimeric peptides showed potent and dose-dependent antinociceptive activities with an ED(50) of 1.34-4.51 (pmol/mouse), nearly comparable to dermorphin alone (ED(50); 1.08 pmol/mouse). In contrast to their micro-opioid receptor binding profiles, intracerebroventricular (i.c.v.) administration of the chimeric peptides resulted in much less potent antinociceptive activity (ED(50) 5.55-100peptides, and the regulation of mu-opioid receptor-mediated antinociception in brain. The present chimeric peptides may be useful as pharmacological tools for studies on micro-opioid receptor/ORL1 receptor heterodimers.

  8. Site-directed alkylation of multiple opioid receptors. I. Binding selectivity

    International Nuclear Information System (INIS)

    James, I.F.; Goldstein, A.

    1984-01-01

    A method for measuring and expressing the binding selectivity of ligands for mu, delta, and kappa opioid binding sites is reported. Radioligands are used that are partially selective for these sites in combination with membrane preparations enriched in each site. Enrichment was obtained by treatment of membranes with the alkylating agent beta-chlornaltrexamine in the presence of appropriate protecting ligands. After enrichment for mu receptors, [ 3 H] dihydromorphine bound to a single type of site as judged by the slope of competition binding curves. After enrichment for delta or kappa receptors, binding sites for [ 3 H] [D-Ala2, D-Leu5]enkephalin and [3H]ethylketocyclazocine, respectively, were still not homogeneous. There were residual mu sites in delta-enriched membranes but no evidence for residual mu or delta sites in kappa-enriched membranes were found. This method was used to identify ligands that are highly selective for each of the three types of sites

  9. Affinity of the enantiomers of. alpha. - and. beta. -cyclazocine for binding to the phencyclidine and. mu. opioid receptors

    Energy Technology Data Exchange (ETDEWEB)

    Todd, S.L.; Balster, R.L.; Martin, B.R. (Virginia Commonwealth Univ., Richmond (USA))

    1990-01-01

    The enantiomers in the {alpha} and {beta} series of cyclazocine were evaluated for their ability to bind to phencyclidine (PCP) and {mu}-opioid receptors in order to determine their receptor selectivity. The affinity of (-)-{beta}-cyclazocine for the PCP receptor was 1.5 greater than PCP itself. In contrast, (-)-{alpha}-cyclazocine, (+)-{alpha}-cyclazocine, and (+)-{beta}-cyclazocine were 3-, 5- and 138-fold less potent than PCP, respectively. Scatchard analysis of saturable binding of ({sup 3}H)Tyr-D-Ala-Gly-N-MePhe-Gly-ol (DAMGO) also exhibited a homogeneous population of binding sites with an apparent K{sub D} of 1.9 nM and an estimated Bmax of 117 pM. (3H)Tyr-D-Ala-Gly-N-MePhe-Gly-ol (DAMGO) binding studies revealed that (-)-{alpha}-cyclazocine (K{sub D} = 0.48 nM) was 31-, 1020- and 12,600-fold more potent than (-)-{beta}-cyclazocine, (+)-{alpha}-cyclazocine and (+)-{beta}-cyclazocine, respectively, for binding to the {mu}-opioid receptor. These data show that, although (-)-{beta}-cyclazocine is a potent PCP receptor ligand consistent with its potent PCP-like discriminative stimulus effects, it shows little selectivity for PCP receptor since it also potently displaces {mu}-opioid binding. However, these cyclazocine isomers, due to their extraordinary degree of stereoselectivity, may be useful in characterizing the structural requirements for benzomorphans having activity at the PCP receptor.

  10. Structural Determinants for the Binding of Morphinan Agonists to the μ-Opioid Receptor.

    Directory of Open Access Journals (Sweden)

    Xiaojing Cong

    Full Text Available Atomistic descriptions of the μ-opioid receptor (μOR noncovalently binding with two of its prototypical morphinan agonists, morphine (MOP and hydromorphone (HMP, are investigated using molecular dynamics (MD simulations. Subtle differences between the binding modes and hydration properties of MOP and HMP emerge from the calculations. Alchemical free energy perturbation calculations show qualitative agreement with in vitro experiments performed in this work: indeed, the binding free energy difference between MOP and HMP computed by forward and backward alchemical transformation is 1.2±1.1 and 0.8±0.8 kcal/mol, respectively, to be compared with 0.4±0.3 kcal/mol from experiment. Comparison with an MD simulation of μOR covalently bound with the antagonist β-funaltrexamine hints to agonist-induced conformational changes associated with an early event of the receptor's activation: a shift of the transmembrane helix 6 relative to the transmembrane helix 3 and a consequent loss of the key R165-T279 interhelical hydrogen bond. This finding is consistent with a previous proposal suggesting that the R165-T279 hydrogen bond between these two helices indicates an inactive receptor conformation.

  11. [3H]naloxone as an opioid receptor label: Analysis of binding site heterogeneity and use for determination of opioid affinities of casomorphin analogues

    International Nuclear Information System (INIS)

    Schnittler, M.; Repke, H.; Liebmann, C.; Schrader, U.; Schulze, H.P.; Neubert, K.

    1990-01-01

    The nonselective antagonist [ 3 H]naloxone was used to identify opioid receptors in rat brain membranes. The multiple naloxone binding sites were related to different opioid receptors by means of selective opiod ligands as well as various β-casomorphin analogues. Analysis of binding site heterogeneity was performed using several computer curve fitting methods. The results indicate that structurally modified casomorphin peptides are able to discriminate between μ 1 and μ 2 binding sites. The affinities to the μ sites obtained with [ 3 H]naloxone as label are in a good agreement with those from experiments with the μ selective radioligand [ 3 H]DAGO. The μ 1 site affinities of these casomorphin derivatives are well correlated with their antinociceptive potencies. This finding suggests the mediation of the analgesic activity via the high-affinity μ 1 subtype. (author)

  12. Adrenergic Agonists Bind to Adrenergic-Receptor-Like Regions of the Mu Opioid Receptor, Enhancing Morphine and Methionine-Enkephalin Binding: A New Approach to “Biased Opioids”?

    Science.gov (United States)

    Turke, Miah; Subhramanyam, Udaya K. Tiruttani; Churchill, Beth; Labahn, Joerg

    2018-01-01

    Extensive evidence demonstrates functional interactions between the adrenergic and opioid systems in a diversity of tissues and organs. While some effects are due to receptor and second messenger cross-talk, recent research has revealed an extracellular, allosteric opioid binding site on adrenergic receptors that enhances adrenergic activity and its duration. The present research addresses whether opioid receptors may have an equivalent extracellular, allosteric adrenergic binding site that has similar enhancing effects on opioid binding. Comparison of adrenergic and opioid receptor sequences revealed that these receptors share very significant regions of similarity, particularly in some of the extracellular and transmembrane regions associated with adrenergic binding in the adrenergic receptors. Five of these shared regions from the mu opioid receptor (muOPR) were synthesized as peptides and tested for binding to adrenergic, opioid and control compounds using ultraviolet spectroscopy. Adrenergic compounds bound to several of these muOPR peptides with low micromolar affinity while acetylcholine, histamine and various adrenergic antagonists did not. Similar studies were then conducted with purified, intact muOPR with similar results. Combinations of epinephrine with methionine enkephalin or morphine increased the binding of both by about half a log unit. These results suggest that muOPR may be allosterically enhanced by adrenergic agonists. PMID:29342106

  13. Adrenergic Agonists Bind to Adrenergic-Receptor-Like Regions of the Mu Opioid Receptor, Enhancing Morphine and Methionine-Enkephalin Binding: A New Approach to “Biased Opioids”?

    Directory of Open Access Journals (Sweden)

    Robert Root-Bernstein

    2018-01-01

    Full Text Available Extensive evidence demonstrates functional interactions between the adrenergic and opioid systems in a diversity of tissues and organs. While some effects are due to receptor and second messenger cross-talk, recent research has revealed an extracellular, allosteric opioid binding site on adrenergic receptors that enhances adrenergic activity and its duration. The present research addresses whether opioid receptors may have an equivalent extracellular, allosteric adrenergic binding site that has similar enhancing effects on opioid binding. Comparison of adrenergic and opioid receptor sequences revealed that these receptors share very significant regions of similarity, particularly in some of the extracellular and transmembrane regions associated with adrenergic binding in the adrenergic receptors. Five of these shared regions from the mu opioid receptor (muOPR were synthesized as peptides and tested for binding to adrenergic, opioid and control compounds using ultraviolet spectroscopy. Adrenergic compounds bound to several of these muOPR peptides with low micromolar affinity while acetylcholine, histamine and various adrenergic antagonists did not. Similar studies were then conducted with purified, intact muOPR with similar results. Combinations of epinephrine with methionine enkephalin or morphine increased the binding of both by about half a log unit. These results suggest that muOPR may be allosterically enhanced by adrenergic agonists.

  14. Human Mu Opioid Receptor (OPRM1A118G) polymorphism is associated with brain mu- opioid receptor binding potential in smokers

    Energy Technology Data Exchange (ETDEWEB)

    Ray, R.; Logan, J.; Ray, R.; Ruparel, K.; Newberg, A.; Wileyto, E.P.; Loughead, J.W.; Divgi, C.; Blendy, J.A.; Logan, J.; Zubieta, J.-K.; Lerman, C.

    2011-04-15

    Evidence points to the endogenous opioid system, and the mu-opioid receptor (MOR) in particular, in mediating the rewarding effects of drugs of abuse, including nicotine. A single nucleotide polymorphism (SNP) in the human MOR gene (OPRM1 A118G) has been shown to alter receptor protein level in preclinical models and smoking behavior in humans. To clarify the underlying mechanisms for these associations, we conducted an in vivo investigation of the effects of OPRM1 A118G genotype on MOR binding potential (BP{sub ND} or receptor availability). Twenty-two smokers prescreened for genotype (12 A/A, 10 */G) completed two [{sup 11}C] carfentanil positron emission tomography (PET) imaging sessions following overnight abstinence and exposure to a nicotine-containing cigarette and a denicotinized cigarette. Independent of session, smokers homozygous for the wild-type OPRM1 A allele exhibited significantly higher levels of MOR BP{sub ND} than smokers carrying the G allele in bilateral amygdala, left thalamus, and left anterior cingulate cortex. Among G allele carriers, the extent of subjective reward difference (denicotinized versus nicotine cigarette) was associated significantly with MOR BP{sub ND} difference in right amygdala, caudate, anterior cingulate cortex, and thalamus. Future translational investigations can elucidate the role of MORs in nicotine addiction, which may lead to development of novel therapeutics.

  15. Human Mu Opioid Receptor (OPRM1A118G) polymorphism is associated with brain mu- opioid receptor binding potential in smokers

    International Nuclear Information System (INIS)

    Ray, R.; Logan, J.; Ruparel, K.; Newberg, A.; Wileyto, E.P.; Loughead, J.W.; Divgi, C.; Blendy, J.A.; Logan, J.; Zubieta, J.-K.; Lerman, C.

    2011-01-01

    Evidence points to the endogenous opioid system, and the mu-opioid receptor (MOR) in particular, in mediating the rewarding effects of drugs of abuse, including nicotine. A single nucleotide polymorphism (SNP) in the human MOR gene (OPRM1 A118G) has been shown to alter receptor protein level in preclinical models and smoking behavior in humans. To clarify the underlying mechanisms for these associations, we conducted an in vivo investigation of the effects of OPRM1 A118G genotype on MOR binding potential (BP ND or receptor availability). Twenty-two smokers prescreened for genotype (12 A/A, 10 */G) completed two [ 11 C] carfentanil positron emission tomography (PET) imaging sessions following overnight abstinence and exposure to a nicotine-containing cigarette and a denicotinized cigarette. Independent of session, smokers homozygous for the wild-type OPRM1 A allele exhibited significantly higher levels of MOR BP ND than smokers carrying the G allele in bilateral amygdala, left thalamus, and left anterior cingulate cortex. Among G allele carriers, the extent of subjective reward difference (denicotinized versus nicotine cigarette) was associated significantly with MOR BP ND difference in right amygdala, caudate, anterior cingulate cortex, and thalamus. Future translational investigations can elucidate the role of MORs in nicotine addiction, which may lead to development of novel therapeutics.

  16. Molecular characterization of opioid receptors

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    Howard, A.D.

    1986-01-01

    The aim of this research was to purify and characterize active opioid receptors and elucidate molecular aspects of opioid receptor heterogeneity. Purification to apparent homogeneity of an opioid binding protein from bovine caudate was achieved by solubilization in the non-ionic detergent, digitonin, followed by sequential chromatography on the opiate affinity matrix, ..beta..-naltrexylethylenediamine-CH-Sepharose 4B, and on the lectine affinity matrix, wheat germ agglutinin-agarose. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE) followed by autoradiography revealed that radioiodinated purified receptor gave a single band. Purified receptor preparations showed a specific activity of 12,000-15,000 fmol of opiate bound per mg of protein. Radioiodinated human beta-endorphin (/sup 125/I-beta-end/sub H/) was used as a probe to investigate the ligand binding subunits of mu and delta opioid receptors. /sup 125/I-beta-end/sub H/ was shown to bind to a variety of opioid receptor-containing tissues with high affinity and specificity with preference for mu and delta sites, and with little, if any, binding to kappa sites. Affinity crosslinking techniques were employed to covalently link /sup 125/I-beta-end/sub H/ to opioid receptors, utilizing derivatives of bis-succinimidyl esters that are bifunctional crosslinkers with specificities for amino and sulfhydryl groups. This, and competition experiments with high type-selective ligands, permitted the assignment of two labeled peptides to their receptor types, namely a peptide of M/sub r/ = 65,000 for mu receptors and one of M/sub r/ = 53,000 for delta receptors.

  17. The potent opioid agonist, (+)-cis-3-methylfentanyl binds pseudoirreversibly to the opioid receptor complex in vitro and in vivo: Evidence for a novel mechanism of action

    Energy Technology Data Exchange (ETDEWEB)

    Band, L.; Xu, Heng; Bykov, V.; Rothman, R.B.; Kim, Chongho; Newman, A.; Jacobson, A.E.; Rice, K.C. (NIDDK, Bethesda, MD (USA)); Greig, N. (NIA, Bethesda, MD (USA))

    1990-01-01

    The present study demonstrates that pretreatment of rat brain membranes with (+)-cis-3-methylfentanyl ((+)-cis-MF), followed by extensive washing of the membranes, produces a wash-resistant decreasing in the binding of ({sup 3}H)-(D-ala{sup 2}, D-leu{sup 5})enkephalin to the d binding site of the opioid receptor complex ({delta}{sub cx} binding site). Intravenous administration of (+)-cis-MF (50 {mu}g/kg) to rats produced a pronounced catalepsy and also produced a wash-resistant masking of {delta}{sub cx} and {mu} binding sites in membranes prepared 120 min post-injection. Administration of 1 mg/kg i.v. of the opioid antagonist, 6-desoxy-6{beta}-fluoronaltrexone (cycloFOXY), 100 min after the injection of (+)-cis-MF (20 min prior to the preparation of membranes) completely reversed the catatonia and restored masked {delta}{sub cx} binding sites to control levels. This was not observed with (+)-cycloFOXY. The implications of these and other findings for the mechanism of action of (+)-cis-MF and models of the opioid receptors are discussed.

  18. Chimeric opioid peptides: Tools for identifying opioid receptor types

    International Nuclear Information System (INIS)

    Xie, G.; Miyajima, A.; Yokota, T.; Arai, K.; Goldstein, A.

    1990-01-01

    The authors synthesized several chimeric [125J-labelled] peptides in which the N-terminal nine residues of dynorphin-32, a peptide selective for the κ opioid receptor, were replaced by opioid peptides selective for other opioid receptor types. Each chimeric peptide retained the high affinity and type selectivity characteristic of its N-terminal sequence. The common C-terminal two-thirds of the chimeric peptides served as an epitope recognized by the same monoclonal antibody. When bound to receptors on a cell surface or membrane preparation, these peptides could still bind specifically to the monoclonal antibody. These chimeric peptides should be useful for isolating μ, δ, and κ opioid receptors and for identifying opioid receptors on transfected cells in expression cloning procedures. The general approach using chimeric peptides should be applicable to other peptide receptors

  19. Kappa-receptor selective binding of opioid ligands with a heterocyclic bicyclo[3.3.1]nonan-9-one structure.

    Science.gov (United States)

    Benyhe, S; Márki, A; Nachtsheim, Corina; Holzgrabe, Ulrike; Borsodi, Anna

    2003-01-01

    Previous pharmacological results have suggested that members of the heterocyclic bicyclo[3.3.1]nonan-9-one-like compounds are potent kappa-opioid receptor specific agonists. One lead molecule of this series. called compound 1 (dimethyl 7-methyl-2,4-di-2-pyridyl-3.7-diazabicyclo[3.3.1]nonan-9-one-1,5-dicarboxylate) exhibited high affinity for [3H]ethylketocyclazocine and [3H]U-69.593 binding sites in guinea pig cerebellar membranes which known to be a good source for kappa1 receptors. It was shown by molecular modelling that heterocyclic bicyclo[3.3.1]nonan-9-ones fit very well with the structure of ketazocine, a prototypic kappa-selective benzomorphan compound; when compared to the arylacetamide structure of U-69.593, a specific kappa1-receptor agonist, a similar geometry was found with a slightly different distribution of the charges. It is postulated, that the essential structural skeleton involved in the opioid activity is an aryl-propyl-amine element distributed along the N7-C6-C5-C4-aryl bonds.

  20. Kinetic analysis of transport and opioid receptor binding of ( sup 3 H)(-)-cyclofoxy in rat brain in vivo: Implications for human studies

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    Sawada, Y.; Kawai, R.; McManaway, M.; Otsuki, H.; Rice, K.C.; Patlak, C.S.; Blasberg, R.G. (National Institutes of Health, Bethesda, MD (USA))

    1991-03-01

    (3H)Cyclofoxy (CF: 17-cyclopropylmethyl-3,14-dihydroxy-4,5-alpha-epoxy-6-beta-fluoromorp hinan) is an opioid antagonist with affinity to both mu and kappa subtypes that was synthesized for quantitative evaluation of opioid receptor binding in vivo. Two sets of experiments in rats were analyzed. The first involved determining the metabolite-corrected blood concentration and tissue distribution of CF in brain 1 to 60 min after i.v. bolus injection. The second involved measuring brain washout for 15 to 120 s following intracarotid artery injection of CF. A physiologically based model and a classical compartmental pharmacokinetic model were compared. The models included different assumptions for transport across the blood-brain barrier (BBB); estimates of nonspecific tissue binding and specific binding to a single opiate receptor site were found to be essentially the same with both models. The nonspecific binding equilibrium constant varied modestly in different brain structures (Keq = 3-9), whereas the binding potential (BP) varied over a much broader range (BP = 0.6-32). In vivo estimates of the opioid receptor dissociation constant were similar for different brain structures (KD = 2.1-5.2 nM), whereas the apparent receptor density (Bmax) varied between 1 (cerebellum) and 78 (thalamus) pmol/g of brain. The receptor dissociation rate constants in cerebrum (k4 = 0.08-0.16 min-1; koff = 0.16-0.23 min-1) and brain vascular permeability (PS = 1.3-3.4 ml/min/g) are sufficiently high to achieve equilibrium conditions within a reasonable period of time. Graphical analysis of the data is inappropriate due to the high tissue-loss rate constant for CF in brain. From these findings, CF should be a very useful opioid receptor ligand for the estimation of the receptor binding parameters in human subjects using (18F)CF and positron emission tomography.

  1. Kinetic analysis of transport and opioid receptor binding of [3H](-)-cyclofoxy in rat brain in vivo: Implications for human studies

    International Nuclear Information System (INIS)

    Sawada, Y.; Kawai, R.; McManaway, M.; Otsuki, H.; Rice, K.C.; Patlak, C.S.; Blasberg, R.G.

    1991-01-01

    [3H]Cyclofoxy (CF: 17-cyclopropylmethyl-3,14-dihydroxy-4,5-alpha-epoxy-6-beta-fluoromorp hinan) is an opioid antagonist with affinity to both mu and kappa subtypes that was synthesized for quantitative evaluation of opioid receptor binding in vivo. Two sets of experiments in rats were analyzed. The first involved determining the metabolite-corrected blood concentration and tissue distribution of CF in brain 1 to 60 min after i.v. bolus injection. The second involved measuring brain washout for 15 to 120 s following intracarotid artery injection of CF. A physiologically based model and a classical compartmental pharmacokinetic model were compared. The models included different assumptions for transport across the blood-brain barrier (BBB); estimates of nonspecific tissue binding and specific binding to a single opiate receptor site were found to be essentially the same with both models. The nonspecific binding equilibrium constant varied modestly in different brain structures (Keq = 3-9), whereas the binding potential (BP) varied over a much broader range (BP = 0.6-32). In vivo estimates of the opioid receptor dissociation constant were similar for different brain structures (KD = 2.1-5.2 nM), whereas the apparent receptor density (Bmax) varied between 1 (cerebellum) and 78 (thalamus) pmol/g of brain. The receptor dissociation rate constants in cerebrum (k4 = 0.08-0.16 min-1; koff = 0.16-0.23 min-1) and brain vascular permeability (PS = 1.3-3.4 ml/min/g) are sufficiently high to achieve equilibrium conditions within a reasonable period of time. Graphical analysis of the data is inappropriate due to the high tissue-loss rate constant for CF in brain. From these findings, CF should be a very useful opioid receptor ligand for the estimation of the receptor binding parameters in human subjects using [18F]CF and positron emission tomography

  2. Identification in the mu-opioid receptor of cysteine residues responsible for inactivation of ligand binding by thiol alkylating and reducing agents.

    Science.gov (United States)

    Gaibelet, G; Capeyrou, R; Dietrich, G; Emorine, L J

    1997-05-19

    Inactivation by thiol reducing and alkylating agents of ligand binding to the human mu-opioid receptor was examined. Dithiothreitol reduced the number of [3H]diprenorphine binding sites. Replacement by seryl residues of either C142 or C219 in extracellular loops 1 and 2 of the mu receptor resulted in a complete loss of opioid binding. A disulfide bound linking C142 to C219 may thus be essential to maintain a functional conformation of the receptor. We also demonstrated that inactivation of ligand binding upon alkylation by N-ethylmaleimide occurred at two sites. Alteration of the more sensitive (IC50 = 20 microM) did not modify antagonists binding but decreased agonist affinity almost 10-fold. Modification of the less reactive site (IC50 = 2 mM) decreased the number of both agonist and antagonist binding sites. The alkylation site of higher sensitivity to N-ethylmaleimide was shown by mutagenesis experiments to be constituted of both C81 and C332 in transmembrane domains 1 and 7 of the mu-opioid receptor.

  3. Opioid receptors in human neuroblastoma SH-SY5Y cells: evidence for distinct morphine (. mu. ) and enkephalin (delta) binding sites

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    Kazmi, S.M.I.; Mishra, R.K.

    1986-06-13

    Human neuroblastoma SH-SY5Y cells exhibited a heterogeneous population of ..mu.. and delta types of opioid binding sites. These specific binding sites displayed the characteristic saturability, stereospecificity and reversibility, expected of a receptor. Scatchard analysis of (/sup 3/H)-D-Ala/sup 2/-D-Leu/sup 5/-enkephalin (DADLE) in the presence of 10/sup -5/M D-Pro/sup 4/-morphiceptin (to block the ..mu.. receptors) and the competitive displacement by various highly selective ligands yielded the binding parameters of delta sites which closely resemble those of the delta receptors in brain and mouse neuroblastoma clones. Similarly, the high affinity binding of (/sup 3/H)-dihydromorphine, together with the higher potency of morphine analogues to displace (/sup 3/H)-naloxone binding established the presence of ..mu.. sites. Guanine nucleotides and NaCl significantly inhibited the association and increased the dissociation of (/sup 3/H)-DADLE binding.

  4. Interleukin-1 interaction with neuroregulatory systems: selective enhancement by recombinant human and mouse interleukin-1 of in vitro opioid peptide receptor binding in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Wiedermann, C.J.

    1989-02-01

    Interleukin-1 (IL-1) exerts a wide variety of biological effects on various cell types and may be regarded as a pleiotropic peptide hormone. Biological evidence suggests that IL-1 participates in the modulation of central nervous system physiology and behavior in a fashion characteristic of neuroendocrine hormones. In this investigation, recombinant (r) human (h) IL-1 and r mouse (m) IL-1 were examined for their modulation of opioid peptide receptor binding in vitro. Experiments were performed on frozen sections of rat brain. Receptor binding of radiolabeled substance P and of radiolabeled neurotensin were not significantly affected by the presence of rIL-1s. Recombinant IL-1s, however, significantly enhanced specific binding of 125I-beta-endorphin (125I-beta-END) and of D-ala2-(tyrosyl-3,5-3H)enkephalin-(5-D-leucine) (3H-D-ALA), equipotently and in a concentration-dependent manner with maximal activity occurring at a concentration of 10 LAF units/ml. The increased binding of 125I-beta-END and 3H-D-ALA was blocked steroselectively by (-)-naloxone and by etorphine, suggesting detection of opiate receptors. In addition, brain distribution patterns of receptors labeled in the presence of rIL-1s corresponded to patterns previously published for opiate receptors. Autoradiographic visualization of receptors revealed that rIL-1s in the different areas of the brain exert their effect on opioid binding with comparable potencies. The data suggest that certain central nervous system effects of IL-1s may be mediated by their selective interaction with opiatergic systems at the receptor level.

  5. Chimeric opioid peptides: tools for identifying opioid receptor types.

    OpenAIRE

    Xie, G X; Miyajima, A; Yokota, T; Arai, K; Goldstein, A

    1990-01-01

    We synthesized several chimeric peptides in which the N-terminal nine residues of dynorphin-32, a peptide selective for the kappa opioid receptor, were replaced by opioid peptides selective for other opioid receptor types. Each chimeric peptide retained the high affinity and type selectivity characteristic of its N-terminal sequence. The common C-terminal two-thirds of the chimeric peptides served as an epitope recognized by the same monoclonal antibody. When bound to receptors on a cell surf...

  6. Naloxone-sensitive, haloperidol-sensitive, [3H](+)SKF-10047-binding protein partially purified from rat liver and rat brain membranes: an opioid/sigma receptor?

    Science.gov (United States)

    Tsao, L I; Su, T P

    1997-02-01

    A naloxone-sensitive, haloperidol-sensitive, [3H](+)SKF-10047-binding protein was partially purified from rat liver and rat brain membranes in an affinity chromatography originally designed to purify sigma receptors. Detergent-solubilized extracts from membranes were adsorbed to Sephadex G-25 resin containing an affinity ligand for sigma receptors: N-(2- 3,4-dichlorophenyl]ethyl)-N-(6-aminohexyl)-(2-[1-pyrrolidinyl]) ethylamine (DAPE). After eluting the resin with haloperidol, a protein that bound [3H](+)SKF-10047 was detected in the eluates. However, the protein was not the sigma receptor. [3H](+)SKF-10047 binding to the protein was inhibited by the following compounds in the order of decreasing potency: (+)pentazocine > (-) pentazocine > (+/-)cyclazocine > (-)morphine > (-)naloxone > haloperidol > (+)SKF-10047 > DADLE > (-)SKF-10047. Further, the prototypic sigma receptor ligands, such as 1,3-di-o-tolylguanidine (DTG), (+)3-PPP, and progesterone, bound poorly to the protein. Tryptic digestion and heat treatment of the affinity-purified protein abolished radioligand binding. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS/PAGE) of the partially-purified protein from the liver revealed a major diffuse band with a molecular mass of 31 kDa, a polypeptide of 65 kDa, and another polypeptide of > 97 kDa. This study demonstrates the existence of a novel protein in the rat liver and rat brain which binds opioids, benzomorphans, and haloperidol with namomolar affinity. The protein resembles the opioid/sigma receptor originally proposed by Martin et al. [(1976): J. Pharmacol. Exp. Ther., 197:517-532.]. A high degree of purification of this protein has been achieved in the present study.

  7. Quantitative analysis of multiple kappa-opioid receptors by selective and nonselective ligand binding in guinea pig spinal cord: Resolution of high and low affinity states of the kappa 2 receptors by a computerized model-fitting technique

    International Nuclear Information System (INIS)

    Tiberi, M.; Magnan, J.

    1990-01-01

    The binding characteristics of selective and nonselective opioids have been studied in whole guinea pig spinal cord, using a computer fitting method to analyze the data obtained from saturation and competition studies. The delineation of specific binding sites labeled by the mu-selective opioid [3H]D-Ala2,MePhe4,Gly-ol5-enkephalin (Kd = 2.58 nM, R = 4.52 pmol/g of tissue) and by the delta-selective opioid [3H]D-Pen2, D-Pen5-enkephalin (Kd = 2.02 nM, R = 1.47 pmol/g of tissue) suggests the presence of mu and delta-receptors in the spinal cord tissue. The presence of kappa receptors was probed by the kappa-selective opioid [3H]U69593 (Kd = 3.31 nM, R = 2.00 pmol/g of tissue). The pharmacological characterization of the sites labeled by [3H]U69593 confirms the assumption that this ligand discriminates kappa receptors in guinea pig spinal cord. The benzomorphan [3H]ethylketazocine labels a population of receptors with one homogeneous affinity state (Kd = 0.65 nM, R = 7.39 pmol/g of tissue). The total binding capacity of this ligand was not different from the sum of the binding capacities of mu, delta-, and kappa-selective ligands. Under mu- and delta-suppressed conditions, [3H]ethylketazocine still binds to receptors with one homogeneous affinity state (Kd = 0.45 nM, R = 1.69 pmol/g of tissue). Competition studies performed against the binding of [3H]ethylketazocine under these experimental conditions reveal that the pharmacological profile of the radiolabeled receptors is similar to the profile of the kappa receptors labeled with [3H]U69593. Saturation studies using the nonselective opioid [3H]bremazocine demonstrate that this ligand binds to spinal cord membranes with heterogeneous affinities (Kd1 = 0.28 nM, R1 = 7.91 pmol/g of tissue; Kd2 = 3.24 nM, R2 = 11.2 pmol/g of tissue)

  8. Quantitative analysis of multiple kappa-opioid receptors by selective and nonselective ligand binding in guinea pig spinal cord: Resolution of high and low affinity states of the kappa 2 receptors by a computerized model-fitting technique

    Energy Technology Data Exchange (ETDEWEB)

    Tiberi, M.; Magnan, J. (Universite de Montreal, Quebec (Canada))

    1990-05-01

    The binding characteristics of selective and nonselective opioids have been studied in whole guinea pig spinal cord, using a computer fitting method to analyze the data obtained from saturation and competition studies. The delineation of specific binding sites labeled by the mu-selective opioid (3H)D-Ala2,MePhe4,Gly-ol5-enkephalin (Kd = 2.58 nM, R = 4.52 pmol/g of tissue) and by the delta-selective opioid (3H)D-Pen2, D-Pen5-enkephalin (Kd = 2.02 nM, R = 1.47 pmol/g of tissue) suggests the presence of mu and delta-receptors in the spinal cord tissue. The presence of kappa receptors was probed by the kappa-selective opioid (3H)U69593 (Kd = 3.31 nM, R = 2.00 pmol/g of tissue). The pharmacological characterization of the sites labeled by (3H)U69593 confirms the assumption that this ligand discriminates kappa receptors in guinea pig spinal cord. The benzomorphan (3H)ethylketazocine labels a population of receptors with one homogeneous affinity state (Kd = 0.65 nM, R = 7.39 pmol/g of tissue). The total binding capacity of this ligand was not different from the sum of the binding capacities of mu, delta-, and kappa-selective ligands. Under mu- and delta-suppressed conditions, (3H)ethylketazocine still binds to receptors with one homogeneous affinity state (Kd = 0.45 nM, R = 1.69 pmol/g of tissue). Competition studies performed against the binding of (3H)ethylketazocine under these experimental conditions reveal that the pharmacological profile of the radiolabeled receptors is similar to the profile of the kappa receptors labeled with (3H)U69593. Saturation studies using the nonselective opioid (3H)bremazocine demonstrate that this ligand binds to spinal cord membranes with heterogeneous affinities (Kd1 = 0.28 nM, R1 = 7.91 pmol/g of tissue; Kd2 = 3.24 nM, R2 = 11.2 pmol/g of tissue).

  9. Comparison of (/sup 125/I)beta-endorphin binding to rat brain and NG108-15 cells using a monoclonal antibody directed against the opioid receptor

    Energy Technology Data Exchange (ETDEWEB)

    Bidlack, J.M.; O' Malley, W.E.; Schulz, R.

    1988-02-01

    The properties of (/sup 125/I)beta h-endorphin-binding sites from rat brain membranes and membranes from the NG108-15 cell line were compared using a monoclonal antibody directed against the opioid receptor and opioid peptides as probes. The binding of (/sup 125/I)beta h-endorphin to both rat brain and NG108-15 membranes yielded linear Scatchard plots with Kd values of 1.2 nM and 1.5 nM, respectively, and Bmax values of 865 fmol/mg rat brain membrane protein and 1077 fmol/mg NG108-15 membrane protein. A monoclonal antibody, OR-689.2.4, capable of inhibiting mu and delta binding but not kappa binding to rat brain membranes, noncompetitively inhibited the binding of 1 nM (/sup 125/I)beta h-endorphin to rat brain and NG108-15 membranes with an IC50 value of 405 nM for rat brain membranes and 543 nM for NG108-15 membranes. The monoclonal antibody also inhibited the binding of 3 nM (/sup 3/H) (D-penicillamine2, D-penicillamine5) enkephalin to NG108-15 membranes with an IC50 value of 370 nM. In addition to blocking the binding of (/sup 125/I)beta h-endorphin to brain membranes, the antibody also displaced (/sup 125/I)beta h-endorphin from membranes. Site-specific opioid peptides had large variations in their IC50 values depending on whether they were inhibiting (/sup 125/I)beta h-endorphin binding to rat brain or the NG108-15 membranes. When the peptides were tested with the monoclonal antibody for their combined ability to inhibit (/sup 125/I)beta h-endorphin binding to both membrane preparations, the peptides and antibody blocked binding as though they were acting at allosterically coupled sites, not two totally independent sites. These studies suggest that mu-, delta-, and beta-endorphin-binding sites share some sequence homology with the 35,000-dalton protein that the antibody is directed against.

  10. Measurement of central μ-opioid receptor binding in vivo with PET and [11C]carfentanil: a test-retest study in healthy subjects

    International Nuclear Information System (INIS)

    Hirvonen, Jussi; Aalto, Sargo; Maksimow, Anu; Oikonen, Vesa; Naagren, Kjell; Hagelberg, Nora; Scheinin, Harry; Ingman, Kimmo; Virkkala, Jussi

    2009-01-01

    [ 11 C]Carfentanil has been widely used in positron emission tomography (PET) studies for measuring μ-opioid receptor binding in humans, but the reproducibility of the binding parameter estimates is unknown. Eight healthy volunteers were scanned twice during the same day with [ 11 C]carfentanil PET, and binding to receptors was assessed with both reference tissue and arterial plasma input-based models using region of interest (ROI) and voxel-based quantification. The two-tissue compartmental model distribution volume (V T ) was highly reproducible as indicated by low variability (VAR 0.93). BP ND (BP relative to the nondisplaceable tissue compartment) was also highly reproducible (VAR 0.90) both at ROI- and voxel-level, and reference tissue-based models provided stable estimates after 40 min. The reproducibility of [ 11 C]carfentanil binding parameter estimates is excellent with outcome measures based on both arterial plasma and reference tissue input, and a scanning time of 40 min appears sufficient. (orig.)

  11. Striatal μ-opioid receptor availability predicts cold pressor pain threshold in healthy human subjects

    DEFF Research Database (Denmark)

    Hagelberg, Nora; Aalto, Sargo; Tuominen, Lauri

    2012-01-01

    the potential associations between μ-opioid receptor BP(ND) and psychophysical measures. The results show that striatal μ-opioid receptor BP(ND) predicts cold pressor pain threshold, but not cold pressor pain tolerance or tactile sensitivity. This finding suggests that striatal μ-opioid receptor density......Previous PET studies in healthy humans have shown that brain μ-opioid receptor activation during experimental pain is associated with reductions in the sensory and affective ratings of the individual pain experience. The aim of this study was to find out whether brain μ-opioid receptor binding...... at the resting state, in absence of painful stimulation, can be a long-term predictor of experimental pain sensitivity. We measured μ-opioid receptor binding potential (BP(ND)) with μ-opioid receptor selective radiotracer [(11)C]carfentanil and positron emission tomography (PET) in 12 healthy male subjects...

  12. Combined autoradiographic-immunocytochemical analysis of opioid receptors and opioid peptide neuronal systems in brain

    Energy Technology Data Exchange (ETDEWEB)

    Lewis, M.E.; Khachaturian, H.; Watson, S.J.

    1985-01-01

    Using adjacent section autoradiography-immunocytochemistry, the distribution of (TH)naloxone binding sites was studied in relation to neuronal systems containing (Leu)enkephalin, dynorphin A, or beta-endorphin immunoreactivity in rat brain. Brain sections from formaldehyde-perfused rats show robust specific binding of (TH)naloxone, the pharmacological (mu-like) properties of which appear unaltered. In contrast, specific binding of the delta ligand (TH)D-Ala2,D-Leu5-enkephalin was virtually totally eliminated as a result of formaldehyde perfusion. Using adjacent section analysis, the authors have noted associations between (TH)naloxone binding sites and one, two, or all three opioid systems in different brain regions; however, in some areas, no apparent relationship could be observed. Within regions, the relationship was complex. The complexity of the association between (TH)naloxone binding sites and the multiple opioid systems, and previous reports of co-localization of mu and kappa receptors in rat brain, are inconsistent with a simple-one-to-one relationship between a given opioid precursor and opioid receptor subtype. Instead, since differential processing of the three precursors gives rise to peptides of varying receptor subtype potencies and selectivities, the multiple peptide-receptor relationships may point to a key role of post-translational processing in determining the physiological consequences of opioid neurotransmission.

  13. PET imaging of human cardiac opioid receptors

    Energy Technology Data Exchange (ETDEWEB)

    Villemagne, Patricia S.R.; Dannals, Robert F. [Department of Radiology, The Johns Hopkins University School of Medicine, 605 N Caroline St., Baltimore, Maryland (United States); Department of Environmental Health Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Ravert, Hayden T. [Department of Radiology, The Johns Hopkins University School of Medicine, 605 N Caroline St., Baltimore, Maryland (United States); Frost, James J. [Department of Radiology, The Johns Hopkins University School of Medicine, 605 N Caroline St., Baltimore, Maryland (United States); Department of Environmental Health Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Department of Neuroscience, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States)

    2002-10-01

    The presence of opioid peptides and receptors and their role in the regulation of cardiovascular function has been previously demonstrated in the mammalian heart. The aim of this study was to image {mu} and {delta} opioid receptors in the human heart using positron emission tomography (PET). Five subjects (three females, two males, 65{+-}8 years old) underwent PET scanning of the chest with [{sup 11}C]carfentanil ([{sup 11}C]CFN) and [{sup 11}C]-N-methyl-naltrindole ([{sup 11}C]MeNTI) and the images were analyzed for evidence of opioid receptor binding in the heart. Either [{sup 11}C]CFN or [{sup 11}C]MeNTI (20 mCi) was injected i.v. with subsequent dynamic acquisitions over 90 min. For the blocking studies, either 0.2 mg/kg or 1 mg/kg of naloxone was injected i.v. 5 min prior to the injection of [{sup 11}C]CFN and [{sup 11}C]MeNTI, respectively. Regions of interest were placed over the left ventricle, left ventricular chamber, lung and skeletal muscle. Graphical analysis demonstrated average baseline myocardial binding potentials (BP) of 4.37{+-}0.91 with [{sup 11}C]CFN and 3.86{+-}0.60 with [{sup 11}C]MeNTI. Administration of 0.2 mg/kg naloxone prior to [{sup 11}C]CFN produced a 25% reduction in BP in one subject in comparison with baseline values, and a 19% decrease in myocardial distribution volume (DV). Administration of 1 mg/kg of naloxone before [{sup 11}C]MeNTI in another subject produced a 14% decrease in BP and a 21% decrease in the myocardial DV. These results demonstrate the ability to image these receptors in vivo by PET. PET imaging of cardiac opioid receptors may help to better understand their role in cardiovascular pathophysiology and the effect of abuse of opioids and drugs on heart function. (orig.)

  14. Mu Opioid Receptor Gene: New Point Mutations in Opioid Addicts

    Directory of Open Access Journals (Sweden)

    Amin Dinarvand

    2014-02-01

    Full Text Available Introduction: Association between single-nucleotide polymorphisms (SNPs in mu opioid receptor gene and drug addiction has been shown in various studies. Here, we have evaluated the existence of polymorphisms in exon 3 of this gene in Iranian population and investigated the possible association between these mutations and opioid addiction.  Methods: 79 opioid-dependent subjects (55 males, 24 females and 134 non-addict or control individuals (74 males, 60 females participated in the study. Genomic DNA was extracted from volunteers’ peripheral blood and exon 3 of the mu opioid receptor gene was amplified by polymerase chain reaction (PCR whose products were then sequenced.  Results: Three different heterozygote polymorphisms were observed in 3 male individuals: 759T>C and 877G>A mutations were found in 2 control volunteers and 1043G>C substitution was observed in an opioid-addicted subject. Association between genotype and opioid addiction for each mutation was not statistically significant.  Discussion: It seems that the sample size used in our study is not enough to confirm or reject any association between 759T>C, 877G>A and 1043G>C substitutions in exon 3 of the mu opioid receptor gene and opioid addiction susceptibility in Iranian population.

  15. Positron Emission Tomography (PET) Imaging of Opioid Receptors

    NARCIS (Netherlands)

    van Waarde, Aren; Absalom, Anthony; Visser, Anniek; Dierckx, Rudi; Dierckx, Rudi AJO; Otte, Andreas; De Vries, Erik FJ; Van Waarde, Aren; Luiten, Paul GM

    2014-01-01

    The opioid system consists of opioid receptors (which mediate the actions of opium), their endogenous ligands (the enkephalins, endorphins, endomorphins, dynorphin, and nociceptin), and the proteins involved in opioid production, transport, and degradation. PET tracers for the various opioid

  16. Opioid binding site in EL-4 thymoma cell line

    International Nuclear Information System (INIS)

    Fiorica, E.; Spector, S.

    1988-01-01

    Using EL-4 thymoma cell-line we found a binding site similar to the k opioid receptor of the nervous system. The Scatchard analysis of the binding of [ 3 H] bremazocine indicated a single site with a K/sub D/ = 60 +/- 17 nM and Bmax = 2.7 +/- 0.8 pmols/10 6 cells. To characterize this binding site, competition studies were performed using selective compounds for the various opioid receptors. The k agonist U-50,488H was the most potent displacer of [ 3 H] bremazocine with an IC 50 value = 0.57μM. The two steroisomers levorphanol and dextrorphan showed the same affinity for this site. While morphine, [D-Pen 2 , D-Pen 5 ] enkephalin and β-endorphin failed to displace, except at very high concentrations, codeine demonstrated a IC 50 = 60μM, that was similar to naloxone. 32 references, 3 figures, 2 tables

  17. Structure of the [delta]-opioid receptor bound to naltrindole

    Energy Technology Data Exchange (ETDEWEB)

    Granier, Sébastien; Manglik, Aashish; Kruse, Andrew C.; Kobilka, Tong Sun; Thian, Foon Sun; Weis, William I.; Kobilka, Brian K. (Stanford-MED)

    2012-07-11

    The opioid receptor family comprises three members, the {mu}-, {delta}- and {kappa}-opioid receptors, which respond to classical opioid alkaloids such as morphine and heroin as well as to endogenous peptide ligands like endorphins. They belong to the G-protein-coupled receptor (GPCR) superfamily, and are excellent therapeutic targets for pain control. The {delta}-opioid receptor ({delta}-OR) has a role in analgesia, as well as in other neurological functions that remain poorly understood. The structures of the {mu}-OR and {kappa}-OR have recently been solved. Here we report the crystal structure of the mouse {delta}-OR, bound to the subtype-selective antagonist naltrindole. Together with the structures of the {mu}-OR and {kappa}-OR, the {delta}-OR structure provides insights into conserved elements of opioid ligand recognition while also revealing structural features associated with ligand-subtype selectivity. The binding pocket of opioid receptors can be divided into two distinct regions. Whereas the lower part of this pocket is highly conserved among opioid receptors, the upper part contains divergent residues that confer subtype selectivity. This provides a structural explanation and validation for the 'message-address' model of opioid receptor pharmacology, in which distinct 'message' (efficacy) and 'address' (selectivity) determinants are contained within a single ligand. Comparison of the address region of the {delta}-OR with other GPCRs reveals that this structural organization may be a more general phenomenon, extending to other GPCR families as well.

  18. The opioid receptors of the rat periaqueductal gray

    Energy Technology Data Exchange (ETDEWEB)

    Fedynyshyn, J.P.

    1989-01-01

    The opioid binding characteristics of the rat (PAG) and the signal transduction mechanisms of the opioid receptors were examined with in vitro radioligand binding, GTPase, adenylyl cyclase, and inositol phosphate assays. The nonselective ligand {sup 3}H-ethylketocyclazocine (EKC), the {mu} and {delta} selective ligand {sup 3}H-(D-Ala{sup 2}, D-Leu{sup 5}) enkephalin (DADLE), the {mu} selective ligand {sup 3}H-(D-Ala{sup 2}, N-methyl Phe{sup 4}, Glyol{sup 5}) enkephalin (DAGO), and the {delta} selective ligand {sup 3}H-(D-Pen{sup 2}, D-Pen{sup 5}) enkephalin (DPDPE) were separately used as tracer ligands to label opioid binding sites in rat PAG enriched P{sub 2} membrane in competition with unlabeled DADLE, DAGO, DPDPE, or the {kappa} selective ligand trans-3,4-dichloro-N-(2-(1-pyrrolidinyl)cyclohexyl)benzeneacetamide, methane sulfonate, hydrate (U50, 488H). Only {mu} selective high affinity opioid binding was observed. No high affinity {delta} or {kappa} selective binding was detected. {sup 3}H-DAGO was used as a tracer ligand to label {mu} selective high affinity opioid binding sites in PAG enriched P{sub 2} membrane in competition with unlabeled {beta}-endorphin, dynorphin A (1-17), BAM-18, methionine enkephalin, dynorphin A (1-8), and leucine enkephalin. Of these endogenous opioid peptides only those with previously reported high affinity {mu} type opioid binding activity competed with {sup 3}H-DAGO for binding sites in rat PAG enriched P{sub 2} membrane with affinities similar to that of unlabeled DAGO.

  19. Quantitative immunolocalization of {mu} opioid receptors: regulation by naltrexone

    Energy Technology Data Exchange (ETDEWEB)

    Evans, C.J.; Lam, H.; To, T.; Anton, B. [Department of Psychiatry and Biobehavioral Sciences, Neuropsychiatric Institute, University of California, Los Angeles, CA (United States); Unterwald, E.M. [Department of Psychiatry, New York University Medical Center, New York, NY (United States)

    1998-04-24

    The present study utilized a newly developed quantitative immunohistochemical assay to measure changes in {mu} opioid receptor abundance following chronic administration of the opioid receptor antagonist naltrexone. These data were compared with those obtained from {mu} receptor radioligand binding on adjacent tissue sections, in order to determine whether the characteristic antagonist-induced increase in radioligand binding is due to an increase in the total number of {mu} receptors and/or to an increase in the proportion of receptors that are in an active binding conformation in the absence of a change in the total number of receptors. Adult male Sprague-Dawley rats were administered naltrexone, 7-8 mg/kg per day, or saline continuously for seven days by osmotic minipumps, after which time their brains were processed for immunohistochemistry and receptor autoradiography on adjacent fresh frozen tissue sections. Semiquantitative immunohistochemistry was performed using a radiolabelled secondary antibody for autoradiographic determination and a set of radioactive standards. Results demonstrate an overall concordance between the distribution of {mu} opioid receptors as measured by the two different methods with a few exceptions. Following naltrexone administration, {mu} receptor immunoreactivity was significantly higher in the amygdala, thalamus, hippocampus, and interpeduncular nucleus as compared with the saline-treated control animals. [{sup 3}H]D-Ala{sup 2},N-Me-Phe{sup 4},Gly-ol{sup 5}-enkephalin binding to {mu} opioid receptors was significantly higher in the globus pallidus, amygdala, thalamus, hypothalamus, hippocampus, substantia nigra, ventral tegmental area, central gray, and interpeduncular nucleus of the naltrexone-treated rats.These findings indicate that in some brain regions chronic naltrexone exposure increases the total number of {mu} opioid receptors, while in other regions there is an increase in the percent of active receptors without an

  20. Quantitative immunolocalization of μ opioid receptors: regulation by naltrexone

    International Nuclear Information System (INIS)

    Evans, C.J.; Lam, H.; To, T.; Anton, B.; Unterwald, E.M.

    1998-01-01

    The present study utilized a newly developed quantitative immunohistochemical assay to measure changes in μ opioid receptor abundance following chronic administration of the opioid receptor antagonist naltrexone. These data were compared with those obtained from μ receptor radioligand binding on adjacent tissue sections, in order to determine whether the characteristic antagonist-induced increase in radioligand binding is due to an increase in the total number of μ receptors and/or to an increase in the proportion of receptors that are in an active binding conformation in the absence of a change in the total number of receptors. Adult male Sprague-Dawley rats were administered naltrexone, 7-8 mg/kg per day, or saline continuously for seven days by osmotic minipumps, after which time their brains were processed for immunohistochemistry and receptor autoradiography on adjacent fresh frozen tissue sections. Semiquantitative immunohistochemistry was performed using a radiolabelled secondary antibody for autoradiographic determination and a set of radioactive standards. Results demonstrate an overall concordance between the distribution of μ opioid receptors as measured by the two different methods with a few exceptions. Following naltrexone administration, μ receptor immunoreactivity was significantly higher in the amygdala, thalamus, hippocampus, and interpeduncular nucleus as compared with the saline-treated control animals. [ 3 H]D-Ala 2 ,N-Me-Phe 4 ,Gly-ol 5 -enkephalin binding to μ opioid receptors was significantly higher in the globus pallidus, amygdala, thalamus, hypothalamus, hippocampus, substantia nigra, ventral tegmental area, central gray, and interpeduncular nucleus of the naltrexone-treated rats.These findings indicate that in some brain regions chronic naltrexone exposure increases the total number of μ opioid receptors, while in other regions there is an increase in the percent of active receptors without an observable change in the total number

  1. Sex differences in opioid analgesia and addiction: interactions among opioid receptors and estrogen receptors

    Science.gov (United States)

    2013-01-01

    Opioids are widely used as the pain reliever and also notorious for being addictive drugs. Sex differences in the opioid analgesia and addiction have been reported and investigated in human subjects and animal models. Yet, the molecular mechanism underlying the differences between males and females is still unclear. Here, we reviewed the literature describing the sex differences in analgesic responses and addiction liabilities to clinically relevant opioids. The reported interactions among opioids, estrogens, opioid receptors, and estrogen receptors are also evaluated. We postulate that the sex differences partly originated from the crosstalk among the estrogen and opioid receptors when stimulated by the exogenous opioids, possibly through common secondary messengers and the downstream gene transcriptional regulators. PMID:24010861

  2. Autoradiographic analysis of mu1, mu2, and delta opioid binding in the central nervous system of C57BL/6BY and CXBK (opioid receptor-deficient) mice

    International Nuclear Information System (INIS)

    Moskowitz, A.S.

    1985-01-01

    In this study the authors used semi-quantitative in vitro autoradiography to compare the levels of binding to central mu 1 , mu 2 , and delta opioid sites in two strains of mice, C57BL/6BY and CXBK. The CXBK strain is known to be deficient in whole brain opioid binding sites and to be less sensitive than the C57 strain to the analgesic and locomotor stimulatory effects of opiates and opioids. Delta sites were visualized using [ 3 H][D-Ala 2 -D-Leu 5 ]-enkephalin (DADL) plus a low concentration of morphine, total mu sites (mu 1 and mu 2 ) were visualized using [ 3 H]dihydromorphine (DHM), and mu 2 sites were visualized using [ 3 H]DHM plus a low concentration of DADL. Binding to mu 1 sites was determined by subtracting mu 2 binding from total mu binding. The authors found that the two strains did not consistently differ in the levels of delta sites. The CXBK strain, however, either had less or the same amount of mu binding as the C57 strain in all areas studied. The CXBK strain was especially deficient in mu 1 binding, particularly in areas involved in pain processing. (Auth.)

  3. Distribution of mu, delta, and kappa opioid receptor binding sites in the brain of the one-day-old domestic chick (Gallus domesticus): An in vitro quantitative autoradiographic study

    Energy Technology Data Exchange (ETDEWEB)

    Csillag, A.; Bourne, R.C.; Stewart, M.G. (Open Univ., Milton Keynes (England))

    1990-12-15

    Three highly specific opioid ligands--(D-Ala2,Gly-ol)-enkephalin (DAGO) for mu (mu) receptor sites, (D-Pen2,D-Pen5)-enkephalin (DPDPE) for delta (delta) sites, and U-69593 for kappa (kappa) sites--were used to determine the regional distribution of the three major subtypes of opioid receptor binding sites in the brains of 1-day-old domestic chicks by the technique of quantitative receptor autoradiography. While there was a degree of heterogeneity in the binding levels of each of the ligands, some notable similarities existed in the binding of the mu and kappa ligands in several forebrain regions, and in the optic tectum of the midbrain where mu and delta binding was very high. In the forebrain there was a high level of binding of mu and kappa ligands in the hyperstriatum, and for the mu ligand there was a very distinct lamination of binding sites in hyperstriatum accessorium, intercalatum supremum, dorsale and ventrale. Levels of binding of the mu and kappa ligands were also high in nucleus basalis, and (for mu only) in the neostriatum. The distribution of binding of the delta specific ligand in the forebrain showed marked differences to that of mu and kappa, being particularly low in the hyperstriatum and neostriatum. Very high levels of labelling of delta binding sites were, however, found in the nucleus rotundus. Binding of the three ligands was generally low or absent in the cerebellum and medulla, apart from a distinct labelling of the granule cell layer by the mu-ligand. A kinetic analysis was made of the binding of the three ligands to whole forebrain sections using scintillation counting methods.

  4. ( sup 3 H)(D-PEN sup 2 , D-PEN sup 5 ) enkephalin binding to delta opioid receptors on intact neuroblastoma-glioma (NG 108-15) hybrid cells

    Energy Technology Data Exchange (ETDEWEB)

    Knapp, R.J.; Yamamura, H.I. (Univ. of Arizona College of Medicine, Tucson (USA))

    1990-01-01

    ({sup 3}H)(D-Pen{sup 2}, D-Pen{sup 5})enkephalin binding to intact NG 108-15 cells has been measured under physiological conditions of temperature and medium. The dissociation constant, receptor density, and Hill slope values measured under these conditions are consistent with values obtained by others using membranes prepared from these cells. Kinetic analysis of the radioligand binding to these cells show biphasic association and monophasic dissociation processes suggesting the presence of different receptor affinity states for the agonist. The data show that the binding affinity of ({sup 3}H)(D-Pen{sup 2}, D-Pen{sup 5})enkephalin under physiological conditions is not substantially different to that measured in 50 mM Tris buffer using cell membrane fractions. Unlike DPDPE, the {mu} opioid agonists morphine, normorphine, PL-17, and DAMGO, have much lower affinity for the {delta} receptor measured under these conditions than is observed by studies using 50 mM Tris buffer. The results described here suggest that this assay may serve as a useful model of {delta} opioid receptor binding in vivo.

  5. Functional characteristics of the naked mole rat μ-opioid receptor.

    Directory of Open Access Journals (Sweden)

    Melanie Busch-Dienstfertig

    Full Text Available While humans and most animals respond to µ-opioid receptor (MOR agonists with analgesia and decreased aggression, in the naked mole rat (NMR opioids induce hyperalgesia and severe aggression. Single nucleotide polymorphisms in the human mu-opioid receptor gene (OPRM1 can underlie altered behavioral responses to opioids. Therefore, we hypothesized that the primary structure of the NMR MOR may differ from other species. Sequencing of the NMR oprm1 revealed strong homology to other mammals, but exposed three unique amino acids that might affect receptor-ligand interactions. The NMR and rat oprm1 sequences were cloned into mammalian expression vectors and transfected into HEK293 cells. Radioligand binding and 3'-5'-cyclic adenosine monophosphate (cAMP enzyme immunoassays were used to compare opioid binding and opioid-mediated cAMP inhibition. At normalized opioid receptor protein levels we detected significantly lower [3H]DAMGO binding to NMR compared to rat MOR, but no significant difference in DAMGO-induced cAMP inhibition. Strong DAMGO-induced MOR internalization was detectable using radioligand binding and confocal imaging in HEK293 cells expressing rat or NMR receptor, while morphine showed weak or no effects. In summary, we found minor functional differences between rat and NMR MOR suggesting that other differences e.g. in anatomical distribution of MOR underlie the NMR's extreme reaction to opioids.

  6. Sigma and opioid receptors in human brain tumors

    International Nuclear Information System (INIS)

    Thomas, G.E.; Szuecs, M.; Mamone, J.Y.; Bem, W.T.; Rush, M.D.; Johnson, F.E.; Coscia, C.J.

    1990-01-01

    Human brain tumors and nude mouse-borne human neuroblastomas and gliomas were analyzed for sigma and opioid receptor content. Sigma binding was assessed using [ 3 H] 1, 3-di-o-tolylguanidine (DTG), whereas opioid receptor subtypes were measured with tritiated forms of the following: μ, [D-ala 2 , mePhe 4 , gly-ol 5 ] enkephalin (DAMGE); κ, ethylketocyclazocine (EKC) or U69,593; δ, [D-pen 2 , D-pen 5 ] enkephalin (DPDPE) or [D-ala 2 , D-leu 5 ] enkephalin (DADLE) with μ suppressor present. Binding parameters were estimated by homologous displacement assays followed by analysis using the LIGAND program. Sigma binding was detected in 15 of 16 tumors examined with very high levels found in a brain metastasis from an adenocarcinoma of lung and a human neuroblastoma (SK-N-MC) passaged in nude mice. κ opioid receptor binding was detected in 4 of 4 glioblastoma multiforme specimens and 2 of 2 human astrocytoma cell lines tested but not in the other brain tumors analyzed

  7. Sigma and opioid receptors in human brain tumors

    Energy Technology Data Exchange (ETDEWEB)

    Thomas, G.E.; Szuecs, M.; Mamone, J.Y.; Bem, W.T.; Rush, M.D.; Johnson, F.E.; Coscia, C.J. (St. Louis Univ. School of Medicine, MO (USA))

    1990-01-01

    Human brain tumors and nude mouse-borne human neuroblastomas and gliomas were analyzed for sigma and opioid receptor content. Sigma binding was assessed using ({sup 3}H) 1, 3-di-o-tolylguanidine (DTG), whereas opioid receptor subtypes were measured with tritiated forms of the following: {mu}, (D-ala{sup 2}, mePhe{sup 4}, gly-ol{sup 5}) enkephalin (DAMGE); {kappa}, ethylketocyclazocine (EKC) or U69,593; {delta}, (D-pen{sup 2}, D-pen{sup 5}) enkephalin (DPDPE) or (D-ala{sup 2}, D-leu{sup 5}) enkephalin (DADLE) with {mu} suppressor present. Binding parameters were estimated by homologous displacement assays followed by analysis using the LIGAND program. Sigma binding was detected in 15 of 16 tumors examined with very high levels found in a brain metastasis from an adenocarcinoma of lung and a human neuroblastoma (SK-N-MC) passaged in nude mice. {kappa} opioid receptor binding was detected in 4 of 4 glioblastoma multiforme specimens and 2 of 2 human astrocytoma cell lines tested but not in the other brain tumors analyzed.

  8. Heterogeneity of Opioid Binding Sites in Guinea Pig Spinal Cord

    Science.gov (United States)

    1984-11-30

    MEDICAL CENTER WILFORD HALL AIR FORCE MEDICAL CENTER Title of Thesis: "Heterogeneity of Opioid Binding Sites in Guinea Pig Spinal Cord" Name of...that the use of any copyrighted material in the dissertation manuscript entitled: "Heterogeneity of Opioid Binding Sites in Guinea Pig Spinal Cord...University of the Health Sciences 11 Abstract Title of Thesis: Heterogenity of Opioid Binding Sites In Guinea Pig Spinal Cord Gary Dean Zarr MAJ/ANC

  9. Biotinylated human. beta. -endorphins as probes for the opioid receptor

    Energy Technology Data Exchange (ETDEWEB)

    Hochhaus, G.; Gibson, B.W.; Sadee, W.

    1988-01-05

    The reaction of human ..beta..-endorphin and biotinyl N-hydroxysuccinimide with or without spacer arm, afforded a series of products that were separated by high performance liquid chromatography (HPLC). Liquid secondary ion mass spectrometry of the biotinylated products and their tryptic digests produced abundant protonated molecular ions (MH/sup +/), which specified the number and location of biotinylation. Between 1 and 4 biotinyl residues were incorporated per human ..beta..-endorphin molecule, at Lys-9, -19, -24, -28, and -29, but not at the amino-terminal Try-1. Three HPLC fractions were isolated for receptor binding studies monobiotinylation of Lys-9, Lys-19, and a mixture of Lys-24, Lys-28, and Lys-29 derivatives. IC/sub 50/ values for binding to ..mu.. and delta opioid receptor sites were 3-8 times higher for monobiotinylated derivatives than for the parent human ..beta..-endorphin. Association with avidin decreased opioid receptor affinities for the C/sub 6/ spacer derivative biotinylated at position Lys-9, which is close to the (1-5) enkephalin receptor region. In contrast, avidin did not affect or even increased apparent affinities to ..mu.. and delta sites for derivatives biotinylated at the ..cap alpha..-helical part of the molecule (Lys-19, -24, -28, and -29). Biotinylated human ..beta..-endorphins also bound to low affinity nonopioid binding sites on NG-108-15 cells; however, affinities to these sites were considerably reduced when derivatives were bound to avidin. The ability of biotinylated human ..beta..-endorphin to cross-link the ..mu.. and delta opioid receptors to avidin allows application of the biotin-avidin system as a molecular probe of the opioid receptor.

  10. The role of opioid antagonist efficacy and constitutive opioid receptor activity in the opioid withdrawal syndrome in mice

    OpenAIRE

    Navani, Dipesh M.; Sirohi, Sunil; Madia, Priyanka A.; Yoburn, Byron C.

    2011-01-01

    On the basis of efficacy, opioid antagonists are classified as inverse opioid agonists (e.g. naltrexone) or neutral opioid antagonists (e.g. 6β-naltrexol). This study examined the interaction between naltrexone and 6β-naltrexol in the precipitated opioid withdrawal syndrome in morphine dependent mice. Furthermore, the possible contribution of constitutive opioid receptor activity to precipitated withdrawal was evaluated using increasing levels of morphine dependence. In the first experiment, ...

  11. Opioid binding site in EL-4 thymoma cell line

    Energy Technology Data Exchange (ETDEWEB)

    Fiorica, E.; Spector, S.

    1988-01-01

    Using EL-4 thymoma cell-line we found a binding site similar to the k opioid receptor of the nervous system. The Scatchard analysis of the binding of (/sup 3/H) bremazocine indicated a single site with a K/sub D/ = 60 +/- 17 nM and Bmax = 2.7 +/- 0.8 pmols/10/sup 6/ cells. To characterize this binding site, competition studies were performed using selective compounds for the various opioid receptors. The k agonist U-50,488H was the most potent displacer of (/sup 3/H) bremazocine with an IC/sub 50/ value = 0.57..mu..M. The two steroisomers levorphanol and dextrorphan showed the same affinity for this site. While morphine, (D-Pen/sup 2/, D-Pen/sup 5/) enkephalin and ..beta..-endorphin failed to displace, except at very high concentrations, codeine demonstrated a IC/sub 50/ = 60..mu..M, that was similar to naloxone. 32 references, 3 figures, 2 tables.

  12. Structure of the Nanobody-Stabilized Active State of the Kappa Opioid Receptor.

    Science.gov (United States)

    Che, Tao; Majumdar, Susruta; Zaidi, Saheem A; Ondachi, Pauline; McCorvy, John D; Wang, Sheng; Mosier, Philip D; Uprety, Rajendra; Vardy, Eyal; Krumm, Brian E; Han, Gye Won; Lee, Ming-Yue; Pardon, Els; Steyaert, Jan; Huang, Xi-Ping; Strachan, Ryan T; Tribo, Alexandra R; Pasternak, Gavril W; Carroll, F Ivy; Stevens, Raymond C; Cherezov, Vadim; Katritch, Vsevolod; Wacker, Daniel; Roth, Bryan L

    2018-01-11

    The κ-opioid receptor (KOP) mediates the actions of opioids with hallucinogenic, dysphoric, and analgesic activities. The design of KOP analgesics devoid of hallucinatory and dysphoric effects has been hindered by an incomplete structural and mechanistic understanding of KOP agonist actions. Here, we provide a crystal structure of human KOP in complex with the potent epoxymorphinan opioid agonist MP1104 and an active-state-stabilizing nanobody. Comparisons between inactive- and active-state opioid receptor structures reveal substantial conformational changes in the binding pocket and intracellular and extracellular regions. Extensive structural analysis and experimental validation illuminate key residues that propagate larger-scale structural rearrangements and transducer binding that, collectively, elucidate the structural determinants of KOP pharmacology, function, and biased signaling. These molecular insights promise to accelerate the structure-guided design of safer and more effective κ-opioid receptor therapeutics. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Opioid receptor mediated anticonvulsant effect of pentazocine.

    Science.gov (United States)

    Khanna, N; Khosla, R; Kohli, J

    1998-01-01

    Intraperitoneal (i.p.) administration of (+/-) pentazocine (10, 30 & 50 mg/kg), a Sigma opioid agonist, resulted in a dose dependent anticonvulsant action against maximal electroshock seizures in mice. This anticonvulsant effect of pentazocine was not antagonized by both the doses of naloxone (1 and 10 mg/kg) suggesting thereby that its anticonvulsant action is probably mediated by Sigma opiate binding sites. Its anticonvulsant effect was potentiated by both the anticonvulsant drugs viz. diazepam and diphenylhydantoin. Morphine, mu opioid agonist, on the other hand, failed to protect the animals against maximal electroshock seizures when it was given in doses of 10-40 mg/kg body wt.

  14. Computer Modeling of Human Delta Opioid Receptor

    Directory of Open Access Journals (Sweden)

    Tatyana Dzimbova

    2013-04-01

    Full Text Available The development of selective agonists of δ-opioid receptor as well as the model of interaction of ligands with this receptor is the subjects of increased interest. In the absence of crystal structures of opioid receptors, 3D homology models with different templates have been reported in the literature. The problem is that these models are not available for widespread use. The aims of our study are: (1 to choose within recently published crystallographic structures templates for homology modeling of the human δ-opioid receptor (DOR; (2 to evaluate the models with different computational tools; and (3 to precise the most reliable model basing on correlation between docking data and in vitro bioassay results. The enkephalin analogues, as ligands used in this study, were previously synthesized by our group and their biological activity was evaluated. Several models of DOR were generated using different templates. All these models were evaluated by PROCHECK and MolProbity and relationship between docking data and in vitro results was determined. The best correlations received for the tested models of DOR were found between efficacy (erel of the compounds, calculated from in vitro experiments and Fitness scoring function from docking studies. New model of DOR was generated and evaluated by different approaches. This model has good GA341 value (0.99 from MODELLER, good values from PROCHECK (92.6% of most favored regions and MolProbity (99.5% of favored regions. Scoring function correlates (Pearson r = -0.7368, p-value = 0.0097 with erel of a series of enkephalin analogues, calculated from in vitro experiments. So, this investigation allows suggesting a reliable model of DOR. Newly generated model of DOR receptor could be used further for in silico experiments and it will give possibility for faster and more correct design of selective and effective ligands for δ-opioid receptor.

  15. Effects of stress and. beta. -funal trexamine pretreatment on morphine analgesia and opioid binding in rats

    Energy Technology Data Exchange (ETDEWEB)

    Adams, J.U.; Andrews, J.S.; Hiller, J.M.; Simon, E.J.; Holtzman, S.G.

    1987-12-28

    This study was essentially an in vivo protection experiment designed to test further the hypothesis that stress induces release of endogenous opiods which then act at opioid receptors. Rats that were either subjected to restraint stress for 1 yr or unstressed were injected ICV with either saline or 2.5 ..mu..g of ..beta..-funaltrexamine (..beta..-FNA), an irreversible opioid antagonist that alkylates the mu-opioid receptor. Twenty-four hours later, subjects were tested unstressed for morphine analgesia or were sacrificed and opioid binding in brain was determined. (/sup 3/H)D-Ala/sup 2/NMePhe/sup 4/-Gly/sup 5/(ol)enkephalin (DAGO) served as a specific ligand for mu-opioid receptors, and (/sup 3/H)-bremazocine as a general ligand for all opioid receptors. Rats injected with saline while stressed were significantly less sensitive to the analgesic action of morphine 24 hr later than were their unstressed counterparts. ..beta..-FNA pretreatment attenuated morphine analgesia in an insurmountable manner. Animals pretreated with ..beta..-FNA while stressed were significantly more sensitive to the analgesic effect of morphine than were animals that received ..beta..-FNA while unstressed. ..beta..-FNA caused small and similar decreases in (/sup 3/H)-DAGO binding in brain of both stressed and unstressed animals. 35 references, 2 figures, 2 tables.

  16. Synthesis of opioid receptor imaging agent 7α-o-IA-DPN

    International Nuclear Information System (INIS)

    Wang Rongfu

    1997-01-01

    A new opioid receptor imaging agent is designed and synthesized. 7α-o-iodoallyl diprenorphine (7α-o-IA-DPN) was obtained in one step by radioiododestannylation, which involved in the selection of DPN as an opioid antagonist, the regioselective protection of the DPN phenol tertiary-OH using acetylation and the introduction of vinylstannane as prosthetic group into the tertiary alcohol group position in the 7α-side chain. The iodinated DPN derivation was possessed of high radiolabeled yield (>90%) with 80 TBq/mmol specific radioactivity and more than 95% radiochemical purity. In vitro opioid receptor binding analysis showed very high affinity (Ki = 0.4 nmol/L). This new radioiodinated opioid ligand is suitable for SPECT study of opioid receptor imaging

  17. Sigma opioid receptor: characterization and co-identity with the phencyclidine receptor

    International Nuclear Information System (INIS)

    Mendelsohn, L.G.; Kalra, V.; Johnson, B.G.; Kerchner, G.A.

    1985-01-01

    The properties of the sigma opioid receptor of rat brain cortex have been characterized using the prototypic ligand (+)-[ 3 H] SKF 10,047. Binding to this receptor was rapid, and equilibrium was obtained within 30 min at 37 degrees C. Specific binding was linear with protein concentration up to 500 micrograms/2 ml and was dependent upon protein integrity. Denaturation by boiling destroyed over 95% of the specific binding. A high-affinity binding site with a KD of 150 +/- 40 nM and a maximum binding of 2.91 +/- 0.84 pmol/mg of protein was determined from a Scatchard plot of the binding data. The addition of salt, either NaCl or CaCl 2 , to the buffers markedly decreased binding, with CaCl 2 being more potent than NaCl. A broad pH optimum for specific binding was observed; maximum binding was at pH 9.0. The affinity of a number of ligands for the sigma site and the phencyclidine receptor were compared. The binding (IC50) of 13 ligands to the sigma site showed a correlation of 0.86 (P less than .01) with binding to the phencyclidine site. The data demonstrate that the biochemical properties of the sigma and phencyclidine receptors are similar and support the view that these receptors are one and the same site

  18. Opioid modulation of immunocompetence: Receptor characterization and second messenger involvement

    International Nuclear Information System (INIS)

    Hemmick, L.M.

    1989-01-01

    The purpose of this thesis was to examine the effects of opioids on several indices of immunocompetence, determined the receptor specificity of these effects, and ascertain whether the actions of opioids on lymphocytes could be correlated with activation of second messenger systems. By measuring 45 Ca 2+ uptake into lymphocytes, it was demonstrated that β-endorphin 1-31 (β-END 1-31) enhanced rat thymocyte Ca 2+ uptake in response to concanavalin A (Con A) but not phytohemagglutinin (PHA). Related opioid peptides and alkaloids were unable to mimic the effect, and naloxone did not block it, suggesting that β-END 1-31 acted by binding to specific, non-opioid receptors on the thymocytes. Rat splenocyte Con A-stimulated Ca 2+ uptake was not affected by β-END 1-31. β-END 1-31 did not affect basal Ca 2+ uptake by either cell type. Using [ 3 H]thymidine uptake as an index of lymphocyte proliferation, β-END 1-31 and several related opioid peptides reversed prostaglandin E 1 (PGE 1 ) suppression of rat lymph node cell Con A- and PHA-stimulated proliferation. Naloxone did not block the reversal. β-END 1-31 was unable to reverse forskolin and cholera toxin suppression of proliferation, indicating that the lowering of cyclic AMP levels was not the mechanism involved. Verapamil inhibition of proliferation was also not reversed by β-END 1-31, suggesting that promotion of Ca 2+ influx was not a major mechanism involved

  19. Opioid modulation of immunocompetence: Receptor characterization and second messenger involvement

    Energy Technology Data Exchange (ETDEWEB)

    Hemmick, L.M.

    1989-01-01

    The purpose of this thesis was to examine the effects of opioids on several indices of immunocompetence, determined the receptor specificity of these effects, and ascertain whether the actions of opioids on lymphocytes could be correlated with activation of second messenger systems. By measuring {sup 45}Ca{sup 2+} uptake into lymphocytes, it was demonstrated that {beta}-endorphin 1-31 ({beta}-END 1-31) enhanced rat thymocyte Ca{sup 2+} uptake in response to concanavalin A (Con A) but not phytohemagglutinin (PHA). Related opioid peptides and alkaloids were unable to mimic the effect, and naloxone did not block it, suggesting that {beta}-END 1-31 acted by binding to specific, non-opioid receptors on the thymocytes. Rat splenocyte Con A-stimulated Ca{sup 2+} uptake was not affected by {beta}-END 1-31. {beta}-END 1-31 did not affect basal Ca{sup 2+} uptake by either cell type. Using ({sup 3}H)thymidine uptake as an index of lymphocyte proliferation, {beta}-END 1-31 and several related opioid peptides reversed prostaglandin E{sub 1} (PGE{sub 1}) suppression of rat lymph node cell Con A- and PHA-stimulated proliferation. Naloxone did not block the reversal. {beta}-END 1-31 was unable to reverse forskolin and cholera toxin suppression of proliferation, indicating that the lowering of cyclic AMP levels was not the mechanism involved. Verapamil inhibition of proliferation was also not reversed by {beta}-END 1-31, suggesting that promotion of Ca{sup 2+} influx was not a major mechanism involved.

  20. Evidence for a non-opioid sigma binding site din the guinea-pig myenteric plexus

    International Nuclear Information System (INIS)

    Roman, F.; Pascaud, X.; Vauche, D.; Junien, J.

    1988-01-01

    The presence of a binding site to (+)-( 3 H)SKF 10,047 was demonstrated in a guinea-pig myenteric plexus (MYP) membrane preparation. Specific binding to this receptor was saturable, reversible, linear with protein concentration and consisted of two components, a high affinity site and a low affinity site. Morphine and naloxone 10 -4 M were unable to displace (+)-( 3 H)SKF 10,047 binding. Haloperidol, imipramine, ethylketocyclazocine and propranolol were among the most potent compounds to inhibit this specific binding. These results suggest the presence of a non-opioid haloperidol sensitive sigma receptor in the MYP of the guinea-pig

  1. Using [11C]diprenorphine to image opioid receptor occupancy by methadone in opioid addiction: clinical and preclinical studies.

    Science.gov (United States)

    Melichar, Jan K; Hume, Susan P; Williams, Tim M; Daglish, Mark R C; Taylor, Lindsay G; Ahmad, Rabia; Malizia, Andrea L; Brooks, David J; Myles, Judith S; Lingford-Hughes, Anne; Nutt, David J

    2005-01-01

    Substitute methadone prescribing is one of the main modes of treatment for opioid dependence with established evidence for improved health and social outcomes. However, the pharmacology underpinning the effects of methadone is little studied despite controversies about dosing in relation to outcome. We therefore examined the relationship between methadone dose and occupation of opioid receptors in brain using the positron emission tomography (PET) radioligand [(11)C]diprenorphine in humans and rats. Eight opioid-dependent subjects stable on their substitute methadone (18-90 mg daily) had an [(11)C]diprenorphine PET scan at predicted peak plasma levels of methadone. These were compared with eight healthy controls. No difference in [(11)C]diprenorphine binding was found between the groups, with no relationship between methadone dose and occupancy. Adult male Sprague-Dawley rats that had been given an acute i.v. injection of methadone hydrochloride (0.35, 0.5, 0.7, or 1.0 mg kg(-1)) before [(11)C]diprenorphine showed a dose-dependent increase in biodistribution but no reduction in [(11)C]diprenorphine binding. We suggest that the lack of a dose-dependent relationship between methadone dose, either given chronically in human or acutely in rat, and occupancy of opioid receptor measured with [(11)C]diprenorphine PET is related to efficacy of this opioid agonist at very low levels of opioid receptor occupancy. This has implications for understanding the actions of methadone in comparison with other opioid drugs such as partial agonists and antagonists.

  2. Purification and characterization of mu-specific opioid receptor from rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Hasegawa, J.; Cho, T.M.; Ge, B.L.; Loh, H.H.

    1986-03-05

    A mu-specific opioid receptor was purified to apparent homogeneity from rat brain membranes by 6-succinylmorphine affinity chromatography, Ultrogel filtration, wheat germ agglutinin affinity chromatography, and isoelectric focusing. The purified receptor had a molecular weight of 58,000 as determined by polyacrylamide gel electrophoresis, and was judged to be homogeneous by the following criteria: (1) a single band on the SDS gel; and (2) a specific opioid binding activity of 17,720 pmole/mg protein, close to the theoretical value. In addition, the 58,000 molecular weight value agrees closely with that determined by covalently labelling purified receptor with bromoacetyl-/sup 3/H-dihydromorphine or with /sup 125/I-beta-endorphin and dimethyl suberimidate. To their knowledge, this is the first complete purification of an opioid receptor that retains its ability to bind opiates.

  3. FMRFamide: low affinity inhibition of opioid binding to rabbit brain membranes

    International Nuclear Information System (INIS)

    Zhu, X.Z.; Raffa, R.B.

    1986-01-01

    FMRFamide (Phe-Met-Arg-Phe-NH 2 ) was first isolated from the ganglia of molluscs by Price and Greenberg in 1977. The peptide was subsequently shown to have diverse actions on various types of molluscan and mammalian tissues. The presence of immunoreactive FMRFamide-like material (irFMRF) in multiple areas of rat brain, spinal cord, and gastrointestinal tract suggests that irFMRF may have a physiological role in mammals. Tang, Yang and Costa recently demonstrated that FMRFamide attenuates morphine antinociception in rats and postulated, based on this and several other lines of evidence, that irFMRF might be an endogenous opioid antagonist. In the present study, they tested the ability of FMRFamide to inhibit the binding of opioid receptor ligands to rabbit membrane preparations. FMRFamide inhibited the specific binding of both 3 [H]-dihydromorphine and 3 [H]-ethylketocyclazocine (IC 50 = 14 μM and 320 μM, respectively) in a dose-related manner, suggesting that FMRFamide may affect binding to at least two types of opioid receptors (mu and kappa). These data are consistent with the concept that irFMRF might act as an endogenous opioid antagonist. However, the low affinity of FMRFamide leaves open the possibility of another mechanism of opioid antagonism, such as neuromodulation

  4. FMRFamide: low affinity inhibition of opioid binding to rabbit brain membranes

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, X.Z.; Raffa, R.B.

    1986-03-05

    FMRFamide (Phe-Met-Arg-Phe-NH/sub 2/) was first isolated from the ganglia of molluscs by Price and Greenberg in 1977. The peptide was subsequently shown to have diverse actions on various types of molluscan and mammalian tissues. The presence of immunoreactive FMRFamide-like material (irFMRF) in multiple areas of rat brain, spinal cord, and gastrointestinal tract suggests that irFMRF may have a physiological role in mammals. Tang, Yang and Costa recently demonstrated that FMRFamide attenuates morphine antinociception in rats and postulated, based on this and several other lines of evidence, that irFMRF might be an endogenous opioid antagonist. In the present study, they tested the ability of FMRFamide to inhibit the binding of opioid receptor ligands to rabbit membrane preparations. FMRFamide inhibited the specific binding of both /sup 3/(H)-dihydromorphine and /sup 3/(H)-ethylketocyclazocine (IC/sub 50/ = 14 ..mu..M and 320 ..mu..M, respectively) in a dose-related manner, suggesting that FMRFamide may affect binding to at least two types of opioid receptors (mu and kappa). These data are consistent with the concept that irFMRF might act as an endogenous opioid antagonist. However, the low affinity of FMRFamide leaves open the possibility of another mechanism of opioid antagonism, such as neuromodulation.

  5. Reconstitution of high-affinity opioid agonist binding in brain membranes

    Energy Technology Data Exchange (ETDEWEB)

    Remmers, A.E.; Medzihradsky, F. (Univ. of Michigan Medical School, Ann Arbor (United States))

    1991-03-15

    In synaptosomal membranes from rat brain cortex, the {mu} selective agonist ({sup 3}H)dihydromorphine in the absence of sodium, and the nonselective antagonist ({sup 3}H)naltrexone in the presence of sodium, bound to two populations of opioid receptor sites with K{sub d} values of 0.69 and 8.7 nM for dihydromorphine, and 0.34 and 5.5 nM for naltrexone. The addition of 5 {mu}M guanosine 5{prime}-({gamma}-thio)triphosphate (GTP({gamma}S)) strongly reduced high-affinity agonist but not antagonist binding. Exposure of the membranes to high pH reduced the number of GTP({gamma}-{sup 35}S) binding sites by 90% and low K{sub m}, opioid-sensitive GTPase activity by 95%. In these membranes, high-affinity agonist binding was abolished and modulation of residual binding by GTP({gamma}S) was diminished. Alkali treatment of the glioma cell membranes prior to fusion inhibited most of the low K{sub m} GTPase activity and prevented the reconstitution of agonist binding. The results show that high-affinity opioid agonist binding reflects the ligand-occupied receptor - guanine nucleotide binding protein complex.

  6. Opioid receptor desensitization: mechanisms and its link to tolerance

    Directory of Open Access Journals (Sweden)

    Stéphane eAllouche

    2014-12-01

    Full Text Available Opioid receptors are part of the class A of G-protein coupled receptors and the target of the opiates, the most powerful analgesic molecules used in clinic. During a protracted use, a tolerance to analgesic effect develops resulting in a reduction of the effectiveness. So understanding mechanisms of tolerance is a great challenge and may help to find new strategies to tackle this side effect. This review will summarize receptor-related mechanisms that could underlie tolerance especially receptor desensitization. We will focus on the latest data obtained on molecular mechanisms involved in opioid receptor desensitization: phosphorylation, receptor uncoupling, internalization and post-endocytic fate of the receptor.

  7. Positive transcriptional regulation of the human micro opioid receptor gene by poly(ADP-ribose) polymerase-1 and increase of its DNA binding affinity based on polymorphism of G-172 -> T.

    Science.gov (United States)

    Ono, Takeshi; Kaneda, Toshio; Muto, Akihiro; Yoshida, Tadashi

    2009-07-24

    Micro opioid receptor (MOR) agonists such as morphine are applied widely in clinical practice as pain therapy. The effects of morphine through MOR, such as analgesia and development of tolerance and dependence, are influenced by individual specificity. Recently, we analyzed single nucleotide polymorphisms on the human MOR gene to investigate the factors that contribute to individual specificity. In process of single nucleotide polymorphisms analysis, we found that specific nuclear proteins bound to G(-172) --> T region in exon 1 in MOR gene, and its affinity to DNA was increased by base substitution from G(-172) to T(-172). The isolated protein was identified by mass spectrometry and was confirmed by Western blotting to be poly(ADP-ribose) polymerase-1 (PARP-1). The overexpressed PARP-1 bound to G(-172) --> T and enhanced the transcription of reporter vectors containing G(-172) and T(-172). Furthermore, PARP-1 inhibitor (benzamide) decreased PARP-1 binding to G(-172) --> T without affecting mRNA or protein expression level of PARP-1 and down-regulated the subsequent MOR gene expression in SH-SY5Y cells. Moreover, we found that tumor necrosis factor-alpha enhanced MOR gene expression as well as increased PARP-1 binding to the G(-172) --> T region and G(-172) --> T-dependent transcription in SH-SY5Y cells. These effects were also inhibited by benzamide. In this study, our data suggest that PARP-1 positively regulates MOR gene transcription via G(-172) --> T, which might influence individual specificity in therapeutic opioid effects.

  8. Expression of μ, κ, and δ opioid receptor messenger RNA in the human CNS: a 33P in situ hybridization study

    International Nuclear Information System (INIS)

    Peckys, D.; Landwehrmeyer, G.B.

    1999-01-01

    The existence of at least three opioid receptor types, referred to as μ, κ, and δ, is well established. Complementary DNAs corresponding to the pharmacologically defined μ, κ, and δ opioid receptors have been isolated in various species including man. The expression patterns of opioid receptor transcripts in human brain has not been established with a cellular resolution, in part because of the low apparent abundance of opioid receptor messenger RNAs in human brain. To visualize opioid receptor messenger RNAs we developed a sensitive in situ hybridization histochemistry method using 33 P-labelled RNA probes. In the present study we report the regional and cellular expression of μ, κ, and δ opioid receptor messenger RNAs in selected areas of the human brain. Hybridization of the different opioid receptor probes resulted in distinct labelling patterns. For the μ and κ opioid receptor probes, the most intense regional signals were observed in striatum, thalamus, hypothalamus, cerebral cortex, cerebellum and certain brainstem areas as well as the spinal cord. The most intense signals for the δ opioid receptor probe were found in cerebral cortex. Expression of opioid receptor transcripts was restricted to subpopulations of neurons within most regions studied demonstrating differences in the cellular expression patterns of μ, κ, and δ opioid receptor messenger RNAs in numerous brain regions. The messenger RNA distribution patterns for each opioid receptor corresponded in general to the distribution of opioid receptor binding sites as visualized by receptor autoradiography. However, some mismatches, for instance between μ opioid receptor receptor binding and μ opioid receptor messenger RNA expression in the anterior striatum, were observed. A comparison of the distribution patterns of opioid receptor messenger RNAs in the human brain and that reported for the rat suggests a homologous expression pattern in many regions. However, in the human brain, κ

  9. Interaction of trimebutine and Jo-1196 (fedotozine) with opioid receptors in the canine ileum

    Energy Technology Data Exchange (ETDEWEB)

    Allescher, H.D.; Ahmad, S.; Classen, M.; Daniel, E.E. (Technical Univ., Munich, (West Germany))

    1991-05-01

    Receptor binding of the opioid receptor antagonist, ({sup 3}H)diprenorphine, which has a similar affinity to the various opioid receptor subtypes, was characterized in subcellular fractions derived from either longitudinal or circular smooth muscle of the canine small intestine with their plexuses (myenteric plexus and deep muscular plexus, respectively) attached. The distribution of opioid binding activity showed a good correlation in the different fractions with the binding of the neuronal marker ({sup 3}H)saxitoxin but no correlation to the smooth muscle plasma membrane marker 5'-nucleotidase. The saturation data (Kd = 0.12 +/- 0.04 nM and maximum binding = 400 +/- 20 fmol/mg) and the data from kinetic experiments (Kd = 0.08 nmol) in the myenteric plexus were in good agreement with results obtained previously from the circular muscle/deep muscular plexus preparation. Competition experiments using selective drugs for mu (morphiceptin-analog (N-MePhe3-D-Pro4)-morphiceptin), delta (D-Pen2,5-enkephalin) and kappa (dynorphin 1-13, U50488-H) ligands showed the existence of all three receptor subtypes. The existence of kappa receptors was confirmed in saturation experiments using ({sup 3}H) ethylketocycloazocine as labeled ligand. Two putative opioid agonists, with effects on gastrointestinal motility, trimebutine and JO-1196 (fedotozin), were also examined. Trimebutine (Ki = 0.18 microM), Des-Met-trimebutine (Ki = 0.72 microM) and Jo-1196 (Ki = 0.19 microM) displaced specific opiate binding. The relative affinity for the opioid receptor subtypes was mu = 0.44, delta = 0.30 and kappa = 0.26 for trimebutine and mu = 0.25, delta = 0.22 and kappa = 0.52 for Jo-1196.

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

    Directory of Open Access Journals (Sweden)

    Li He

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

  11. [11C]-MeJDTic: a novel radioligand for κ-opioid receptor positron emission tomography imaging

    International Nuclear Information System (INIS)

    Poisnel, Geraldine; Oueslati, Farhana; Dhilly, Martine; Delamare, Jerome; Perrio, Cecile; Debruyne, Daniele; Barre, Louisa

    2008-01-01

    Introduction: Radiopharmaceuticals that can bind selectively the κ-opioid receptor may present opportunities for staging clinical brain disorders and evaluating the efficiency of new therapies related to stroke, neurodegenerative diseases or opiate addiction. The N-methylated derivative of JDTic (named MeJDTic), which has been recently described as a potent and selective antagonist of κ-opioid receptor in vitro, was labeled with carbon-11 and evaluated for in vivo imaging the κ-opioid receptor in mice. Methods: [ 11 C]-MeJDTic was prepared by methylation of JDTic with [ 11 C]-methyl triflate. The binding of [ 11 C]-MeJDTic to κ-opioid receptor was investigated ex vivo by biodistribution and competition studies using nonfasted male CD1 mice. Results: [ 11 C]-MeJDTic exhibited a high and rapid distribution in peripheral organs. The uptake was maximal in lung where the κ receptor is largely expressed. [ 11 C]-MeJDTic rapidly crossed the blood-brain barrier and accumulated in the brain regions of interest (hypothalamus). The parent ligand remained the major radioactive compound in brain during the experiment. Chase studies with U50,488 (a κ referring agonist), morphine (a μ agonist) and naltrindole (a δ antagonist) demonstrated that this uptake was the result of specific binding to the κ-opioid receptor. Conclusion: These findings suggested that [ 11 C]-MeJDTic appeared to be a promising selective 'lead' radioligand for κ-opioid receptor PET imaging

  12. Site-specific effects of the nonsteroidal anti-inflammatory drug lysine clonixinate on rat brain opioid receptors.

    Science.gov (United States)

    Ortí, E; Coirini, H; Pico, J C

    1999-04-01

    In addition to effects in the periphery through inhibition of prostaglandin synthesis, several lines of evidence suggest that nonsteroidal anti-inflammatory drugs (NSAIDs) act in the central nervous system. The possibility that the central action of NSAIDs involves regulation of opioid receptors was investigated by quantitative autoradiography of mu, delta, and kappa sites in rat brain slices. Increased (p lysine clonixinate. Labeling of delta receptors was lower in the lateral septum, and kappa sites decreased in thalamic nuclei. These effects were not mediated through direct interaction with opioid-binding sites, since receptor-binding assays using rat brain membranes confirmed that clonixinate up to 1 x 10(-4) mol/l does not inhibit mu, delta, and kappa receptor specific binding. Central effects of NSAIDs might, therefore, involve interaction with the opioid receptor system through indirect mechanisms.

  13. Opioid receptors in midbrain dopaminergic regions of the rat. 1. Mu receptor autoradiography

    International Nuclear Information System (INIS)

    German, D.C.; Speciale, S.G.; Manaye, K.F.; Sadeq, M.

    1993-01-01

    Several lines of evidence indicate that an interaction exists between opioid peptides and midbrain dopaminergic neurons. The purpose of this study was to map and quantify the density of the mu opioid receptor subtype relative to the location of the dopaminergic (DA) neurons in the retrorubral field (nucleus A8), substantia nigra (nucleus A9), and ventral tegmental area and related nuclei (nucleus A10) in the rat. Sections through the rostral-caudal extent of the midbrain were stained with an antibody against tyrosine hydroxylase, as a DA cell marker, and comparable sections were processed for in vitro receptor autoradiography using the mu-selective ligand, 3 H-Tyr-D-Ala-N-MePhe-Gyl-ol enkephalin. In the nucleus A8 region, there were low levels of mu binding. In the rostral portion of nucleus A9, there was prominent mu binding both in the ventral pars compacta, which contains numerous DA neurons, and in regions that correspond to the location of the DA dendrites which project ventrally into the underlying substantia nigra pars reticulata. In the caudal portion of nucleus A9, mu binding was greatest in the substantia nigra pars reticulata, but also in the same region that contains DA neurons. In nucleus A10, mu receptor densities differed depending upon the nucleus A10 subdivision, and the rostral-caudal position in the nucleus. Low receptor densities were observed in rostral portions of the ventral tegmental area and interfascicular nucleus, and there was negligible binding in the parabrachial pigmented nucleus and paranigral nucleus at the level of the interpeduncular nucleus; all regions where there are high densities of DA somata. Mu binding was relatively high in the central linear nucleus, and in the dorsal and medial divisions of the medial terminal nucleus of the accessory optic system, which has been shown to contain DA dendrites. These data indicate that mu opioid receptors are located in certain regions occupied by all three midbrain DA nuclei, but in a

  14. Opioid binding sites in the guinea pig and rat kidney: Radioligand homogenate binding and autoradiography

    Energy Technology Data Exchange (ETDEWEB)

    Dissanayake, V.U.; Hughes, J.; Hunter, J.C. (Parke-Davis Research Unit, Addenbrookes Hospital Site, Cambridge (England))

    1991-07-01

    The specific binding of the selective {mu}-, {delta}-, and {kappa}-opioid ligands (3H)(D-Ala2,MePhe4,Gly-ol5)enkephalin ((3H) DAGOL), (3H)(D-Pen2,D-Pen5)enkephalin ((3H)DPDPE), and (3H)U69593, respectively, to crude membranes of the guinea pig and rat whole kidney, kidney cortex, and kidney medulla was investigated. In addition, the distribution of specific 3H-opioid binding sites in the guinea pig and rat kidney was visualized by autoradiography. Homogenate binding and autoradiography demonstrated the absence of {mu}- and {kappa}-opioid binding sites in the guinea pig kidney. No opioid binding sites were demonstrable in the rat kidney. In the guinea pig whole kidney, cortex, and medulla, saturation studies demonstrated that (3H)DPDPE bound with high affinity (KD = 2.6-3.5 nM) to an apparently homogeneous population of binding sites (Bmax = 8.4-30 fmol/mg of protein). Competition studies using several opioid compounds confirmed the nature of the {delta}-opioid binding site. Autoradiography experiments demonstrated that specific (3H)DPDPE binding sites were distributed radially in regions of the inner and outer medulla and at the corticomedullary junction of the guinea pig kidney. Computer-assisted image analysis of saturation data yielded KD values (4.5-5.0 nM) that were in good agreement with those obtained from the homogenate binding studies. Further investigation of the {delta}-opioid binding site in medulla homogenates, using agonist ((3H)DPDPE) and antagonist ((3H)diprenorphine) binding in the presence of Na+, Mg2+, and nucleotides, suggested that the {delta}-opioid site is linked to a second messenger system via a GTP-binding protein. Further studies are required to establish the precise localization of the {delta} binding site in the guinea pig kidney and to determine the nature of the second messenger linked to the GTP-binding protein in the medulla.

  15. Biased Agonism of Endogenous Opioid Peptides at the μ-Opioid Receptor.

    Science.gov (United States)

    Thompson, Georgina L; Lane, J Robert; Coudrat, Thomas; Sexton, Patrick M; Christopoulos, Arthur; Canals, Meritxell

    2015-08-01

    Biased agonism is having a major impact on modern drug discovery, and describes the ability of distinct G protein-coupled receptor (GPCR) ligands to activate different cell signaling pathways, and to result in different physiologic outcomes. To date, most studies of biased agonism have focused on synthetic molecules targeting various GPCRs; however, many of these receptors have multiple endogenous ligands, suggesting that "natural" bias may be an unappreciated feature of these GPCRs. The μ-opioid receptor (MOP) is activated by numerous endogenous opioid peptides, remains an attractive therapeutic target for the treatment of pain, and exhibits biased agonism in response to synthetic opiates. The aim of this study was to rigorously assess the potential for biased agonism in the actions of endogenous opioids at the MOP in a common cellular background, and compare these to the effects of the agonist d-Ala2-N-MePhe4-Gly-ol enkephalin (DAMGO). We investigated activation of G proteins, inhibition of cAMP production, extracellular signal-regulated kinase 1 and 2 phosphorylation, β-arrestin 1/2 recruitment, and MOP trafficking, and applied a novel analytical method to quantify biased agonism. Although many endogenous opioids displayed signaling profiles similar to that of DAMGO, α-neoendorphin, Met-enkephalin-Arg-Phe, and the putatively endogenous peptide endomorphin-1 displayed particularly distinct bias profiles. These may represent examples of natural bias if it can be shown that they have different signaling properties and physiologic effects in vivo compared with other endogenous opioids. Understanding how endogenous opioids control physiologic processes through biased agonism can reveal vital information required to enable the design of biased opioids with improved pharmacological profiles and treat diseases involving dysfunction of the endogenous opioid system. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

  16. Effect of prenatal methadone and ethanol on opioid receptor development in rats

    Energy Technology Data Exchange (ETDEWEB)

    Peters, M.A.; Braun, R.L. (Loma Linda Univ., CA (United States))

    1991-03-11

    The current literature shows that the offspring of female rats exposed to methadone or ethanol display similar neurochemical and neurobehavioral alterations, and suggests that these drugs may be operating through a common mechanism. If this hypothesis is true, their effect on the endogenous opioid systems should be qualitatively similar. In this study virgin females were treated with methadone or 10% ethanol oral solution starting prior to conception and continued throughout gestation. When the offspring had reached 15 or 30 days of age they were sacrificed, the brain was removed and prepared for opioid receptor binding studies. ({sup 3}H)DAGO and ({sup 3}H)DADLE were used as ligands for the mu and delta receptors, respectively. These studies show significant treatment-related differences in both the number of mu and delta binding sites as well as in apparent receptor affinity. Significant sex- and age-related differences between treatments were also observed. These data show that methadone and ethanol, while manifesting some similar neurochemical and behavioral effects, have unique effects on opioid receptor binding, suggesting that they may be acting by different mechanisms.

  17. GRK2 Constitutively Governs Peripheral Delta Opioid Receptor Activity

    Directory of Open Access Journals (Sweden)

    Allison Doyle Brackley

    2016-09-01

    Full Text Available Opioids remain the standard for analgesic care; however, adverse effects of systemic treatments contraindicate long-term administration. While most clinical opioids target mu opioid receptors (MOR, those that target the delta class (DOR also demonstrate analgesic efficacy. Furthermore, peripherally restrictive opioids represent an attractive direction for analgesia. However, opioid receptors including DOR are analgesically incompetent in the absence of inflammation. Here, we report that G protein-coupled receptor kinase 2 (GRK2 naively associates with plasma membrane DOR in peripheral sensory neurons to inhibit analgesic agonist efficacy. This interaction prevents optimal Gβ subunit association with the receptor, thereby reducing DOR activity. Importantly, bradykinin stimulates GRK2 movement away from DOR and onto Raf kinase inhibitory protein (RKIP. protein kinase C (PKC-dependent RKIP phosphorylation induces GRK2 sequestration, restoring DOR functionality in sensory neurons. Together, these results expand the known function of GRK2, identifying a non-internalizing role to maintain peripheral DOR in an analgesically incompetent state.

  18. Dermorphin-related peptides from the skin of Phyllomedusa bicolor and their amidated analogs activate two mu opioid receptor subtypes that modulate antinociception and catalepsy in the rat.

    OpenAIRE

    Negri, L; Erspamer, G F; Severini, C; Potenza, R L; Melchiorri, P; Erspamer, V

    1992-01-01

    Three naturally occurring dermorphin-like peptides from the skin of the frog Phyllomedusa bicolor, the related carboxyl-terminal amides, and some substituted analogs were synthesized, their binding profiles to opioid receptors were determined, and their biological activities were studied in isolated organ preparations and intact animals. The opioid binding profile revealed a very high selectivity of these peptides for mu sites and suggested the existence of two receptor subtypes, of high and ...

  19. Non-analgesic effects of opioids: management of opioid-induced constipation by peripheral opioid receptor antagonists: prevention or withdrawal?

    Science.gov (United States)

    Holzer, Peter

    2012-01-01

    The therapeutic action of opioid analgesics is compromised by peripheral adverse effects among which opioid-induced constipation (OIC) is the most disabling, with a prevalence reported to vary between 15 and 90 %. Although OIC is usually treated with laxatives, there is insufficient clinical evidence that laxatives are efficacious in this indication. In contrast, there is ample evidence from double- blind, randomized and placebo-controlled trials that peripheral opioid receptor antagonists (PORAs) counteract OIC. This specific treatment modality is currently based on subcutaneous methylnaltrexone for the interruption of OIC in patients with advanced illness, and a fixed combination of oral prolonged-release naloxone with prolonged-release oxycodone for the prevention of OIC in the treatment of non-cancer and cancer pain. Both drugs counteract OIC while the analgesic effect of opioids remains unabated. The clinical studies show that more than 50 % of the patients with constipation under opioid therapy may benefit from the use of PORAs, while PORA-resistant patients are likely to suffer from non-opioid-induced constipation, the prevalence of which increases with age. While the addition of naloxone to oxycodone seems to act by preventing OIC, the intermittent dosing of methylnaltrexone every other day seems to stimulate defaecation by provoking an intestinal withdrawal response. The availability of PORAs provides a novel opportunity to specifically control OIC and other peripheral adverse effects of opioid analgesics (e.g., urinary retention and pruritus). The continuous dosing of a PORA has the advantage of few adverse effects, while intermittent dosing of a PORA can be associated with abdominal cramp-like pain.

  20. Peripheral δ-opioid receptors attenuate the exercise pressor reflex.

    Science.gov (United States)

    Leal, Anna K; Yamauchi, Katsuya; Kim, Joyce; Ruiz-Velasco, Victor; Kaufman, Marc P

    2013-10-15

    In rats with ligated femoral arteries, the exercise pressor reflex is exaggerated, an effect that is attenuated by stimulation of peripheral μ-opioid receptors on group IV metabosensitive afferents. In contrast, δ-opioid receptors are expressed mostly on group III mechanosensitive afferents, a finding that prompted us to determine whether stimulation of these opioid receptors could also attenuate the exaggerated exercise pressor reflex in "ligated" rats. We found femoral arterial injection of [D-Pen2,D-Pen5]enkephalin (DPDPE; 1.0 μg), a δ-opioid agonist, significantly attenuated the pressor and cardioaccelerator components of the exercise pressor reflex evoked by hindlimb muscle contraction in both rats with ligated and patent femoral arteries. DPDPE significantly decreased the pressor responses to muscle mechanoreflex activation, evoked by tendon stretch, in ligated rats only. DPDPE (1.0 μg) had no effect in either group on the pressor and cardioaccelerator responses to capsaicin (0.2 μg), which primarily stimulates group IV afferents. DPDPE (1.0 μg) had no effect on the pressor and cardioaccelerator responses to lactic acid (24 mM), which stimulates group III and IV afferents, in rats with patent femoral arteries but significantly decreased the pressor response in ligated rats. Western blots revealed the amount of protein comprising the δ-opioid receptor was greater in dorsal root ganglia innervating hindlimbs with ligated femoral arteries than in dorsal root ganglia innervating hindlimbs with patent femoral arteries. Our findings support the hypothesis that stimulation of δ-opioid receptors on group III afferents attenuated the exercise pressor reflex.

  1. Crystal structure of the[mu]-opioid receptor bound to a morphinan antagonist

    Energy Technology Data Exchange (ETDEWEB)

    Manglik, Aashish; Kruse, Andrew C.; Kobilka, Tong Sun; Thian, Foon Sun; Mathiesen, Jesper M.; Sunahara, Roger K.; Pardo, Leonardo; Weis, William I.; Kobilka, Brian K.; Granier, Sébastien (Michigan-Med); (Stanford-MED); (UAB, Spain)

    2012-06-27

    Opium is one of the world's oldest drugs, and its derivatives morphine and codeine are among the most used clinical drugs to relieve severe pain. These prototypical opioids produce analgesia as well as many undesirable side effects (sedation, apnoea and dependence) by binding to and activating the G-protein-coupled {mu}-opioid receptor ({mu}-OR) in the central nervous system. Here we describe the 2.8 {angstrom} crystal structure of the mouse {mu}-OR in complex with an irreversible morphinan antagonist. Compared to the buried binding pocket observed in most G-protein-coupled receptors published so far, the morphinan ligand binds deeply within a large solvent-exposed pocket. Of particular interest, the {mu}-OR crystallizes as a two-fold symmetrical dimer through a four-helix bundle motif formed by transmembrane segments 5 and 6. These high-resolution insights into opioid receptor structure will enable the application of structure-based approaches to develop better drugs for the management of pain and addiction.

  2. Methamphetamine-induced changes in the striatal dopamine pathway in μ-opioid receptor knockout mice

    Directory of Open Access Journals (Sweden)

    Park Sang Won

    2011-11-01

    Full Text Available Abstract Background Repeated exposure to methamphetamine (METH can cause not only neurotoxicity but also addiction. Behavioral sensitization is widely used as an animal model for the study of drug addiction. We previously reported that the μ-opioid receptor knockout mice were resistant to METH-induced behavioral sensitization but the mechanism is unknown. Methods The present study determined whether resistance of the μ-opioid receptor (μ-OR knockout mice to behavioral sensitization is due to differential expression of the stimulatory G protein α subunit (Gαs or regulators of G-protein signaling (RGS coupled to the dopamine D1 receptor. Mice received daily intraperitoneal injections of saline or METH (10 mg/kg for 7 consecutive days to induce sensitization. On day 11(following 4 abstinent days, mice were either given a test dose of METH (10 mg/kg for behavioral testing or sacrificed for neurochemical assays without additional METH treatment. Results METH challenge-induced stereotyped behaviors were significantly reduced in the μ-opioid receptor knockout mice when compared with those in wild-type mice. Neurochemical assays indicated that there is a decrease in dopamine D1 receptor ligand binding and an increase in the expression of RGS4 mRNA in the striatum of METH-treated μ-opioid receptor knockout mice but not of METH-treated wild-type mice. METH treatment had no effect on the expression of Gαs and RGS2 mRNA in the striatum of either strain of mice. Conclusions These results indicate that down-regulation of the expression of the dopamine D1 receptor and up-regulation of RGS4 mRNA expression in the striatum may contribute to the reduced response to METH-induced stereotypy behavior in μ-opioid receptor knockout mice. Our results highlight the interactions of the μ-opioid receptor system to METH-induced behavioral responses by influencing the expression of RGS of dopamine D1 receptors.

  3. [{sup 11}C]-MeJDTic: a novel radioligand for {kappa}-opioid receptor positron emission tomography imaging

    Energy Technology Data Exchange (ETDEWEB)

    Poisnel, Geraldine; Oueslati, Farhana; Dhilly, Martine; Delamare, Jerome [Groupe de Developpements Methodologiques en Tomographie par Emission de Positons, DSV/DRM UMR CEA 2E, Universite de Caen-Basse Normandie, Centre Cyceron, 14074 Caen Cedex (France); Perrio, Cecile [Groupe de Developpements Methodologiques en Tomographie par Emission de Positons, DSV/DRM UMR CEA 2E, Universite de Caen-Basse Normandie, Centre Cyceron, 14074 Caen Cedex (France)], E-mail: perrio@cyceron.fr; Debruyne, Daniele [Groupe de Developpements Methodologiques en Tomographie par Emission de Positons, DSV/DRM UMR CEA 2E, Universite de Caen-Basse Normandie, Centre Cyceron, 14074 Caen Cedex (France)], E-mail: debruyne@cyceron.fr; Barre, Louisa [Groupe de Developpements Methodologiques en Tomographie par Emission de Positons, DSV/DRM UMR CEA 2E, Universite de Caen-Basse Normandie, Centre Cyceron, 14074 Caen Cedex (France)

    2008-07-15

    Introduction: Radiopharmaceuticals that can bind selectively the {kappa}-opioid receptor may present opportunities for staging clinical brain disorders and evaluating the efficiency of new therapies related to stroke, neurodegenerative diseases or opiate addiction. The N-methylated derivative of JDTic (named MeJDTic), which has been recently described as a potent and selective antagonist of {kappa}-opioid receptor in vitro, was labeled with carbon-11 and evaluated for in vivo imaging the {kappa}-opioid receptor in mice. Methods: [{sup 11}C]-MeJDTic was prepared by methylation of JDTic with [{sup 11}C]-methyl triflate. The binding of [{sup 11}C]-MeJDTic to {kappa}-opioid receptor was investigated ex vivo by biodistribution and competition studies using nonfasted male CD1 mice. Results: [{sup 11}C]-MeJDTic exhibited a high and rapid distribution in peripheral organs. The uptake was maximal in lung where the {kappa} receptor is largely expressed. [{sup 11}C]-MeJDTic rapidly crossed the blood-brain barrier and accumulated in the brain regions of interest (hypothalamus). The parent ligand remained the major radioactive compound in brain during the experiment. Chase studies with U50,488 (a {kappa} referring agonist), morphine (a {mu} agonist) and naltrindole (a {delta} antagonist) demonstrated that this uptake was the result of specific binding to the {kappa}-opioid receptor. Conclusion: These findings suggested that [{sup 11}C]-MeJDTic appeared to be a promising selective 'lead' radioligand for {kappa}-opioid receptor PET imaging.

  4. Pharmacology of morphine and morphine-3-glucuronide at opioid, excitatory amino acid, GABA and glycine binding sites

    Energy Technology Data Exchange (ETDEWEB)

    Bartlett, S.E.; Smith, M.T. (Department of Pharmacy, The University of Queensland (Australia)); Dood, P.R. (Clinical Research Centre, Royal Brisbane Hospital Foundation, Brisbane (Australia))

    1994-07-01

    Morphine in high doses and its major metabolite, morphine-3-glucuronide, cause CNS excitation following intrathecal and intracerebroventricular administration by an unknown mechanism. This study investigated whether morphine and morphine-3-glucuronide interact at major excitatory (glutamate), major inhibitory (GABA or glycine), or opioid binding sites. Homogenate binding assays were performed using specific radioligands. At opioid receptors, morphine-3-glucuronide and morphine caused an equipotent sodium shift, consistent with morphine-3-glucuronide behaving as an agonist. This suggests that morphine-3-glucuronide-mediated excitation is not caused by an interaction at opioid receptors. Morphine-3-glucuronide and morphine caused a weak inhibition of the binding of [sup 3]H-MK801 (non-competitive antagonist) and [sup 125]I-ifenprodil (polyamine site antagonist), but at unphysiologically high concentrations. This suggests that CNS excitation would not result from an interaction of morphine-3-glucuronide and high-dose morphine with these sites on the NMDA receptor. Morphine-3-glucuronide and morphine inhibited the binding of [sup 3]H-muscimol (GABA receptor agonist), [sup 3]H-diazepam and [sup 3]H-flunitraxepam (benzodiazepine agonists) binding very weakly, suggesting the excitatory effects of morphine-3-glucuronide and high-dose morphine are not elicited through GABA[sub A] receptors. Morphine-3-glucuronide and high-dose morphine did not prevent re-uptake of glutamate into presynaptic nerve terminals. In addition, morphine-3-glucuronide and morphine did not inhibit the binding of [sup 3]H-strychnine (glycine receptor antagonist) to synaptic membranes prepared from bovine spinal cord. It is concluded that excitation caused by high-dose morphine and morphine-3-glucuronide is not mediated by an interaction with postsynaptic amino acid receptors. (au) (30 refs.).

  5. Sodium modulates opioid receptors through a membrane component different from G-proteins. Demonstration by target size analysis

    International Nuclear Information System (INIS)

    Ott, S.; Costa, T.; Herz, A.

    1988-01-01

    The target size for opioid receptor binding was studied after manipulations known to affect the interactions between receptor and GTP-binding regulatory proteins (G-proteins). Addition of GTP or its analogs to the binding reaction, exposure of intact cells to pertussis toxin prior to irradiation, or treatment of irradiated membranes with N-ethylmaleimide did not change the target size (approximately equal to 100 kDa) for opioid receptors in NG 108-15 cells and rat brain. These data suggest that the 100-kDa species does not include an active subunit of a G-protein or alternatively that GTP does not promote the dissociation of the receptor-G-protein complex. The presence of Na+ (100 mM) in the radioligand binding assay induced a biphasic decay curve for agonist binding and a flattening of the monoexponential decay curve for a partial agonist. In both cases the effect was explained by an irradiation-induced loss of the low affinity state of the opioid receptor produced by the addition of Na+. This suggests that an allosteric inhibitor that mediates the effect of sodium on the receptor is destroyed at low doses of irradiation, leaving receptors which are no longer regulated by sodium. The effect of Na+ on target size was slightly increased by the simultaneous addition of GTP but was not altered by pertussis toxin treatment. Thus, the sodium unit is distinct from G-proteins and may represent a new component of the opioid receptor complex. Assuming a simple bimolecular model of one Na+ unit/receptor, the size of this inhibitor can be measured as 168 kDa

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

    Science.gov (United States)

    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; Patel, Hemal H

    2010-11-01

    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 NH(2)-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

  7. 3D-QSAR comparative molecular field analysis on opioid receptor antagonists: pooling data from different studies.

    Science.gov (United States)

    Peng, Youyi; Keenan, Susan M; Zhang, Qiang; Kholodovych, Vladyslav; Welsh, William J

    2005-03-10

    Three-dimensional quantitative structure-activity relationship (3D-QSAR) models were constructed using comparative molecular field analysis (CoMFA) on a series of opioid receptor antagonists. To obtain statistically significant and robust CoMFA models, a sizable data set of naltrindole and naltrexone analogues was assembled by pooling biological and structural data from independent studies. A process of "leave one data set out", similar to the traditional "leave one out" cross-validation procedure employed in partial least squares (PLS) analysis, was utilized to study the feasibility of pooling data in the present case. These studies indicate that our approach yields statistically significant and highly predictive CoMFA models from the pooled data set of delta, mu, and kappa opioid receptor antagonists. All models showed excellent internal predictability and self-consistency: q(2) = 0.69/r(2) = 0.91 (delta), q(2) = 0.67/r(2) = 0.92 (mu), and q(2) = 0.60/r(2) = 0.96 (kappa). The CoMFA models were further validated using two separate test sets: one test set was selected randomly from the pooled data set, while the other test set was retrieved from other published sources. The overall excellent agreement between CoMFA-predicted and experimental binding affinities for a structurally diverse array of ligands across all three opioid receptor subtypes gives testimony to the superb predictive power of these models. CoMFA field analysis demonstrated that the variations in binding affinity of opioid antagonists are dominated by steric rather than electrostatic interactions with the three opioid receptor binding sites. The CoMFA steric-electrostatic contour maps corresponding to the delta, mu, and kappa opioid receptor subtypes reflected the characteristic similarities and differences in the familiar "message-address" concept of opioid receptor ligands. Structural modifications to increase selectivity for the delta over mu and kappa opioid receptors have been predicted on the

  8. Somatostatin and opioid receptors do not regulate proliferation or apoptosis of the human multiple myeloma U266 cells

    Directory of Open Access Journals (Sweden)

    Allouche Stéphane

    2009-06-01

    Full Text Available Abstract Background opioid and somatostatin receptors (SSTRs that can assemble as heterodimer were individually reported to modulate malignant cell proliferation and to favour apoptosis. Materials and methods: SSTRs and opioid receptors expression were examined by RT-PCR, western-blot and binding assays, cell proliferation was studied by XTT assay and propidium iodide (PI staining and apoptosis by annexin V-PI labelling. Results almost all human malignant haematological cell lines studied here expressed the five SSTRs. Further experiments were conducted on the human U266 multiple myeloma cells, which express also μ-opioid receptors (MOP-R. XTT assays and cell cycle studies provide no evidence for a significant effect upon opioid or somatostatin receptors stimulation. Furthermore, neither direct effect nor potentiation of the Fas-receptor pathway was detected on apoptosis after these treatments. Conclusion these data suggest that SSTRs or opioid receptors expression is not a guaranty for an anti-tumoral action in U266 cell line.

  9. Atypical Opioid Mechanisms of Control of Injury-Induced Cutaneous Pain by Delta Receptors

    Science.gov (United States)

    2016-07-01

    treat, and current opioids (i.e. mu opioid receptor agonists such as morphine) cause unacceptable side effects including addiction . Injuries suffered...treat, and current opioids that act on mu opioid receptors such as morphine generate significant side effects including addiction . War-related...al., J Neurosci Methods, 1994), starting with 0.1 g and ending with 2.0 g filament as cutoff value. As shown in Figure 2, our preliminary experiments

  10. Enhanced efficacy (intrinsic activity) of cyclic opioid peptide analogs at the μ-receptor

    International Nuclear Information System (INIS)

    Schiller, P.W.; Lemieux, C.; Nguyen, T.M.D.; Maziak, L.A.

    1986-01-01

    Side-chain to end group cyclized enkephalin analogs (e.g. H-Tyr-cyclo[-D-Lys-Gly-Phe-Leu-] and cyclic opioid peptide analogs obtained through covalent linkage of two side-chains (e.g. H-Tyr-D-Cys-Gly-Phe-Cys-NH 2 or H-Tyr-D-Lys-Gly-Phe-Glu-NH 3 ) were tested in the μ-receptor-representative guinea pig ileum (GPI) bioassay and in a binding assay based on displacement of the μ-ligand [ 3 H]DAGO from rat brain membranes. The cyclic analogs were 5 to 70 times more potent in the GPI assay than in the binding assay, whereas linear analogs showed equal potency in the two assays. These results suggest that the efficacy (intrinsic activity) of cyclic opioid peptide analogs at the μ-receptor is increased as a consequence of the conformation constraint imposed through ring closure. This effect was most pronounced in analogs containing a long hydrophobic sidechain as part of the ring structure in the 2-position of the peptide sequence. Further experimental evidence ruled out the possibilities that these potency discrepancies may be due to differences in enzymatic degradation, dissimilar exposure of the receptors in their lipid environment or interaction with different receptor types in the two assay systems. It can be hypothesized that the semi-rigid cyclic analogs may induce a more productive conformational change in the receptor protein than the linear peptides

  11. Identification of rat brain opioid (enkephalin) receptor by photoaffinity labeling

    International Nuclear Information System (INIS)

    Yeung, C.W.

    1986-01-01

    A photoreactive, radioactive enkephalin derivative was prepared and purified by high performance liquid chromatography. Rat brain and spinal cord plasma membranes were incubated with this radioiodinated photoprobe and were subsequently photolysed. Autoradiography of the sodium dodecyl sulfate gel electrophoresis of the solubilized and reduced membranes showed that a protein having an apparent molecular weight of 46,000 daltons was specifically labeled, suggesting that this protein may be the opioid (enkephalin) receptor

  12. Effect of Iboga alkaloids on µ-opioid receptor-coupled G protein activation.

    Directory of Open Access Journals (Sweden)

    Tamara Antonio

    Full Text Available The iboga alkaloids are a class of small molecules defined structurally on the basis of a common ibogamine skeleton, some of which modify opioid withdrawal and drug self-administration in humans and preclinical models. These compounds may represent an innovative approach to neurobiological investigation and development of addiction pharmacotherapy. In particular, the use of the prototypic iboga alkaloid ibogaine for opioid detoxification in humans raises the question of whether its effect is mediated by an opioid agonist action, or if it represents alternative and possibly novel mechanism of action. The aim of this study was to independently replicate and extend evidence regarding the activation of μ-opioid receptor (MOR-related G proteins by iboga alkaloids.Ibogaine, its major metabolite noribogaine, and 18-methoxycoronaridine (18-MC, a synthetic congener, were evaluated by agonist-stimulated guanosine-5´-O-(γ-thio-triphosphate ([(35S]GTPγS binding in cells overexpressing the recombinant MOR, in rat thalamic membranes, and autoradiography in rat brain slices.In rat thalamic membranes ibogaine, noribogaine and 18-MC were MOR antagonists with functional Ke values ranging from 3 uM (ibogaine to 13 uM (noribogaine and 18MC. Noribogaine and 18-MC did not stimulate [(35S]GTPγS binding in Chinese hamster ovary cells expressing human or rat MORs, and had only limited partial agonist effects in human embryonic kidney cells expressing mouse MORs. Ibogaine did not did not stimulate [(35S]GTPγS binding in any MOR expressing cells. Noribogaine did not stimulate [(35S]GTPγS binding in brain slices using autoradiography. An MOR agonist action does not appear to account for the effect of these iboga alkaloids on opioid withdrawal. Taken together with existing evidence that their mechanism of action also differs from that of other non-opioids with clinical effects on opioid tolerance and withdrawal, these findings suggest a novel mechanism of action, and

  13. Does the kappa opioid receptor system contribute to pain aversion?

    Directory of Open Access Journals (Sweden)

    Catherine M Cahill

    2014-11-01

    Full Text Available The kappa opioid receptor (KOR and the endogenous peptide-ligand dynorphin have received significant attention due the involvement in mediating a variety of behavioral and neurophysiological responses, including opposing the rewarding properties of drugs of abuse including opioids. Accumulating evidence indicates this system is involved in regulating states of motivation and emotion. Acute activation of the KOR produces an increase in motivational behavior to escape a threat, however, KOR activation associated with chronic stress leads to the expression of symptoms indicative of mood disorders. It is well accepted that KOR can produce analgesia and is engaged in chronic pain states including neuropathic pain. Spinal studies have revealed KOR-induced analgesia in reversing pain hypersensitivities associated with peripheral nerve injury. While systemic administration of KOR agonists attenuates nociceptive sensory transmission, this effect appears to be a stress-induced effect as anxiolytic agents, including delta opioid receptor agonists, mitigate KOR agonist-induced analgesia. Additionally, while the role of KOR and dynorphin in driving the dysphoric and aversive components of stress and drug withdrawal has been well characterized, how this system mediates the negative emotional states associated with chronic pain is relatively unexplored. This review provides evidence that dynorphin and the KOR system contribute to the negative affective component of pain and that this receptor system likely contributes to the high comorbidity of mood disorders associated with chronic neuropathic pain.

  14. Characterization of kappa opioid binding using dynorphin A1-13 and U69,593 in the rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Devlin, T.; Shoemaker, W.J. (Univ. of Connecticut Health Center, Farmington (USA))

    1990-05-01

    Previous studies of kappa opioid binding sites have suggested heterogeneous binding to this class of opioid receptors. To further investigate kappa receptor heterogeneity, we analyzed the binding properties of various kappa-selective ligands in rat brain homogenates. Displacement assays were carried out using (3H)bremazocine in the presence of various displacing ligands under mu and delta receptor-blocked conditions. Homologous displacement of (3H)bremazocine produced shallow displacement which best fit a two-site model of drug-receptor interaction. Dynorphin A1-13 and U69,593 exhibited similar biphasic displacement of (3H)bremazocine. Maximal displacement by these ligands, however, represented only approximately 55% of total (3H)bremazocine binding, which suggests the existence of a third component of (3H)bremazocine binding. Biphasic displacement by dynorphin A1-13 was detected in tissue throughout the brain and the spinal cord, whereas the dynorphin-resistant component of (3H)bremazocine binding was uniquely absent in the spinal cord. U50,488H, tifluadom and ethylketocyclazocine appeared to displace from additional, dynorphin-insensitive sites, as their maximal displacement exceeded that seen with either dynorphin A1-13 or U69,593. These results strongly suggest the existence of at least three components of non-mu, non-delta (3H)bremazocine binding in the rat brain: two with differential affinity for dynorphin A1-13 and U69-593 (kappa-1 and kappa-2 sites), and a third (termed here R1) that was further resolved into two binding sites by bremazocine. Preliminary analysis of the R1 component using naloxone revealed one high-affinity site, which may be opiate in nature, and a second site whose binding properties closely resemble those of the sigma receptor described by others.

  15. A kinetic analysis of kappa-opioid agonist binding using the selective radioligand (/sup 3/H)U69593

    Energy Technology Data Exchange (ETDEWEB)

    Smith, J.A.; Hunter, J.C.; Hill, R.G.; Hughes, J.

    1989-07-01

    The interaction of the nonselective opioid ligand (3H)bremazocine and of the kappa-opioid (3H)U69593 with the kappa-receptor was investigated in guinea-pig cortical membranes. Each radioligand bound to a single population of high-affinity sites, although (3H)U69593 apparently recognised only 70% of those sites labelled by (3H)bremazocine. Naloxone and the kappa-selective ligands U69593 and PD117302 exhibited full inhibition of the binding of both radioligands. Kinetic analysis demonstrated biphasic rates of association and dissociation for both (3H)bremazocine and (3H)U69593. Detailed analysis of the binding of (3H)U69593 revealed that the fast rate of association was dependent on radioligand concentration, in contrast to the slow rate, which was independent of ligand concentration. Guanylyl-5'-imidodiphosphate (GppNHp) inhibited binding of (3H)U69593; saturation analysis demonstrated that the inhibitory effects of GppNHp resulted in a decrease in affinity without any significant change in binding capacity. GppNHp attenuated the formation of the slow component of (3H)U69593 binding, while accelerating the fast component. The data are consistent with the formation of a high-affinity complex between the kappa-receptor and a guanine nucleotide binding protein. Guanine nucleotides promote the dissociation of this ternary complex and the stabilisation of a lower-affinity state of the receptor.

  16. Characterization of kappa 1 and kappa 2 opioid binding sites in frog (Rana esculenta) brain membrane preparation

    Energy Technology Data Exchange (ETDEWEB)

    Benyhe, S.; Varga, E.; Hepp, J.; Magyar, A.; Borsodi, A.; Wollemann, M.

    1990-09-01

    The distribution and properties of frog brain kappa-opioid receptor subtypes differ not only from those of the guinea pig brain, but also from that of the rat brain. In guinea pig cerebellum the kappa 1 is the dominant receptor subtype, frog brain contains mainly the kappa 2 subtype, and the distribution of the rat brain subtypes is intermediate between the two others. In competition experiments it has been established that ethylketocyclazocine and N-cyclopropylmethyl-norazidomorphine, which are nonselective kappa-ligands, have relatively high affinities to frog brain membranes. The kappa 2 ligands (Met5)enkephalin-Arg6-Phe7 and etorphine also show high affinities to the frog brain. Kappa 1 binding sites measured in the presence of 5 microM/D-Ala2-Leu5/enkephalin represent 25-30% of (3H)ethylketocyclazocine binding in frog brain membranes. The kappa 2 subtype in frog brain resembles more to the mu subtype than the delta subtype of opioid receptors, but it differs from the mu subtype in displaying low affinity toward beta-endorphin and /D-Ala2-(Me)Phe4-Gly5-ol/enkephalin (DAGO). From our data it is evident that the opioid receptor subtypes are already present in the amphibian brain but the differences among them are less pronounced than in mammalian brain.

  17. Characterization of kappa 1 and kappa 2 opioid binding sites in frog (Rana esculenta) brain membrane preparation

    International Nuclear Information System (INIS)

    Benyhe, S.; Varga, E.; Hepp, J.; Magyar, A.; Borsodi, A.; Wollemann, M.

    1990-01-01

    The distribution and properties of frog brain kappa-opioid receptor subtypes differ not only from those of the guinea pig brain, but also from that of the rat brain. In guinea pig cerebellum the kappa 1 is the dominant receptor subtype, frog brain contains mainly the kappa 2 subtype, and the distribution of the rat brain subtypes is intermediate between the two others. In competition experiments it has been established that ethylketocyclazocine and N-cyclopropylmethyl-norazidomorphine, which are nonselective kappa-ligands, have relatively high affinities to frog brain membranes. The kappa 2 ligands (Met5)enkephalin-Arg6-Phe7 and etorphine also show high affinities to the frog brain. Kappa 1 binding sites measured in the presence of 5 microM/D-Ala2-Leu5/enkephalin represent 25-30% of [3H]ethylketocyclazocine binding in frog brain membranes. The kappa 2 subtype in frog brain resembles more to the mu subtype than the delta subtype of opioid receptors, but it differs from the mu subtype in displaying low affinity toward beta-endorphin and /D-Ala2-(Me)Phe4-Gly5-ol/enkephalin (DAGO). From our data it is evident that the opioid receptor subtypes are already present in the amphibian brain but the differences among them are less pronounced than in mammalian brain

  18. CARBOHYDRATE-CONTAINING COMPOUNDS WHICH BIND TO CARBOHYDRATE BINDING RECEPTORS

    DEFF Research Database (Denmark)

    1995-01-01

    Carbohydrate-containing compounds which contain saccharides or derivatives thereof and which bind to carbohydrate binding receptors are useful in pharmaceutical products for treatment of inflammatory diseases and other diseases.......Carbohydrate-containing compounds which contain saccharides or derivatives thereof and which bind to carbohydrate binding receptors are useful in pharmaceutical products for treatment of inflammatory diseases and other diseases....

  19. Development of concepts on the interaction of drugs with opioid receptors

    Energy Technology Data Exchange (ETDEWEB)

    Kuzmina, N E; Kuzmin, V S

    2011-02-28

    The development of concepts on the molecular mechanisms of the action of medicinal drugs on the opioid receptors is briefly surveyed. The modern point of view on the mechanism of activation of opioid receptors is given based on the data from chimeric and site-directed mutagenesis of the cloned opioid receptors and the computer-aided simulations of the reception zone and ligand-receptor complexes. Three-dimensional models of the opioid pharmacophore derived by both conventional methods and a comparative analysis of molecular fields are described in detail.

  20. Opioid mediated activity and expression of mu and delta opioid receptors in isolated human term non-labouring myometrium.

    LENUS (Irish Health Repository)

    Fanning, Rebecca A

    2013-01-05

    The existence of opioid receptors in mammalian myometrial tissue is now widely accepted. Previously enkephalin degrading enzymes have been shown to be elevated in pregnant rat uterus and a met-enkephalin analogue has been shown to alter spontaneous contractility of rat myometrium. Here we have undertaken studies to determine the effects of met-enkephalin on in vitro human myometrial contractility and investigate the expression of opioid receptors in pregnant myometrium. Myometrial biopsies were taken from women undergoing elective caesarean delivery at term. Organ bath experiments were used to investigate the effect of the met-enkephalin analogue [d-Ala 2, d-met 5] enkephalin (DAMEA) on spontaneous contractility. A confocal immunofluorescent technique and real time PCR were used to determine the expression of protein and mRNA, respectively for two opioid receptor subtypes, mu and delta. DAMEA had a concentration dependent inhibitory effect on contractile activity (1 × 10(-7)M-1 × 10(-4)M; 54% reduction in contractile activity, P<0.001 at 1 × 10(-4)M concentration). Mu and delta opioid receptor protein sub-types and their respective mRNA were identified in all tissues sampled. This is the first report of opioid receptor expression and of an opioid mediated uterorelaxant action in term human non-labouring myometrium in vitro.

  1. Binge eating disorder and morbid obesity are associated with lowered mu-opioid receptor availability in the brain.

    Science.gov (United States)

    Joutsa, Juho; Karlsson, Henry K; Majuri, Joonas; Nuutila, Pirjo; Helin, Semi; Kaasinen, Valtteri; Nummenmaa, Lauri

    2018-03-09

    Both morbid obesity and binge eating disorder (BED) have previously been linked with aberrant brain opioid function. Behaviorally these two conditions are however different suggesting also differences in neurotransmitter function. Here we directly compared mu-opioid receptor (MOR) availability between morbidly obese and BED subjects. Seven BED and nineteen morbidly obese (non-BED) patients, and thirty matched control subjects underwent positron emission tomography (PET) with MOR-specific ligand [ 11 C]carfentanil. Both subjects with morbid obesity and BED had widespread reduction in [ 11 C]carfentanil binding compared to control subjects. However, there was no significant difference in brain MOR binding between subjects with morbid obesity and BED. Thus, our results indicate that there is common brain opioid abnormality in behaviorally different eating disorders involving obesity. Copyright © 2018 Elsevier B.V. All rights reserved.

  2. Radioreceptor opioid assay

    International Nuclear Information System (INIS)

    Miller, R.J.; Chang, K.-J.

    1981-01-01

    A radioreceptor assay is described for assaying opioid drugs in biological fluids. The method enables the assay of total opioid activity, being specific for opioids as a class but lacking specificity within the class. A radio-iodinated opioid and the liquid test sample are incubated with an opiate receptor material. The percentage inhibition of the binding of the radio-iodinated compound to the opiate receptor is calculated and the opioid activity of the test liquid determined from a standard curve. Examples of preparing radio-iodinated opioids and assaying opioid activity are given. A test kit for the assay is described. Compared to other methods, this assay is cheap, easy and rapid. (U.K.)

  3. Differential regulation of. mu. , delta, kappa opioid receptors by Mn/sup + +/

    Energy Technology Data Exchange (ETDEWEB)

    Szuecs, M.; Oetting, G.M.; Coscia, C.J.

    1986-03-05

    Differential effects of Mn/sup + +/ on three opioid receptor subtypes of rat brain membranes were evaluated. Concentration dependency studies performed with 0.05-20 mM Mn/sup + +/ revealed that only the delta receptors are stimulated at any concentration. The binding of 1 nM /sup 3/H-DAGO was not stimulated by low concentrations (< 1mM) of Mn/sup + +/, and was significantly inhibited at higher concentrations (40% at 20 mM). 1 nM /sup 3/H-EKC (+100nM DAGO and 100nM DADLE) binding was inhibited by Mn/sup + +/ in the entire concentration range. While regulation of ..mu.. receptor binding did not change during postnatal development, delta and kappa binding displayed a pronounced developmental time-dependency. Kappa sites were hardly affected by Mn/sup + +/ at day 5, and adult levels of inhibition were reached only after the third week postnatal. In contrast, 1 nM /sup 3/H-DADLE (+10nM DAGO) binding was most sensitive to Mn/sup + +/ on day 5 after birth (100% stimulation with 5-20 mM). The ED/sub 50/ of Mn/sup + +/ stimulation was unchanged during maturation. These immature delta sites displayed a similar extent of Mn/sup + +/ reversal of Gpp(NH)p inhibition as seen in microsomes, which represent a good model of N/sub i/-uncoupled receptors. These data suggest that ..mu.., delta and kappa receptors are differently coupled to N/sub i/. Moreover, a second divalent cation binding site, in addition to that on N/sub i/ might exist for delta receptors.

  4. Opioid withdrawal increases transient receptor potential vanilloid 1 activity in a protein kinase A-dependent manner.

    Science.gov (United States)

    Spahn, Viola; Fischer, Oliver; Endres-Becker, Jeannette; Schäfer, Michael; Stein, Christoph; Zöllner, Christian

    2013-04-01

    Hyperalgesia is a cardinal symptom of opioid withdrawal. The transient receptor potential vanilloid 1 (TRPV1) is a ligand-gated ion channel expressed on sensory neurons responding to noxious heat, protons, and chemical stimuli such as capsaicin. TRPV1 can be inhibited via μ-opioid receptor (MOR)-mediated reduced activity of adenylyl cyclases (ACs) and decreased cyclic adenosine monophosphate (cAMP) levels. In contrast, opioid withdrawal following chronic activation of MOR uncovers AC superactivation and subsequent increases in cAMP and protein kinase A (PKA) activity. Here we investigated (1) whether an increase in cAMP during opioid withdrawal increases the activity of TRPV1 and (2) how opioid withdrawal modulates capsaicin-induced nocifensive behavior in rats. We applied whole-cell patch clamp, microfluorimetry, cAMP assays, radioligand binding, site-directed mutagenesis, and behavioral experiments. Opioid withdrawal significantly increased cAMP levels and capsaicin-induced TRPV1 activity in both transfected human embryonic kidney 293 cells and dissociated dorsal root ganglion (DRG) neurons. Inhibition of AC and PKA, as well as mutations of the PKA phosphorylation sites threonine 144 and serine 774, prevented the enhanced TRPV1 activity. Finally, capsaicin-induced nocifensive behavior was increased during opioid withdrawal in vivo. In summary, our results demonstrate an increased activity of TRPV1 in DRG neurons as a new mechanism contributing to opioid withdrawal-induced hyperalgesia. Copyright © 2013 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  5. Deltorphins: a family of naturally occurring peptides with high affinity and selectivity for delta opioid binding sites.

    Science.gov (United States)

    Erspamer, V; Melchiorri, P; Falconieri-Erspamer, G; Negri, L; Corsi, R; Severini, C; Barra, D; Simmaco, M; Kreil, G

    1989-07-01

    Deltorphins are endogenous linear heptapeptides, isolated from skin extracts of frogs belonging to the genus Phyllomedusa, that have a higher affinity and selectivity for delta opioid binding sites than any other natural compound known. Two deltorphins with the sequence Tyr-Ala-Phe-Asp(or Glu)-Val-Val-Gly-NH2 have been isolated from skin extracts of Phyllomedusa bicolor. The alanine in position 2 is in the D configuration. These peptides, [D-Ala2]deltorphins I and II, show an even higher affinity for delta receptors than the previously characterized deltorphin, which contains D-methionine as the second amino acid. These peptides show some similarity to another constituent of Phyllomedusa skin, dermorphin, which is highly selective for mu-opioid receptors. These peptides all have the N-terminal sequence Tyr-D-Xaa-Phe, where D-Xaa is either D-alanine or D-methionine. While this structure seems to be capable of activating both mu and delta opioid receptors, differences in the C-terminal regions of these peptides are probably responsible for the observed high receptor selectivity of dermorphin and deltorphin.

  6. Mu-opioid receptor knockout mice show diminished food-anticipatory activity

    NARCIS (Netherlands)

    Kas, Martien J H; van den Bos, Ruud; Baars, Annemarie M; Lubbers, Marianne; Lesscher, Heidi M B; Hillebrand, Jacquelien J G; Schuller, Alwin G; Pintar, John E; Spruijt, Berry M

    We have previously suggested that during or prior to activation of anticipatory behaviour to a coming reward, mu-opioid receptors are activated. To test this hypothesis schedule induced food-anticipatory activity in mu-opioid receptor knockout mice was measured using running wheels. We hypothesized

  7. Expression and purification of functional human mu opioid receptor from E.coli.

    Directory of Open Access Journals (Sweden)

    Yanbin Ma

    Full Text Available N-terminally his-tagged human mu opioid receptor, a G protein-coupled receptor was produced in E.coli employing synthetic codon-usage optimized constructs. The receptor was expressed in inclusion bodies and membrane-inserted in different E.coli strains. By optimizing the expression conditions the expression level for the membrane-integrated receptor was raised to 0.3-0.5 mg per liter of culture. Milligram quantities of receptor could be enriched by affinity chromatography from IPTG induced cultures grown at 18°C. By size exclusion chromatography the protein fraction with the fraction of alpha-helical secondary structure expected for a 7-TM receptor was isolated, by CD-spectroscopy an alpha-helical content of ca. 45% was found for protein solubilised in the detergent Fos-12. Receptor in Fos-12 micelles was shown to bind endomorphin-1 with a K(D of 61 nM. A final yield of 0.17 mg functional protein per liter of culture was obtained.

  8. N1'-fluoroethyl-naltrindole (BU97001) and N1'-fluoroethyl-(14-formylamino)-naltrindole (BU97018) potential δ-opioid receptor PET ligands

    International Nuclear Information System (INIS)

    Tyacke, Robin J.; Robinson, Emma S.J.; Schnabel, Rebecca; Lewis, John W.; Husbands, Stephen M.; Nutt, David J.; Hudson, Alan L.

    2002-01-01

    The properties of two prospective positron emission tomography (PET) ligands for the δ-opioid receptor, N1'-fluoroethyl-naltrindole (BU97001) and N1'-fluoroethyl-(14-formylamino)-naltrindole (BU97018) were investigated. Both were antagonists in the mouse vas deferens, and showed high affinity and selectivity, 1.81 nM and 3.09 nM respectively. [ 3 H]BU97001 binding to rat whole brain was also of high affinity, K D of 0.42 nM of and B MAX of 59.95 fmol mg of protein -1 . In autoradiographic studies, it was found to bind to brain areas previously shown to be associated with the δ-opioid receptor and good correlations were found to exist with naltrindole and DPDPE. BU97018 and especially BU97001 appear to show good potential as δ-opioid receptor PET ligands with the incorporation of 18 F

  9. Novel approaches for the treatment of psychostimulant and opioid abuse - focus on opioid receptor-based therapies.

    Science.gov (United States)

    Bailey, Chris P; Husbands, Stephen M

    2014-11-01

    Psychostimulant and opioid addiction are poorly treated. The majority of abstinent users relapse back to drug-taking within a year of abstinence, making 'anti-relapse' therapies the focus of much current research. There are two fundamental challenges to developing novel treatments for drug addiction. First, there are three key stimuli that precipitate relapse back to drug-taking: stress, presentation of drug-conditioned cue, taking a small dose of drug. The most successful novel treatment would be effective against all three stimuli. Second, a large number of drug users are poly-drug users: taking more than one drug of abuse at a time. The ideal anti-addiction treatment would, therefore, be effective against all classes of drugs of abuse. In this review, the authors discuss the clinical need and animal models used to uncover potential novel treatments. There is a very broad range of potential treatment approaches and targets currently being examined as potential anti-relapse therapies. These broadly fit into two categories: 'memory-based' and 'receptor-based' and the authors discuss the key targets here within. Opioid receptors and ligands have been widely studied, and research into how different opioid subtypes affect behaviours related to addiction (reward, dysphoria, motivation) suggests that they are tractable targets as anti-relapse treatments. Regarding opioid ligands as novel 'anti-relapse' medication targets, research suggests that a 'non-selective' approach to targeting opioid receptors will be the most effective.

  10. Autoradiographic localization of mu and delta opioid receptors in the mesocorticolimbic dopamine system

    Energy Technology Data Exchange (ETDEWEB)

    Dilts, R.P. Jr.

    1989-01-01

    In vitro autoradiographic techniques were coupled with selective chemical lesions of the A10 dopamine cells and intrinsic perikarya of the region to delineate the anatomical localization of mu and delta opioid receptors, as well as, neurotensin receptors. Mu opioid receptors were labeled with {sup 125}I-DAGO. Delta receptors were labeled with {sup 125}I-DPDPE. Neurotensin receptors were labeled with {sup 125}I-NT3. Unilateral lesions of the dopamine perikarya were produced by injections of 6-OHDA administered in the ventral mesencephalon. Unilateral lesions of intrinsic perikarya were induced by injections of quinolinic acid in to the A10 dopamine cell region. Unilateral lesions produced with 6-OHDA resulted in the loss of neurotensin receptors in the A10 region and within the terminal fields. Mu opioid receptors were unaffected by this treatment, but delta opioid receptors increased in the contralateral striatum and nucleus accumbens following 6-OHDA administration. Quinolinic acid produced a reduction of mu opioid receptors within the A10 region with a concomitant reduction in neurotensin receptors in both the cell body region and terminal fields. These results are consistent with a variety of biochemical and behavioral data which suggest the indirect modulation of dopamine transmission by the opioids. In contrast these results strongly indicate a direct modulation of the mesolimbic dopamine system by neurotensin.

  11. [Effect of opioid receptors on acute stress-induced changes in recognition memory].

    Science.gov (United States)

    Liu, Ying; Wu, Yu-Wei; Qian, Zhao-Qiang; Yan, Cai-Fang; Fan, Ka-Min; Xu, Jin-Hui; Li, Xiao; Liu, Zhi-Qiang

    2016-12-25

    Although ample evidence has shown that acute stress impairs memory, the influences of acute stress on different phases of memory, such as acquisition, consolidation and retrieval, are different. Experimental data from both human and animals support that endogenous opioid system plays a role in stress, as endogenous opioid release is increased and opioid receptors are activated during stress experience. On the other hand, endogenous opioid system mediates learning and memory. The aim of the present study was to investigate the effect of acute forced swimming stress on recognition memory of C57 mice and the role of opioid receptors in this process by using a three-day pattern of new object recognition task. The results showed that 15-min acute forced swimming damaged the retrieval of recognition memory, but had no effect on acquisition and consolidation of recognition memory. No significant change of object recognition memory was found in mice that were given naloxone, an opioid receptor antagonist, by intraperitoneal injection. But intraperitoneal injection of naloxone before forced swimming stress could inhibit the impairment of recognition memory retrieval caused by forced swimming stress. The results of real-time PCR showed that acute forced swimming decreased the μ opioid receptor mRNA levels in whole brain and hippocampus, while the injection of naloxone before stress could reverse this change. These results suggest that acute stress may impair recognition memory retrieval via opioid receptors.

  12. Rapid agonist-induced loss of sup 125 I-. beta. -endorphin opioid receptor sites in NG108-15, but not SK-N-SH neuroblastoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Cone, R.I.; Lameh, J.; Sadee, W. (Univ. of California, San Francisco (United States))

    1991-01-01

    The authors have measured {mu} and {delta} opioid receptor sites on intact SK-N-SH and NG108-15 neuroblastoma cells, respectively, in culture. Use of {sup 125}I-{beta}-endorphin ({beta}E) as a tracer, together with {beta}E(6-31) to block high-affinity non-opioid binding in both cell lines, permitted the measurement of cell surface {mu} and {delta} opioid receptor sites. Labeling was at {delta} sites in NG108-15 cells and predominantly at {mu} sites in SK-N-SH cells. Pretreatment with the {mu} and {delta} agonist, DADLE, caused a rapid loss of cell surface {delta} receptor sites in NG108-15 cells, but failed to reduce significantly {mu} receptor density in SK-N-SH cells.

  13. Long-lasting, distinct changes in central opioid receptor and urinary bladder functions in models of schizophrenia in rats.

    Science.gov (United States)

    Kekesi, Orsolya; Tuboly, Gabor; Szucs, Maria; Birkas, Erika; Morvay, Zita; Benedek, Gyorgy; Horvath, Gyongyi

    2011-07-01

    Ketamine treatments and social isolation of rats reflect certain features of schizophrenia, among them altered pain sensitivity. To study the underlying mechanisms of these phenomena, rats were either housed individually or grouped for 33 days after weaning, and treated with either ketamine or saline for 14 days. After one month re-socialization, the urinary bladder capacity by ultrasound examination in the anesthetized animals, and changes of μ-opioid receptors by saturation binding experiments using a specific μ-opioid agonist [(3)H]DAMGO were determined. G-protein signaling was investigated in DAMGO-stimulated [(35)S]GTPγS functional assays. Ketamine treatment significantly decreased the bladder volume and isolation decreased the receptor density in cortical membranes. Among all groups, the only change in binding affinity was an increase induced by social isolation in the cortex. G-protein signaling was significantly decreased by either ketamine or social isolation in this tissue. Ketamine treatment, but not housing, significantly increased μ-opioid receptor densities in hippocampal membranes. Both ketamine and isolation increased the efficacy, while the potency of signaling was decreased by any treatment. Ketamine increased the receptor density and G-protein activation; while isolation decreased the efficacy of G-protein signaling in hippocampal membranes. The changes in the co-treated group were similar to those of the isolated animals in most tests. The distinct changes of opioid receptor functioning in different areas of the CNS may, at least partially, explain the augmented nociceptive threshold and morphine potency observed in these animals. Changes in the relative urinary bladder suggest a detrusor hyperreflexia, another sign of schizophrenia. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Identification of endogenous opioid receptor components in rat brain using a monoclonal antibody

    Energy Technology Data Exchange (ETDEWEB)

    Bero, L.A.; Roy, S.; Lee, N.M.

    1988-11-01

    A monoclonal antibody generated against the tertiary structure of a partially purified opioid binding protein was used to probe the structure of the dynorphin and beta-endorphin receptors. The Fab fragment 3B4F11 inhibited completely the binding of 125I-beta-endorphin and (3H)dynorphin to rat brain P2 membranes with IC50 values of 26 ng/ml and 40 ng/ml, respectively. To explore further the interaction of 3B4F11 with the beta-endorphin receptor, the effect of the Fab fragment on 125I-beta-endorphin cross-linking to rat brain membranes was examined. 125I-beta-endorphin was covalently bound to three major species of approximate molecular weights 108,000, 73,000, and 49,000. The delta-selective ligand D-Pen2, D-pen5enkephalin was least effective at inhibiting the cross-linking of beta-endorphin, whereas the micro-selective ligand Tyr-D-Ala-Gly-NMe-Phe-Gly-ol and kappa-selective ligand U50488 inhibited beta-endorphin cross-linking to the 108,000 and 73,000 Da species. Both 3B4F11 and beta-endorphin prevented the covalent binding of 125I-beta-endorphin to all three labeled species. These findings suggest that micro and kappa receptor types might have some structural similarities, whereas the delta receptor type might differ in molecular size. In addition, the micro, kappa, and delta ligands might have different primary sequences, whereas their tertiary structures might share regions of molecular homology with all three receptor constituents labeled by 125I-beta-endorphin. 3B4F11 will be a valuable tool for the purification and isolation of the several components of the beta-endorphin receptor complex.

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

    Science.gov (United States)

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

    2016-11-01

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

  16. Discrete mapping of brain Mu and delta opioid receptors using selective peptides: Quantitative autoradiography, species differences and comparison with kappa receptors

    Energy Technology Data Exchange (ETDEWEB)

    Sharif, N.A.; Hughes, J. (Addenbrookes Hospital Site, Cambridge (England))

    1989-05-01

    The opioid peptides, (3H)DAGO and (3H)DPDPE, bound to rat and guinea pig brain homogenates with a high, nanomolar affinity and to a high density of mu and delta receptors, respectively. (3H)DAGO binding to mu receptors was competitively inhibited by unlabelled opioids with the following rank order of potency: DAGO greater than morphine greater than DADLE greater than naloxone greater than etorphine much greater than U50488 much greater than DPDPE. In contrast, (3H)DPDPE binding to delta receptors was inhibited by compounds with the following rank order of potency: DPDPE greater than DADLE greater than etorphine greater than dynorphin(1-8) greater than naloxone much greater than U50488 much greater than DAGO. These profiles were consistent with specific labelling of the mu and delta opioid receptors, respectively. In vitro autoradiographic techniques coupled with computer-assisted image analyses revealed a discrete but differential anatomical localization of mu and delta receptors in the rat and guinea pig brain. In general, mu and delta receptor density in the rat exceeded that in the guinea pig brain and differed markedly from that of kappa receptors in these species. However, while mu receptors were distributed throughout the brain with hotspots in the fore-, mid- and hindbrain of the two rodents, the delta sites were relatively diffusely distributed, and were mainly concentrated in the forebrain with particularly high levels within the olfactory bulb (OB), n. accumbens and striatum. Notable regions of high density of mu receptors in the rat and guinea pig brain were the accessory olfactory bulb, striatal patches and streaks, amygdaloid nuclei, ventral hippocampal subiculum and dentate gyrus, numerous thalamic nuclei, geniculate bodies, central grey, superior and inferior colliculi, solitary and pontine nuclei and s. nigra.

  17. Evaluation of the kappa-opioid receptor-selective tracer [{sup 11}C]GR103545 in awake rhesus macaques

    Energy Technology Data Exchange (ETDEWEB)

    Schoultz, Bent W. [University of Oslo, Department of Chemistry, Oslo (Norway); Hjornevik, Trine; Willoch, Frode [University of Oslo, Centre for Molecular Biology and Neuroscience and Institute of Basic Medical Sciences, Oslo (Norway); Akershus University Hospital, Department of Nuclear Medicine, Loerenskog (Norway); Marton, Janos [ABX Advanced Biochemical Compounds GmbH, Radeberg (Germany); Noda, Akihiro; Murakami, Yoshihiro; Miyoshi, Sosuke; Nishimura, Shintaro [Medical and Pharmacological Research Center Foundation, Basic Research Department, Hakui City, Ishikawa (Japan); Aarstad, Erik [University College of London, Institute of Nuclear Medicine, London (United Kingdom); Drzezga, Alexander [Technische Universitaet Muenchen, Department of Nuclear Medicine, Klinikum rechts der Isar, Munich (Germany); Matsunari, Ichiro [Medical and Pharmacological Research Center Foundation, Clinical Research Department, Hakui City, Ishikawa (Japan); Henriksen, Gjermund [University of Oslo, Department of Chemistry, Oslo (Norway); Technische Universitaet Muenchen, Department of Nuclear Medicine, Klinikum rechts der Isar, Munich (Germany)

    2010-06-15

    The recent development in radiosynthesis of the {sup 11}C-carbamate function increases the potential of [{sup 11}C]GR103545, which for the last decade has been regarded as promising for imaging the kappa-opioid receptor ({kappa}-OR) with PET. In the present study, [{sup 11}C]GR103545 was evaluated in awake rhesus macaques. Separate investigations were performed to clarify the OR subtype selectivity of this compound. Regional brain uptake kinetics of [{sup 11}C]GR103545 was studied 0-120 min after injection. The binding affinity and opioid subtype selectivity of [{sup 11}C]GR103545 was determined in cells transfected with cloned human opioid receptors. In vitro binding assays demonstrated a high affinity of GR103545 for {kappa}-OR (K{sub i} = 0.02 {+-}0.01 nM) with excellent selectivity over {mu}-OR (6 x 10{sup 2}-fold) and {delta}-OR (2 x 10{sup 4}-fold). PET imaging revealed a volume of distribution (V{sub T}) pattern consistent with the known distribution of {kappa}-OR, with striatum = temporal cortex > cingulate cortex > frontal cortex > parietal cortex > thalamus > cerebellum. [{sup 11}C]GR103545 is selective for {kappa}-OR and holds promise for use to selectively depict and quantify this receptor in humans by means of PET. (orig.)

  18. Evaluation of the kappa-opioid receptor-selective tracer [11C]GR103545 in awake rhesus macaques

    International Nuclear Information System (INIS)

    Schoultz, Bent W.; Hjornevik, Trine; Willoch, Frode; Marton, Janos; Noda, Akihiro; Murakami, Yoshihiro; Miyoshi, Sosuke; Nishimura, Shintaro; Aarstad, Erik; Drzezga, Alexander; Matsunari, Ichiro; Henriksen, Gjermund

    2010-01-01

    The recent development in radiosynthesis of the 11 C-carbamate function increases the potential of [ 11 C]GR103545, which for the last decade has been regarded as promising for imaging the kappa-opioid receptor (κ-OR) with PET. In the present study, [ 11 C]GR103545 was evaluated in awake rhesus macaques. Separate investigations were performed to clarify the OR subtype selectivity of this compound. Regional brain uptake kinetics of [ 11 C]GR103545 was studied 0-120 min after injection. The binding affinity and opioid subtype selectivity of [ 11 C]GR103545 was determined in cells transfected with cloned human opioid receptors. In vitro binding assays demonstrated a high affinity of GR103545 for κ-OR (K i = 0.02 ±0.01 nM) with excellent selectivity over μ-OR (6 x 10 2 -fold) and δ-OR (2 x 10 4 -fold). PET imaging revealed a volume of distribution (V T ) pattern consistent with the known distribution of κ-OR, with striatum = temporal cortex > cingulate cortex > frontal cortex > parietal cortex > thalamus > cerebellum. [ 11 C]GR103545 is selective for κ-OR and holds promise for use to selectively depict and quantify this receptor in humans by means of PET. (orig.)

  19. Effects of combining opioids and clinically available NMDA receptor antagonists in the treatment of pain.

    NARCIS (Netherlands)

    Snijdelaar, D.G.

    2005-01-01

    This thesis concerns the effects of combining opioids with clinically available NMDA receptor antagonists in the treatment of acute and chronic pain. There are a number of problems with the use of opioids, such as, the development of tolerance/hyperalgesia, the reduced effectiveness in (central)

  20. Morphine withdrawal enhances constitutive μ-opioid receptor activity in the ventral tegmental area

    NARCIS (Netherlands)

    Meye, F.J.; van Zessen, R.; Smidt, M.P.; Adan, R.A.H.; Ramakers, G.M.J.

    2012-01-01

    μ-opioid receptors (MORs) in the ventral tegmental area (VTA) are pivotally involved in addictive behavior. While MORs are typically activated by opioids, they can also become constitutively active in the absence of any agonist. In the current study, we present evidence that MOR constitutive

  1. Interaction of 3,8-diazabicyclo (3.2.1) octanes with mu and delta opioid receptors.

    Science.gov (United States)

    Cignarella, G; Barlocco, D; Tranquillini, M E; Volterra, A; Brunello, N; Racagni, G

    1988-05-01

    A series of 3,8-diazabicyclo (3.2.1) octanes (DBO) (1) substituted at the nitrogen atoms by acyl and aralkenyl groups, were tested in in vitro binding assays towards mu and delta opioid receptors. The most representative terms (1a, 1d, 1g, 1j,) were also evaluated for the analgesic potency in vivo by the hot plate method. Among the compounds tested the most potent was the p.nitrocinnamyl DBO (1d) which displayed a mu/delta selectivity and an analgesic activity respectively 25 and 17 fold those of morphine. On the contrary, the m.hydroxycinnamyl DBO (1g) was markedly less active as agonist than the parent 1a, thus suggesting that structure 1 interacts with opioid receptors in a different fashion than morphine. Compound 1j isomer of 1a which is provided with high mu affinity, but lower analgesic potency, was found to possess a mixed agonist-antagonist activity.

  2. Mycobacteria attenuate nociceptive responses by formyl peptide receptor triggered opioid peptide release from neutrophils.

    Directory of Open Access Journals (Sweden)

    Heike L Rittner

    2009-04-01

    Full Text Available In inflammation, pain is regulated by a balance of pro- and analgesic mediators. Analgesic mediators include opioid peptides which are secreted by neutrophils at the site of inflammation, leading to activation of opioid receptors on peripheral sensory neurons. In humans, local opioids and opioid peptides significantly downregulate postoperative as well as arthritic pain. In rats, inflammatory pain is induced by intraplantar injection of heat inactivated Mycobacterium butyricum, a component of complete Freund's adjuvant. We hypothesized that mycobacterially derived formyl peptide receptor (FPR and/or toll like receptor (TLR agonists could activate neutrophils, leading to opioid peptide release and inhibition of inflammatory pain. In complete Freund's adjuvant-induced inflammation, thermal and mechanical nociceptive thresholds of the paw were quantified (Hargreaves and Randall-Selitto methods, respectively. Withdrawal time to heat was decreased following systemic neutrophil depletion as well as local injection of opioid receptor antagonists or anti-opioid peptide (i.e. Met-enkephalin, beta-endorphin antibodies indicating an increase in pain. In vitro, opioid peptide release from human and rat neutrophils was measured by radioimmunoassay. Met-enkephalin release was triggered by Mycobacterium butyricum and formyl peptides but not by TLR-2 or TLR-4 agonists. Mycobacterium butyricum induced a rise in intracellular calcium as determined by FURA loading and calcium imaging. Opioid peptide release was blocked by intracellular calcium chelation as well as phosphoinositol-3-kinase inhibition. The FPR antagonists Boc-FLFLF and cyclosporine H reduced opioid peptide release in vitro and increased inflammatory pain in vivo while TLR 2/4 did not appear to be involved. In summary, mycobacteria activate FPR on neutrophils, resulting in tonic secretion of opioid peptides from neutrophils and in a decrease in inflammatory pain. Future therapeutic strategies may aim

  3. Gold Nanorods Targeted to Delta Opioid Receptor: Plasmon-Resonant Contrast and Photothermal Agents

    Directory of Open Access Journals (Sweden)

    Kvar C. Black

    2008-01-01

    Full Text Available Molecularly targeted gold nanorods were investigated for applications in both diagnostic imaging and disease treatment with cellular resolution. The nanorods were tested in two genetically engineered cell lines derived from the human colon carcinoma HCT-116, a model for studying ligand-receptor interactions. One of these lines was modified to express delta opioid receptor (δOR and green fluorescent protein, whereas the other was receptor free and expressed a red fluorescent protein, to serve as the control. Deltorphin, a high-affinity ligand for δOR, was stably attached to the gold nanorods through a thiol-terminated linker. In a mixed population of cells, we demonstrated selective imaging and destruction of receptor-expressing cells while sparing those cells that did not express the receptor. The molecularly targeted nanorods can be used as an in vitro ligand-binding and cytotoxic treatment assay platform and could potentially be applied in vivo for diagnostic and therapeutic purposes with endoscopic technology.

  4. Dermorphin-related peptides from the skin of Phyllomedusa bicolor and their amidated analogs activate two mu opioid receptor subtypes that modulate antinociception and catalepsy in the rat.

    Science.gov (United States)

    Negri, L; Erspamer, G F; Severini, C; Potenza, R L; Melchiorri, P; Erspamer, V

    1992-08-01

    Three naturally occurring dermorphin-like peptides from the skin of the frog Phyllomedusa bicolor, the related carboxyl-terminal amides, and some substituted analogs were synthesized, their binding profiles to opioid receptors were determined, and their biological activities were studied in isolated organ preparations and intact animals. The opioid binding profile revealed a very high selectivity of these peptides for mu sites and suggested the existence of two receptor subtypes, of high and low affinity. The peptides tested acted as potent mu opioid agonists on isolated organ preparations. They were several times more active in inhibiting electrically evoked contractions in guinea pig ileum than in mouse vas deferens. When injected into the lateral brain ventricle or peritoneum of rats, the high-affinity-site-preferring ligand, [Lys7-NH2]dermorphin, behaved as a potent analgesic agent. By contrast, the low-affinity-site-preferring ligand, [Trp4,Asn7-NH2]dermorphin, produced a weak antinociception but an intense catalepsy.

  5. Potent μ-Opioid Receptor Agonists from Cyclic Peptides Tyr-c[D-Lys-Xxx-Tyr-Gly]: Synthesis, Biological, and Structural Evaluation.

    Science.gov (United States)

    Li, Yangmei; Cazares, Margret; Wu, Jinhua; Houghten, Richard A; Toll, Laurence; Dooley, Colette

    2016-02-11

    To optimize the structure of a μ-opioid receptor ligand, analogs H-Tyr-c[D-Lys-Xxx-Tyr-Gly] were synthesized and their biological activity was tested. The analog containing a Phe(3) was identified as not only exhibiting binding affinity 14-fold higher than the original hit but also producing agonist activity 3-fold more potent than morphine. NMR study suggested that a trans conformation at D-Lys(2)-Xxx(3) is crucial for these cyclic peptides to maintain high affinity, selectivity, and functional activity toward the μ-opioid receptor.

  6. The opioid receptor pharmacology of GSK1521498 compared to other ligands with differential effects on compulsive reward-related behaviours.

    Science.gov (United States)

    Kelly, Eamonn; Mundell, Stuart J; Sava, Anna; Roth, Adelheid L; Felici, Antonio; Maltby, Kay; Nathan, Pradeep J; Bullmore, Edward T; Henderson, Graeme

    2015-01-01

    The novel opioid receptor antagonist, GSK1421498, has been shown to attenuate reward-driven compulsive behaviours, such as stimulant drug seeking or binge eating, in animals and humans. Here, we report new data on the receptor pharmacology of GSK121498, in comparison to naltrexone, naloxone, 6-β-naltrexol and nalmefene. To determine whether the novel opioid antagonist, GSK1521498, is an orthosteric or allosteric antagonist at the μ opioid receptor (MOPr) and whether it has neutral antagonist or inverse agonist properties. A combination of radioligand binding assays and [(35)S]GTPγS binding assays was employed. GSK1521498 completely displaced [(3)H]naloxone binding to MOPr and did not alter the rate of [(3)H]naloxone dissociation from MOPr observations compatible with it binding to the orthosteric site on MOPr. GSK1521498 exhibited inverse agonism when MOPr was overexpressed but not when the level of MOPr expression was low. In parallel studies under conditions of high receptor expression density, naloxone, naltrexone, 6-β-naltrexol and nalmefene exhibited partial agonism, not inverse agonism as has been reported previously for naloxone and naltrexone. In brain tissue from mice receiving a prolonged morphine pre-treatment, GSK1521498 exhibited slight inverse agonism. Differences between GSK1521498 and naltrexone in their effects on compulsive reward seeking are arguably linked to the more selective and complete MOPr antagonism of GSK1521498 versus the partial MOPr agonism of naltrexone. GSK1521498 is also pharmacologically differentiated by its inverse agonist efficacy at high levels of MOPr expression, but this may be less likely to contribute to behavioural differentiation at patho-physiological levels of expression.

  7. INTERACTION BETWEEN DELTA OPIOID RECEPTORS AND BENZODIAZEPINES IN CO2- INDUCED RESPIRATORY RESPONSES IN MICE

    Science.gov (United States)

    Borkowski, Anne H.; Barnes, Dylan C.; Blanchette, Derek R.; Castellanos, F. Xavier; Klein, Donald F.; Wilson, Donald A.

    2011-01-01

    The false-suffocation hypothesis of panic disorder (Klein, 1993) suggested δ-opioid receptors as a possible source of the respiratory dysfunction manifested in panic attacks occurring in panic disorder (Preter and Klein, 2008). This study sought to determine if a lack of δ-opioid receptors in a mouse model affects respiratory response to elevated CO2, and whether the response is modulated by benzodiazepines, which are widely used to treat panic disorder. In a whole-body plethysmograph, respiratory responses to 5% CO2 were compared between δ-opioid receptor knockout mice and wild-type mice after saline, diazepam (1 mg/kg), and alprazolam (0.3 mg/kg) injection. The results show that lack of δ-opioid receptors does not affect normal response to elevated CO2, but does prevent benzodiazepines from modulating that response. Thus, in the presence of benzodiazepine agonists, respiratory responses to elevated CO2 were enhanced in δ-opioid receptor knockout mice compared to wild-type mice. This suggests an interplay between benzodiazepine receptors and δ-opioid receptors in regulating the respiratory effects of elevated CO2, which might be related to CO2 induced panic. PMID:21561601

  8. Optimisation of in silico derived 2-aminobenzimidazole hits as unprecedented selective kappa opioid receptor agonists

    DEFF Research Database (Denmark)

    Sasmal, Pradip K; Krishna, C Vamsee; Sudheerkumar Adabala, S

    2015-01-01

    Kappa opioid receptor (KOR) is an important mediator of pain signaling and it is targeted for the treatment of various pains. Pharmacophore based mining of databases led to the identification of 2-aminobenzimidazole derivative as KOR agonists with selectivity over the other opioid receptors DOR a...... of novel benzimidazole derivatives as KOR agonists are described. The in vivo proof of principle for anti-nociceptive effect with a lead compound from this series is exemplified....

  9. 14-O-Methylmorphine: A Novel Selective Mu-Opioid Receptor Agonist with High Efficacy and Affinity.

    Science.gov (United States)

    Zádor, Ferenc; Balogh, Mihály; Váradi, András; Zádori, Zoltán S; Király, Kornél; Szűcs, Edina; Varga, Bence; Lázár, Bernadette; Hosztafi, Sándor; Riba, Pál; Benyhe, Sándor; Fürst, Susanna; Al-Khrasani, Mahmoud

    2017-11-05

    14-O-methyl (14-O-Me) group in morphine-6-O-sulfate (M6SU) or oxymorphone has been reported to be essential for enhanced affinity, potency and antinociceptive effect of these opioids. Herein we report on the pharmacological properties (potency, affinity and efficacy) of the new compound, 14-O-methylmorphine (14-O-MeM) in in vitro. Additionally, we also investigated the antinociceptive effect of the novel compound, as well as its inhibitory action on gastrointestinal transit in in vivo. The potency and efficacy of test compound were measured by [ 35 S]GTPγS binding, isolated mouse vas deferens (MVD) and rat vas deferens (RVD) assays. The affinity of 14-O-MeM for opioid receptors was assessed by radioligand binding and MVD assays. The antinociceptive and gastrointestinal effects of the novel compound were evaluated in the rat tail-flick test and charcoal meal test, respectively. Morphine, DAMGO, Ile 5,6 deltorphin II, deltorphin II and U-69593 were used as reference compounds. 14-O-MeM showed higher efficacy (E max ) and potency (EC 50 ) than morphine in MVD, RVD or [ 35 S]GTPγS binding. In addition, 14-O-MeM compared to morphine showed higher affinity for μ-opioid receptor (MOR). In vivo, in rat tail-flick test 14-O-MeM proved to be stronger antinociceptive agent than morphine after peripheral or central administration. Additionally, both compounds inhibited the gastrointestinal peristalsis. However, when the antinociceptive and antitransit doses for each test compound are compared, 14-O-MeM proved to have slightly more favorable pharmacological profile. Our results affirm that 14-O-MeM, an opioid of high efficacy and affinity for MOR can be considered as a novel analgesic agent of potential clinical value. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. A DFT and semiempirical model-based study of opioid receptor affinity and selectivity in a group of molecules with a morphine structural core.

    Science.gov (United States)

    Bruna-Larenas, Tamara; Gómez-Jeria, Juan S

    2012-01-01

    We report the results of a search for model-based relationships between mu, delta, and kappa opioid receptor binding affinity and molecular structure for a group of molecules having in common a morphine structural core. The wave functions and local reactivity indices were obtained at the ZINDO/1 and B3LYP/6-31G(∗∗) levels of theory for comparison. New developments in the expression for the drug-receptor interaction energy expression allowed several local atomic reactivity indices to be included, such as local electronic chemical potential, local hardness, and local electrophilicity. These indices, together with a new proposal for the ordering of the independent variables, were incorporated in the statistical study. We found and discussed several statistically significant relationships for mu, delta, and kappa opioid receptor binding affinity at both levels of theory. Some of the new local reactivity indices incorporated in the theory appear in several equations for the first time in the history of model-based equations. Interaction pharmacophores were generated for mu, delta, and kappa receptors. We discuss possible differences regulating binding and selectivity in opioid receptor subtypes. This study, contrarily to the statistically backed ones, is able to provide a microscopic insight of the mechanisms involved in the binding process.

  11. A DFT and Semiempirical Model-Based Study of Opioid Receptor Affinity and Selectivity in a Group of Molecules with a Morphine Structural Core

    Directory of Open Access Journals (Sweden)

    Tamara Bruna-Larenas

    2012-01-01

    Full Text Available We report the results of a search for model-based relationships between mu, delta, and kappa opioid receptor binding affinity and molecular structure for a group of molecules having in common a morphine structural core. The wave functions and local reactivity indices were obtained at the ZINDO/1 and B3LYP/6-31 levels of theory for comparison. New developments in the expression for the drug-receptor interaction energy expression allowed several local atomic reactivity indices to be included, such as local electronic chemical potential, local hardness, and local electrophilicity. These indices, together with a new proposal for the ordering of the independent variables, were incorporated in the statistical study. We found and discussed several statistically significant relationships for mu, delta, and kappa opioid receptor binding affinity at both levels of theory. Some of the new local reactivity indices incorporated in the theory appear in several equations for the first time in the history of model-based equations. Interaction pharmacophores were generated for mu, delta, and kappa receptors. We discuss possible differences regulating binding and selectivity in opioid receptor subtypes. This study, contrarily to the statistically backed ones, is able to provide a microscopic insight of the mechanisms involved in the binding process.

  12. Opioid and nicotine receptors affect growth regulation of human lung cancer cell lines

    International Nuclear Information System (INIS)

    Maneckjee, R.; Minna, J.D.

    1990-01-01

    Using specific radioactively-labeled ligands, the authors find that lung cancer cell lines of diverse histologic types express multiple, high-affinity membrane receptors for μ, δ, and κ opioid agonists and for nicotine and α-bungarotoxin. These receptors are biologically active because cAMP levels decreased in lung cancer cells after opioid and nicotine application. Nicotine at concentrations found in the blood of smokers had no effect on in vitro lung cancer cell growth, whereas μ, δ, and κ opioid agonists at low concentrations inhibited lung cancer growth in vitro. They also found that lung cancer cells expressed various combinations of immunoreactive opioid peptides (β-endorphin, enkephalin, or dynorphin), suggesting the participation of opioids in a negative autocrine loop or tumor-suppressing system. Due to the almost universal exposure of patients with lung cancer to nicotine, they tested whether nicotine affected the response of lung cancer cell growth to opioids and found that nicotine at concentrations of 100-200 nM partially or totally reversed opioid-induced growth inhibition in 9/14 lung cancer cell lines. These in vitro results for lung cancer cells suggest that opioids could function as part of a tumor suppressor system and that nicotine can function to circumvent this system in the pathogenesis of lung cancer

  13. Opioid and nicotine receptors affect growth regulation of human lung cancer cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Maneckjee, R.; Minna, J.D. (National Cancer Institute-Navy Medical Oncology Branch, Bethesda, MD (USA) Uniformed Services Univ. of the Health Sciences, Bethesda, MD (USA))

    1990-05-01

    Using specific radioactively-labeled ligands, the authors find that lung cancer cell lines of diverse histologic types express multiple, high-affinity membrane receptors for {mu}, {delta}, and {kappa} opioid agonists and for nicotine and {alpha}-bungarotoxin. These receptors are biologically active because cAMP levels decreased in lung cancer cells after opioid and nicotine application. Nicotine at concentrations found in the blood of smokers had no effect on in vitro lung cancer cell growth, whereas {mu}, {delta}, and {kappa} opioid agonists at low concentrations inhibited lung cancer growth in vitro. They also found that lung cancer cells expressed various combinations of immunoreactive opioid peptides ({beta}-endorphin, enkephalin, or dynorphin), suggesting the participation of opioids in a negative autocrine loop or tumor-suppressing system. Due to the almost universal exposure of patients with lung cancer to nicotine, they tested whether nicotine affected the response of lung cancer cell growth to opioids and found that nicotine at concentrations of 100-200 nM partially or totally reversed opioid-induced growth inhibition in 9/14 lung cancer cell lines. These in vitro results for lung cancer cells suggest that opioids could function as part of a tumor suppressor system and that nicotine can function to circumvent this system in the pathogenesis of lung cancer.

  14. Reduced basal ganglia μ-opioid receptor availability in trigeminal neuropathic pain: A pilot study

    Directory of Open Access Journals (Sweden)

    DosSantos Marcos

    2012-09-01

    Full Text Available Abstract Background Although neuroimaging techniques have provided insights into the function of brain regions involved in Trigeminal Neuropathic Pain (TNP in humans, there is little understanding of the molecular mechanisms affected during the course of this disorder. Understanding these processes is crucial to determine the systems involved in the development and persistence of TNP. Findings In this study, we examined the regional μ-opioid receptor (μOR availability in vivo (non-displaceable binding potential BPND of TNP patients with positron emission tomography (PET using the μOR selective radioligand [11C]carfentanil. Four TNP patients and eight gender and age-matched healthy controls were examined with PET. Patients with TNP showed reduced μOR BPND in the left nucleus accumbens (NAc, an area known to be involved in pain modulation and reward/aversive behaviors. In addition, the μOR BPND in the NAc was negatively correlated with the McGill sensory and total pain ratings in the TNP patients. Conclusions Our findings give preliminary evidence that the clinical pain in TNP patients can be related to alterations in the endogenous μ-opioid system, rather than only to the peripheral pathology. The decreased availability of μORs found in TNP patients, and its inverse relationship to clinical pain levels, provide insights into the central mechanisms related to this condition. The results also expand our understanding about the impact of chronic pain on the limbic system.

  15. Receptor binding studies of the living heart

    International Nuclear Information System (INIS)

    Syrota, A.

    1988-01-01

    Receptors form a class of intrinsic membrane proteins (or glycoproteins) defined by the high affinity and specificity with which they bind ligands. Many receptors are associated directly or indirectly with membrane ion channels that open or close after a conformational change of the receptor induced by the binding of the neurotransmitter. Changes in number and/or affinity of cardiac neurotransmitter receptors have been associated with myocardial ischemia and infarction, congestive heart failure, and cardiomyopathy as well as diabetes or thyroid-induced heart muscle disease. These alterations of cardiac receptors have been demonstrated in vitro on membrane homogenates from samples collected mainly during surgery or postmortem. The disadvantage of these in vitro binding techniques is that receptors lose their natural environment and their relationships with the other components of the tissue

  16. Affinity crosslinking of /sup 125/I-human beta-endorphin to cell lines possessing either mu or delta type opioid binding sites

    Energy Technology Data Exchange (ETDEWEB)

    Keren, O.; Gioannini, T.L.; Hiller, J.M.; Simon, E.J.

    1986-01-01

    Affinity crosslinking of human /sup 125/I-beta-Endorphin to cell lines possessing either mu or delta binding sites was carried out. Autoradiography of SDS-PAGE gels from these crosslinked cell lines revealed that these two sites contain major peptide subunits that differ in molecular size. This confirms our earlier finding in mammalian brain which demonstrated separate and distinct subunits for mu and delta opioid receptors.

  17. Kappa opioid receptors stimulate phosphoinositide turnover in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Periyasamy, S.; Hoss, W. (Univ. of Toledo, OH (USA))

    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.

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

    Science.gov (United States)

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

    2017-11-15

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

  19. Inhibition of GABAergic Neurotransmission by HIV-1 Tat and Opioid Treatment in the Striatum Involves μ-opioid Receptors

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    Changqing Xu

    2016-11-01

    Full Text Available Due to combined antiretroviral therapy (cART, human immunodeficiency virus type 1 (HIV-1 is considered a chronic disease with high prevalence of mild forms of neurocognitive impairments, also referred to as HIV-associated neurocognitive disorders (HAND. Although opiate drug use can exacerbate HIV-1 Tat-induced neuronal damage, it remains unknown how and to what extent opioids interact with Tat on the GABAergic system. We conducted whole-cell recordings in mouse striatal slices and examined the effects of HIV-1 Tat in the presence and absence of morphine (1 μM and damgo (1 μM on GABAergic neurotransmission. Results indicated a decrease in the frequency and amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs and miniature IPSCs (mIPSCs by Tat (5 – 50 nM in a concentration-dependent manner. The significant Tat-induced decrease in IPSCs was abolished when removing extracellular and/or intracellular calcium. Treatment with morphine or damgo alone significantly decreased the frequency, but not amplitude of IPSCs. Interestingly, morphine but not damgo indicated an additional downregulation of the mean frequency of mIPSCs in combination with Tat. Pretreatment with naloxone (1 μM and CTAP (1 μM prevented the Tat-induced decrease in sIPSCs frequency but only naloxone prevented the combined Tat and morphine effect on mIPSCs frequency. Results indicate a Tat- or opioid-induced decrease in GABAergic neurotransmission via µ-opioid receptors with combined Tat and morphine effects involving additional opioid receptor-related mechanisms. Exploring the interactions between Tat and opioids on the GABAergic system may help to guide future research on HAND in the context of opiate drug use.

  20. South African guideline for the use of chronic opioid therapy for ...

    African Journals Online (AJOL)

    pain and chronic pain associated with cancer and at the end of life. Although .... Opioid drugs are agonists that bind to endogenous opioid receptors and mimic ..... interstitial cystitis/painful bladder syndrome, chronic prostate pain, and irritable ...

  1. Hypothalamic kappa opioid receptor mediates both diet- and MCH-induced liver damage through inflammation and ER stress

    NARCIS (Netherlands)

    Imbernon, Monica; Sanchez-Rebordelo, Estrella; Romero-Picó, Amparo; Kalló, Imre; Chee, Melissa J; Porteiro, Begoña; Al-Massadi, Omar; Contreras, Cristina; Fernø, Johan; Senra, Ana; Gallego, Rosalia; Folgueira, Cintia; Seoane, Luisa M; van Gestel, Margriet; Adan, Roger A; Liposits, Zsolt; Dieguez, Carlos; Lopez, Miguel; Nogueiras, Ruben

    2016-01-01

    The opioid system is widely known to modulate the brain reward system and thus affect human and animal behaviour, including feeding. We hypothesized that the hypothalamic opioid system might also control energy metabolism in peripheral tissues. Mice lacking the kappa opioid receptor (κOR) and

  2. Mu opioid receptor availability in people with psychiatric disorders who died by suicide: a case control study

    Directory of Open Access Journals (Sweden)

    Scarr Elizabeth

    2012-08-01

    Full Text Available Abstract Background Mu opioid receptors have previously been shown to be altered in people with affective disorders who died as a result of suicide. We wished to determine whether these changes were more widespread and independent of psychiatric diagnoses. Methods Mu receptor levels were determined using [3 H]DAMGO binding in BA24 from 51 control subjects; 38 people with schizophrenia (12 suicides; 20 people with major depressive disorder (15 suicides; 13 people with bipolar disorder (5 suicides and 9 people who had no history of psychiatric disorders but who died as a result of suicide. Mu receptor levels were further determined in BA9 and caudate-putamen from 38 people with schizophrenia and 20 control subjects using [3 H]DAMGO binding and, in all three regions, using Western blots. Data was analysed using one-way ANOVAs with Bonferroni’s Multiple Comparison Test or, where data either didn’t approximate to a binomial distribution or the sample size was too small to determine distribution, a Kruskal-Wallis test with Dunn’s Multiple Comparison Test. Results [3 H]DAMGO binding density was lower in people who had died as a result of suicide (p3 H]DAMGO binding densities, but not mu protein levels, were significantly decreased in BA9 from people with schizophrenia who died as a result of suicide (p Conclusions Overall these data suggest that mu opioid receptor availability is decreased in the brains of people with schizophrenia who died as a result of suicide, which would be consistent with increased levels of endogenous ligands occupying these receptors.

  3. Antinociceptive Action of Isolated Mitragynine from Mitragyna Speciosa through Activation of Opioid Receptor System

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    Mohamad Aris Mohd Moklas

    2012-09-01

    Full Text Available Cannabinoids and opioids systems share numerous pharmacological properties and antinociception is one of them. Previous findings have shown that mitragynine (MG, a major indole alkaloid found in Mitragyna speciosa (MS can exert its antinociceptive effects through the opioids system. In the present study, the action of MG was investigated as the antinociceptive agent acting on Cannabinoid receptor type 1 (CB1 and effects on the opioids receptor. The latency time was recorded until the mice showed pain responses such as shaking, licking or jumping and the duration of latency was measured for 2 h at every 15 min interval by hot plate analysis. To investigate the beneficial effects of MG as antinociceptive agent, it was administered intraperitoneally 15 min prior to pain induction with a single dosage (3, 10, 15, 30, and 35 mg/kg b.wt. In this investigation, 35 mg/kg of MG showed significant increase in the latency time and this dosage was used in the antagonist receptor study. The treated groups were administered with AM251 (cannabinoid receptor-1 antagonist, naloxone (non-selective opioid antagonist, naltrindole (δ-opioid antagonist naloxonazine (µ1-receptor antagonist and norbinaltorpimine (κ-opioid antagonist respectively, prior to administration of MG (35 mg/kg. The results showed that the antinociceptive effect of MG was not antagonized by AM251; naloxone and naltrindole were effectively blocked; and norbinaltorpimine partially blocked the antinociceptive effect of MG. Naloxonazine did inhibit the effect of MG, but it was not statistically significant. These results demonstrate that CB1 does not directly have a role in the antinociceptive action of MG where the effect was observed with the activation of opioid receptor.

  4. Amygdala mu-opioid receptors mediate the motivating influence of cue-triggered reward expectations.

    Science.gov (United States)

    Lichtenberg, Nina T; Wassum, Kate M

    2017-02-01

    Environmental reward-predictive stimuli can retrieve from memory a specific reward expectation that allows them to motivate action and guide choice. This process requires the basolateral amygdala (BLA), but little is known about the signaling systems necessary within this structure. Here we examined the role of the neuromodulatory opioid receptor system in the BLA in such cue-directed action using the outcome-specific Pavlovian-to-instrumental transfer (PIT) test in rats. Inactivation of BLA mu-, but not delta-opioid receptors was found to dose-dependently attenuate the ability of a reward-predictive cue to selectively invigorate the performance of actions directed at the same unique predicted reward (i.e. to express outcome-specific PIT). BLA mu-opioid receptor inactivation did not affect the ability of a reward itself to similarly motivate action (outcome-specific reinstatement), suggesting a more selective role for the BLA mu-opioid receptor in the motivating influence of currently unobservable rewarding events. These data reveal a new role for BLA mu-opioid receptor activation in the cued recall of precise reward memories and the use of this information to motivate specific action plans. © 2016 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  5. Evaluation of Analgesic Activity of Papaver libanoticum Extract in Mice: Involvement of Opioids Receptors

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    Mohamad Ali Hijazi

    2017-01-01

    Full Text Available Papaver libanoticum is an endemic plant to Lebanese region (family Papaveraceae that has not been investigated before. The present study aimed to explore the analgesic activity of dried ethanolic extract of Papaver libanoticum (PLE using tail flick, hot plate, and acetic acid induced writhing models in mice. The involvement of opioid receptors in the analgesic mechanism was investigated using naloxone antagonism. Results demonstrated that PLE exhibited a potent dose dependent analgesic activity in all tested models for analgesia. The analgesic effect involved activation of opioid receptors in the central nervous system, where both spinal and supraspinal components might be involved. The time course for analgesia revealed maximum activity after three hours in both tail flick and hot plate methods, which was prolonged to 24 hours. Metabolites of PLE could be responsible for activation of opioid receptors. The EC50 of PLE was 79 and 50 mg/kg in tail flick and hot plate tests, respectively. The total coverage of analgesia by PLE was double that of morphine in both tests. In conclusion, PLE proved to have opioid agonistic activity with a novel feature of slow and prolonged effect. The present study could add a potential tool in the armaments of opioid drugs as a natural potent analgesic and for treatment of opioid withdrawal syndrome.

  6. Endogenous opioid peptide-mediated neurotransmission in central and pericentral nuclei of the inferior colliculus recruits μ1-opioid receptor to modulate post-ictal antinociception.

    Science.gov (United States)

    Felippotti, Tatiana Tocchini; de Freitas, Renato Leonardo; Coimbra, Norberto Cysne

    2012-02-01

    The aim of the present work was to investigate the involvement of the μ1-endogenous opioid peptide receptor-mediated system in post-ictal antinociception. Antinociceptive responses were determined by the tail-flick test after pre-treatment with the selective μ1-opioid receptor antagonist naloxonazine, peripherally or centrally administered at different doses. Peripheral subchronic (24 h) pre-treatment with naloxonazine antagonised the antinociception elicited by tonic-clonic seizures. Acute (10 min) pre-treatment, however, did not have the same effect. In addition, microinjections of naloxonazine into the central, dorsal cortical and external cortical nuclei of the inferior colliculus antagonised tonic-clonic seizure-induced antinociception. Neither acute (10-min) peripheral pre-treatment with naloxonazine nor subchronic intramesencephalic blockade of μ1-opioid receptors resulted in consistent statistically significant differences in the severity of tonic-clonic seizures shown by Racine's index (1972), although the intracollicular specific antagonism of μ1-opioid receptor decreased the duration of seizures. μ1-Opioid receptors and the inferior colliculus have been implicated in several endogenous opioid peptide-mediated responses such as antinociception and convulsion. The present findings suggest the involvement of μ1-opiate receptors of central and pericentral nuclei of the inferior colliculus in the modulation of tonic-clonic seizures and in the organisation of post-ictal antinociception. Copyright © 2011 Elsevier Ltd. All rights reserved.

  7. Opioid and GABAB receptors differentially couple to an adenylyl cyclase/protein kinase A downstream effector after chronic morphine treatment.

    Directory of Open Access Journals (Sweden)

    Elena Elizabeth Bagley

    2014-06-01

    Full Text Available Opioids are intensely addictive, and cessation of their chronic use is associated with a highly aversive withdrawal syndrome. A cellular hallmark of withdrawal is an opioid sensitive protein kinase A-dependent increase in GABA transporter-1 (GAT-1 currents in periaqueductal gray (PAG neurons. Elevated GAT-1 activity directly increases GABAergic neuronal excitability and synaptic GABA release, which will enhance GABAergic inhibition of PAG output neurons. This reduced activity of PAG output neurons to several brain regions, including the hypothalamus and medulla, contributes to many of the PAG-mediated signs of opioid withdrawal. The GABAB receptor agonist baclofen reduces some of the PAG mediated signs of opioid withdrawal. Like the opioid receptors the GABAB receptor is a Gi/Go coupled G-protein coupled receptor. This suggests it could be modulating GAT-1 activity in PAG neurons through its inhibition of the adenylyl cyclase/protein kinase A pathway. Opioid modulation of the GAT-1 activity can be detected by changes in the reversal potential of opioid membrane currents. We found that when opioids are reducing the GAT-1 cation conductance and increasing the GIRK conductance the opioid agonist reversal potential is much more negative than Ek. Using this approach for GABAB receptors we show that the GABAB receptor agonist, baclofen, does not couple to inhibition of GAT-1 currents during opioid withdrawal. It is possible this differential signaling of the two Gi/Go coupled G-protein coupled receptors is due to the strong compartmentalization of the GABAB receptor that does not favor signaling to the adenylyl cyclase/protein kinase A/GAT-1 pathway. This highlights the importance of studying the effects of G-protein coupled receptors in native tissue with endogenous G-protein coupled receptors and the full complement of relevant proteins and signaling molecules. This study suggests that baclofen reduces opioid withdrawal symptoms through a non-GAT-1

  8. New features of the delta opioid receptor: conformational properties of deltorphin I analogues.

    Science.gov (United States)

    Balboni, G; Marastoni, M; Picone, D; Salvadori, S; Tancredi, T; Temussi, P A; Tomatis, R

    1990-06-15

    Deltorphin I is an opioid peptide of sequence H-Tyr-D-Ala-Phe-Asp-Val-Val-Gly-NH2, recently isolated from the skin of Phyllomedusa bicolor. Its enormous selectivity towards the delta opioid receptor and the similarity of the conformation of the N-terminal part of the sequence with that of dermorphin (H-Tyr-D-Ala-he-Gly-Tyr-Pro-Ser-NH2), a mu selective peptide, prompted the synthesis, biological evaluation and comparative conformational study of four analogs. A 1H-NMR study showed that the conformational preferences of the N-terminal sequences of all peptides are similar. The different selectivities towards opioid receptors have been interpreted in terms of charge effects in the interaction with the membrane and at the receptor site and of hydrophobicity of the C-terminal part, when structured in a folded conformation.

  9. Autoradiographic localization of delta opioid receptors within the mesocorticolimbic dopamine system using radioiodinated (2-D-penicillamine, 5-D-penicillamine)enkephalin ( sup 125 I-DPDPE)

    Energy Technology Data Exchange (ETDEWEB)

    Dilts, R.P.; Kalivas, P.W. (Washington State Univ., Pullman (USA))

    1990-01-01

    The enkephalin analog (2-D-penicillamine, 5-D-penicillamine)enkephalin was radioiodinated (125I-DPDPE) and shown to retain a pharmacological selectivity characteristic of the delta opioid receptor in in vitro binding studies. The distributions of 125I-DPDPE binding, using in vitro autoradiographic techniques, were similar to those previously reported for the delta opioid receptor. The nucleus accumbens, striatum, and medial prefrontal cortex contain dense gradients of 125I-DPDPE binding in regions known to receive dopaminergic afferents emanating from the mesencephalic tegmentum. Selective chemical lesions of the ventral tegmental area and substantia nigra were employed to deduce the location of the 125I-DPDPE binding within particular regions of the mesocorticolimbic dopamine system. Unilateral lesions of dopamine perikarya (A9 and A10) within the ventral tegmental area and substantia nigra produced by mesencephalic injection of 6-hydroxydopamine resulted in significant (20-30%) increases in 125I-DPDPE binding contralateral to the lesion within the striatum and nucleus accumbens. Lesions of the perikarya (dopaminergic and nondopaminergic) of the ventral tegmental area, induced by quinolinic acid injections, caused increases of less magnitude within these same nuclei. No significant alterations in 125I-DPDPE binding were observed within the mesencephalon as a result of either treatment. The specificity of the lesions was confirmed by immunocytochemistry for tyrosine hydroxylase. These results suggest that the enkephalins and opioid agonists acting through delta opioid receptors do not directly modulate dopaminergic afferents but do regulate postsynaptic targets of the mesocorticolimbic dopamine system.

  10. Regulation of extinction-related plasticity by opioid receptors in the ventrolateral periaqueductal gray matter

    Directory of Open Access Journals (Sweden)

    Ryan Parsons

    2010-08-01

    Full Text Available Recent work has led to a better understanding of the neural mechanisms underlying the extinction of Pavlovian fear conditioning. Long-term synaptic changes in the medial prefrontal cortex (mPFC are critical for extinction learning, but very little is currently known about how the mPFC and other brain areas interact during extinction. The current study examined the effect of drugs that impair the extinction of fear conditioning on the activation of the extracellular-related kinase/mitogen-activated protein kinase (ERK/MAPK in brain regions that likely participate in the consolidation of extinction learning. Inhibitors of opioid and N-methyl-D-aspartic acid (NMDA receptors were applied to the ventrolateral periaqueductal gray matter (vlPAG and amygdala shortly before extinction training. Results from these experiments show that blocking opioid receptors in the vlPAG prevented the formation of extinction memory, whereas NMDA receptor blockade had no effect. Conversely, blocking NMDA receptors in the amygdala disrupted the formation of fear extinction memory, but opioid receptor blockade in the same brain area did not. Subsequent experiments tested the effect of these drug treatments on the activation of the ERK/MAPK signaling pathway in various brain regions following extinction training. Only opioid receptor blockade in the vlPAG disrupted ERK phosphorylation in the mPFC and amygdala. These data support the idea that opiodergic signaling derived from the vlPAG affects plasticity across the brain circuit responsible for the formation of extinction memory.

  11. Electroacupuncture-Induced Dynamic Processes of Gene Expression Levels of Endogenous Opioid Peptide Precursors and Opioid Receptors in the CNS of Goats

    Directory of Open Access Journals (Sweden)

    Li-Li Cheng

    2013-01-01

    Full Text Available In order to investigate the dynamic processes of mRNA levels of proenkephalin, proopiomelanocortin, prodynorphin, and opioid receptors (δ-, μ-, and κ-receptor induced by electroacupuncture (EA in the central nerve system, goats were stimulated by EA of 60 Hz for 0.5 h at a set of Baihui, Santai, Ergen, and Sanyangluo points. The pain threshold was measured using the method of potassium iontophoresis. The mRNA levels of the three opioid peptide precursors and three opioid receptors were determined with quantitative real-time PCR and the levels of Met-enkephalin with SABC immunohistochemistry at 0.5 h before and at 0, 2, 4, 6, 8, 12, and 24 h after EA. The results showed that the pain threshold correlated (P<0.01 with Met-enkephalin immunoactivities in the measured nuclei and areas of goats. The analgesic aftereffect lasted for 12 h at least. The mRNA levels of the three opioid peptide precursors and three opioid receptors began to increase at 0 h, reached the peak during the time from 4 h to 6 h or at 12 h, and remained higher at 24 h after EA was discontinued. These results suggested that the initiation of gene expression of opioid peptides and the three receptors may be associated with EA-induced analgesic aftereffect.

  12. Synthesis of [3]DIPPA: a potent irreversible antagonist selective for the κ opioid receptor

    International Nuclear Information System (INIS)

    Chang, Anchih; Portoghese, P.S.

    1995-01-01

    2-(3,4-Dichlorophenyl)-N-methyl-N-[(1S)-1-(3-isothiocyanatophe nyl)-2-(1-pyrrolidinyl)ethyl]acetamide (1,DIPPA) has been previously reported to be an opioid receptor affinity label that produces selective and long-lasting κ opioid receptor antagonism in mice. High specific activity [ 3 H]DIPPA (39.7 Ci/mmol) was prepared by bromination and catalytic tritiation of the amino precursor of DIPPA followed by conversion to the isothiocyanate with thiophosgene. (Author)

  13. Supersensitive Kappa Opioid Receptors Promotes Ethanol Withdrawal-Related Behaviors and Reduce Dopamine Signaling in the Nucleus Accumbens.

    Science.gov (United States)

    Rose, Jamie H; Karkhanis, Anushree N; Chen, Rong; Gioia, Dominic; Lopez, Marcelo F; Becker, Howard C; McCool, Brian A; Jones, Sara R

    2016-05-01

    Chronic ethanol exposure reduces dopamine transmission in the nucleus accumbens, which may contribute to the negative affective symptoms associated with ethanol withdrawal. Kappa opioid receptors have been implicated in withdrawal-induced excessive drinking and anxiety-like behaviors and are known to inhibit dopamine release in the nucleus accumbens. The effects of chronic ethanol exposure on kappa opioid receptor-mediated changes in dopamine transmission at the level of the dopamine terminal and withdrawal-related behaviors were examined. Five weeks of chronic intermittent ethanol exposure in male C57BL/6 mice were used to examine the role of kappa opioid receptors in chronic ethanol-induced increases in ethanol intake and marble burying, a measure of anxiety/compulsive-like behavior. Drinking and marble burying were evaluated before and after chronic intermittent ethanol exposure, with and without kappa opioid receptor blockade by nor-binaltorphimine (10mg/kg i.p.). Functional alterations in kappa opioid receptors were assessed using fast scan cyclic voltammetry in brain slices containing the nucleus accumbens. Chronic intermittent ethanol-exposed mice showed increased ethanol drinking and marble burying compared with controls, which was attenuated with kappa opioid receptor blockade. Chronic intermittent ethanol-induced increases in behavior were replicated with kappa opioid receptor activation in naïve mice. Fast scan cyclic voltammetry revealed that chronic intermittent ethanol reduced accumbal dopamine release and increased uptake rates, promoting a hypodopaminergic state of this region. Kappa opioid receptor activation with U50,488H concentration-dependently decreased dopamine release in both groups; however, this effect was greater in chronic intermittent ethanol-treated mice, indicating kappa opioid receptor supersensitivity in this group. These data suggest that the chronic intermittent ethanol-induced increase in ethanol intake and anxiety

  14. Dopamine D1 receptor gene variation modulates opioid dependence risk by affecting transition to addiction.

    Directory of Open Access Journals (Sweden)

    Feng Zhu

    Full Text Available Dopamine D1 receptor (DRD1 modulates opioid reinforcement, reward, and opioid-induced neuroadaptation. We propose that DRD1 polymorphism affects susceptibility to opioid dependence (OD, the efficiency of transition to OD, and opioid-induced pleasure response. We analyzed potential association between seven DRD1 polymorphisms with the following traits: duration of transition from the first use to dependence (DTFUD, subjective pleasure responses to opioid on first use and post-dependence use, and OD risk in 425 Chinese with OD and 514 healthy controls. DTFUD and level of pleasure responses were examined using a semi-structured interview. The DTFUD of opioid addicts ranged from 5 days to 11 years. Most addicts (64.0% reported non-comfortable response upon first opioid use, while after dependence, most addicts (53.0% felt strong opioid-induced pleasure. Survival analysis revealed a correlation of prolonged DTFUD with the minor allele-carrying genotypes of DRD1 rs4532 (hazard ratios (HR = 0.694; p = 0.001 and rs686 (HR = 0.681, p = 0.0003. Binary logistic regression indicated that rs10063995 GT genotype (vs. GG+TT, OR = 0.261 could predict decreased pleasure response to first-time use and the minor alleles of rs686 (OR = 0.535 and rs4532 (OR = 0.537 could predict decreased post-dependence pleasure. Moreover, rs686 minor allele was associated with a decreased risk for rapid transition from initial use to dependence (DTFUD≤30 days; OR = 0.603 or post-dependence euphoria (OR = 0.603 relative to major allele. In conclusion, DRD1 rs686 minor allele decreases the OD risk by prolonging the transition to dependence and attenuating opioid-induced pleasure in Chinese.

  15. Mu opioid receptor availability in people with psychiatric disorders who died by suicide: a case control study.

    Science.gov (United States)

    Scarr, Elizabeth; Money, Tammie Terese; Pavey, Geoffrey; Neo, Jaclyn; Dean, Brian

    2012-08-28

    Mu opioid receptors have previously been shown to be altered in people with affective disorders who died as a result of suicide. We wished to determine whether these changes were more widespread and independent of psychiatric diagnoses. Mu receptor levels were determined using [3 H]DAMGO binding in BA24 from 51 control subjects; 38 people with schizophrenia (12 suicides); 20 people with major depressive disorder (15 suicides); 13 people with bipolar disorder (5 suicides) and 9 people who had no history of psychiatric disorders but who died as a result of suicide. Mu receptor levels were further determined in BA9 and caudate-putamen from 38 people with schizophrenia and 20 control subjects using [3 H]DAMGO binding and, in all three regions, using Western blots. Data was analysed using one-way ANOVAs with Bonferroni's Multiple Comparison Test or, where data either didn't approximate to a binomial distribution or the sample size was too small to determine distribution, a Kruskal-Wallis test with Dunn's Multiple Comparison Test. [3 H]DAMGO binding density was lower in people who had died as a result of suicide (pendogenous ligands occupying these receptors.

  16. Mu and delta opioid receptors oppositely regulate motor impulsivity in the signaled nose poke task.

    Directory of Open Access Journals (Sweden)

    Mary C Olmstead

    Full Text Available Impulsivity is a primary feature of many psychiatric disorders, most notably attention deficit hyperactivity disorder and drug addiction. Impulsivity includes a number of processes such as the inability to delay gratification, the inability to withhold a motor response, or acting before all of the relevant information is available. These processes are mediated by neural systems that include dopamine, serotonin, norepinephrine, glutamate and cannabinoids. We examine, for the first time, the role of opioid systems in impulsivity by testing whether inactivation of the mu- (Oprm1 or delta- (Oprd1 opioid receptor gene alters motor impulsivity in mice. Wild-type and knockout mice were examined on either a pure C57BL6/J (BL6 or a hybrid 50% C57Bl/6J-50% 129Sv/pas (HYB background. Mice were trained to respond for sucrose in a signaled nose poke task that provides independent measures of associative learning (responses to the reward-paired cue and motor impulsivity (premature responses. Oprm1 knockout mice displayed a remarkable decrease in motor impulsivity. This was observed on the two genetic backgrounds and did not result from impaired associative learning, as responses to the cue signaling reward did not differ across genotypes. Furthermore, mutant mice were insensitive to the effects of ethanol, which increased disinhibition and decreased conditioned responding in wild-type mice. In sharp contrast, mice lacking the Oprd1 gene were more impulsive than controls. Again, mutant animals showed no deficit in associative learning. Ethanol completely disrupted performance in these animals. Together, our results suggest that mu-opioid receptors enhance, whereas delta-opioid receptors inhibit, motor impulsivity. This reveals an unanticipated contribution of endogenous opioid receptor activity to disinhibition. In a broader context, these data suggest that alterations in mu- or delta-opioid receptor function may contribute to impulse control disorders.

  17. Anti-analgesic effect of the mu/delta opioid receptor heteromer revealed by ligand-biased antagonism.

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    Laura Milan-Lobo

    Full Text Available Delta (DOR and mu opioid receptors (MOR can complex as heteromers, conferring functional properties in agonist binding, signaling and trafficking that can differ markedly from their homomeric counterparts. Because of these differences, DOR/MOR heteromers may be a novel therapeutic target in the treatment of pain. However, there are currently no ligands selective for DOR/MOR heteromers, and, consequently, their role in nociception remains unknown. In this study, we used a pharmacological opioid cocktail that selectively activates and stabilizes the DOR/MOR heteromer at the cell surface by blocking its endocytosis to assess its role in antinociception. We found that mice treated chronically with this drug cocktail showed a significant right shift in the ED50 for opioid-mediated analgesia, while mice treated with a drug that promotes degradation of the heteromer did not. Furthermore, promoting degradation of the DOR/MOR heteromer after the right shift in the ED50 had occurred, or blocking signal transduction from the stabilized DOR/MOR heteromer, shifted the ED50 for analgesia back to the left. Taken together, these data suggest an anti-analgesic role for the DOR/MOR heteromer in pain. In conclusion, antagonists selective for DOR/MOR heteromer could provide an avenue for alleviating reduced analgesic response during chronic pain treatment.

  18. Panicolytic-like effect of tramadol is mediated by opioid receptors in the dorsal periaqueductal grey.

    Science.gov (United States)

    Fiaes, Gislaine Cardoso de Souza; Roncon, Camila Marroni; Sestile, Caio Cesar; Maraschin, Jhonatan Christian; Souza, Rodolfo Luis Silva; Porcu, Mauro; Audi, Elisabeth Aparecida

    2017-05-30

    Tramadol is a synthetic opioid prescribed for the treatment of moderate to severe pain, acting as agonist of μ-opioid receptors and serotonin (5-HT) and noradrenaline (NE) reuptake inhibitor. This study evaluated the effects of tramadol in rats submitted to the elevated T-maze (ETM), an animal model that evaluates behavioural parameters such as anxiety and panic. Male Wistar rats were intraperitoneally (i.p.) treated acutely with tramadol (16 and 32mg/kg) and were submitted to the ETM. Tramadol (32mg/kg) promoted a panicolytic-like effect. Considering that dorsal periaqueductal grey (dPAG) is the main brain structure related to the pathophysiology of panic disorder (PD), this study also evaluated the participation of 5-HT and opioid receptors located in the dPAG in the panicolytic-like effect of tramadol. Seven days after stereotaxic surgery for implantation of a cannula in the dPAG, the animals were submitted to the test. To assess the involvement of 5-HT 1A receptors on the effect of tramadol, we combined the 5-HT 1A receptor antagonist, WAY100635 (0.37nmol), microinjected intra-dPAG, 10min prior to the administration of tramadol (32mg/kg, i.p.). WAY100635 did not block the panicolytic-like effect of tramadol. We also associated the non-selective opioid receptor antagonist, naloxone, systemically (1mg/kg, i.p.) or intra-dPAG (0.5nmol) administered 10min prior to tramadol (32mg/kg, i.p.). Naloxone blocked the panicolytic-like effect of tramadol in both routes of administrations, showing that tramadol modulates acute panic defensive behaviours through its interaction with opioid receptors located in the dPAG. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Kappa-opioid receptors mediate the antidepressant-like activity of hesperidin in the mouse forced swimming test.

    Science.gov (United States)

    Filho, Carlos B; Del Fabbro, Lucian; de Gomes, Marcelo G; Goes, André T R; Souza, Leandro C; Boeira, Silvana P; Jesse, Cristiano R

    2013-01-05

    The opioid system has been implicated as a contributing factor for major depression and is thought to play a role in the mechanism of action of antidepressants. This study investigated the involvement of the opioid system in the antidepressant-like effect of hesperidin in the mouse forced swimming test. Our results demonstrate that hesperidin (0.1, 0.3 and 1 mg/kg; intraperitoneal) decreased the immobility time in the forced swimming test without affecting locomotor activity in the open field test. The antidepressant-like effect of hesperidin (0.3 mg/kg) in the forced swimming test was prevented by pretreating mice with naloxone (1 mg/kg, a nonselective opioid receptor antagonist) and 2-(3,4-dichlorophenyl)-Nmethyl-N-[(1S)-1-(3-isothiocyanatophenyl)-2-(1-pyrrolidinyl)ethyl] acetamide (DIPPA (1 mg/kg), a selective κ-opioid receptor antagonist), but not with naloxone methiodide (1 mg/kg, a peripherally acting opioid receptor antagonist), naltrindole (3 mg/kg, a selective δ-opioid receptor antagonist), clocinnamox (1 mg/kg, a selective μ-opioid receptor antagonist) or caffeine (3 mg/kg, a nonselective adenosine receptor antagonist). In addition, a sub-effective dose of hesperidin (0.01 mg/kg) produced a synergistic antidepressant-like effect in the forced swimming test when combined with a sub-effective dose of morphine (1 mg/kg). The antidepressant-like effect of hesperidin in the forced swimming test on mice was dependent on its interaction with the κ-opioid receptor, but not with the δ-opioid, μ-opioid or adenosinergic receptors. Taken together, these results suggest that hesperidin possesses antidepressant-like properties and may be of interest as a therapeutic agent for the treatment of depressive disorders. Published by Elsevier B.V.

  20. δ-opioid receptor and somatostatin receptor-4 heterodimerization: possible implications in modulation of pain associated signaling.

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    Rishi K Somvanshi

    Full Text Available Pain relief is the principal action of opioids. Somatostatin (SST, a growth hormone inhibitory peptide is also known to alleviate pain even in cases when opioids fail. Recent studies have shown that mice are prone to sustained pain and devoid of analgesic effect in the absence of somatostatin receptor 4 (SSTR4. In the present study, using brain slices, cultured neurons and HEK-293 cells, we showed that SSTR4 and δ-Opioid receptor (δOR exist in a heteromeric complex and function in synergistic manner. SSTR4 and δOR co-expressed in cortical/striatal brain regions and spinal cord. Using cultured neuronal cells, we describe the heterogeneous complex formation of SSTR4 and δOR at neuronal cell body and processes. Cotransfected cells display inhibition of cAMP/PKA and co-activation of SSTR4 and δOR oppose receptor trafficking induced by individual receptor activation. Furthermore, downstream signaling pathways either associated with withdrawal or pain relief are modulated synergistically with a predominant role of SSTR4. Inhibition of cAMP/PKA and activation of ERK1/2 are the possible cellular adaptations to prevent withdrawal induced by chronic morphine use. Our results reveal direct intra-membrane interaction between SSTR4 and δOR and provide insights for the molecular mechanism for the anti-nociceptive property of SST in combination with opioids as a potential therapeutic approach to avoid undesirable withdrawal symptoms.

  1. Kappa Opioid Receptors Mediate where Fear Is Expressed Following Extinction Training

    Science.gov (United States)

    Cole, Sindy; Richardson, Rick; McNally, Gavan P.

    2011-01-01

    Six experiments used a within-subjects renewal design to examine the involvement of kappa opioid receptors (KORs) in regulating the expression and recovery of extinguished fear. Rats were trained to fear a tone conditioned stimulus (CS) via pairings with foot shock in a distinctive context (A). This was followed by extinction training of the CS in…

  2. Opioid regulation of mu receptor internalisation: relevance to the development of tolerance and dependence.

    Science.gov (United States)

    Lopez-Gimenez, Juan F; Milligan, Graeme

    2010-11-01

    Internalisation of the mu opioid receptor from the surface of cells is generally achieved by receptor occupancy with agonist ligands of high efficacy. However, in many situations the potent analgesic morphine fails to promote internalisation effectively and whether there is a direct link between this and the propensity for the sustained use of morphine to result in both tolerance and dependence has been studied intensely. Although frequently described as a partial agonist, this characteristic appears insufficient to explain the poor capacity of morphine to promote internalisation of the mu opioid receptor. Experiments performed using both transfected cell systems and ex vivo/in vivo models have provided evidence that when morphine can promote internalisation of the mu receptor there is a decrease in the development of tolerance and dependence. Although aspects of this model are controversial, such observations suggest a number of approaches to further enhance the use of morphine as an analgesic.

  3. Neuroanatomical patterns of the mu, delta, and kappa opioid receptors of rat brain as determined by quantitative in vitro autoradiography

    International Nuclear Information System (INIS)

    Tempel, A.; Zukin, R.S.

    1987-01-01

    Highly specific radioligands and quantitative autoradiography reveal strikingly different neuroanatomical patterns for the mu, delta, and kappa opioid receptors of rat brain. The mu receptors are most densely localized in patches in the striatum, layers I and III of the cortex, the pyramidal cell layer of the hippocampal formation, specific nuclei of the thalamus, the pars reticulata of the substantia nigra, the interpeduncular nucleus, and the locus coeruleus. In contrast, delta receptors are highly confined, exhibiting selective localization in layers I, II, and VIa of the neocortex, a diffuse pattern in the striatum, and moderate concentration in the pars reticulata of the substantia nigra and in the interpeduncular nucleus. delta receptors are absent in most other brain structures. This distribution is unexpected in that the enkephalins, the putative endogenous ligands of the delta receptor, occur essentially throughout the brain. The kappa receptors of rat brain exhibit a third pattern distinct from that of the mu and delta receptors. kappa receptors occur at low density in patches in the striatum and at particularly high density in the nucleus accumbens, along the pyramidal and molecular layers of the hippocampus, in the granular cell layer of the dentate gyrus, specific midline nuclei of the thalamus, and hindbrain regions. kappa receptors appear to be uniformly distributed across regions in the neocortex with the exception of layer III, which revealed only trace levels of binding. An important conclusion of the present study is that delta receptors occur at high density only in the forebrain and in two midbrain structures, whereas mu and kappa receptors exhibit discrete patterns in most major brain regions

  4. Dopamine D4 Receptor Counteracts Morphine-Induced Changes in µ Opioid Receptor Signaling in the Striosomes of the Rat Caudate Putamen

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    Diana Suárez-Boomgaard

    2014-01-01

    Full Text Available The mu opioid receptor (MOR is critical in mediating morphine analgesia. However, prolonged exposure to morphine induces adaptive changes in this receptor leading to the development of tolerance and addiction. In the present work we have studied whether the continuous administration of morphine induces changes in MOR protein levels, its pharmacological profile, and MOR-mediated G-protein activation in the striosomal compartment of the rat CPu, by using immunohistochemistry and receptor and DAMGO-stimulated [35S]GTPγS autoradiography. MOR immunoreactivity, agonist binding density and its coupling to G proteins are up-regulated in the striosomes by continuous morphine treatment in the absence of changes in enkephalin and dynorphin mRNA levels. In addition, co-treatment of morphine with the dopamine D4 receptor (D4R agonist PD168,077 fully counteracts these adaptive changes in MOR, in spite of the fact that continuous PD168,077 treatment increases the [3H]DAMGO Bmax values to the same degree as seen after continuous morphine treatment. Thus, in spite of the fact that both receptors can be coupled to Gi/0 protein, the present results give support for the existence of antagonistic functional D4R-MOR receptor-receptor interactions in the adaptive changes occurring in MOR of striosomes on continuous administration of morphine.

  5. Functional μ-Opioid-Galanin Receptor Heteromers in the Ventral Tegmental Area.

    Science.gov (United States)

    Moreno, Estefanía; Quiroz, César; Rea, William; Cai, Ning-Sheng; Mallol, Josefa; Cortés, Antoni; Lluís, Carme; Canela, Enric I; Casadó, Vicent; Ferré, Sergi

    2017-02-01

    The neuropeptide galanin has been shown to interact with the opioid system. More specifically, galanin counteracts the behavioral effects of the systemic administration of μ-opioid receptor (MOR) agonists. Yet the mechanism responsible for this galanin-opioid interaction has remained elusive. Using biophysical techniques in mammalian transfected cells, we found evidence for selective heteromerization of MOR and the galanin receptor subtype Gal1 (Gal1R). Also in transfected cells, a synthetic peptide selectively disrupted MOR-Gal1R heteromerization as well as specific interactions between MOR and Gal1R ligands: a negative cross talk, by which galanin counteracted MAPK activation induced by the endogenous MOR agonist endomorphin-1, and a cross-antagonism, by which a MOR antagonist counteracted MAPK activation induced by galanin. These specific interactions, which represented biochemical properties of the MOR-Gal1R heteromer, could then be identified in situ in slices of rat ventral tegmental area (VTA) with MAPK activation and two additional cell signaling pathways, AKT and CREB phosphorylation. Furthermore, in vivo microdialysis experiments showed that the disruptive peptide selectively counteracted the ability of galanin to block the dendritic dopamine release in the rat VTA induced by local infusion of endomorphin-1, demonstrating a key role of MOR-Gal1R heteromers localized in the VTA in the direct control of dopamine cell function and their ability to mediate antagonistic interactions between MOR and Gal1R ligands. The results also indicate that MOR-Gal1R heteromers should be viewed as targets for the treatment of opioid use disorders. The μ-opioid receptor (MOR) localized in the ventral tegmental area (VTA) plays a key role in the reinforcing and addictive properties of opioids. With parallel in vitro experiments in mammalian transfected cells and in situ and in vivo experiments in rat VTA, we demonstrate that a significant population of these MORs form

  6. Spinal antinociceptive effects of [D-Ala2]deltorphin II, a novel and highly selective delta-opioid receptor agonist.

    Science.gov (United States)

    Improta, G; Broccardo, M

    1992-01-01

    Pharmacological assays in isolated tissues and binding tests have recently shown that two peptides, with the sequence Tyr-D-Ala-Phe-Asp-(or Glu)- Val-Val-Gly-NH2, isolated from skin extracts of Phyllomedusa bicolor and named [D-Ala2]deltorphin I and II, respectively, possess a higher affinity and selectivity for delta-opioid receptors than any other known natural compound. Since much evidence supports the role of spinal delta-opioid sites in producing antinociceptive effects, we investigated whether analgesia might be detected by direct spinal cord administration of [D-Ala2]deltorphin II (DADELT II) in the rat. The thermal antinociceptive effects of intrathecal DADELT II and dermorphin, a potent mu-selective agonist, were compared at different postinjection times by means of the tail-flick test. The DADELT II produced a dose-related inhibition of the tail-flick response, which lasted 10-60 min depending on the dose and appeared to be of shorter duration than the analgesia produced in rats after intrathecal injection of dermorphin (20-120 min). The analgesic effect of infused or injected DADELT II was completely abolished by naltrindole, the highly selective delta antagonist. These results confirm the involvement of delta receptors in spinal analgesic activity in the rat.

  7. ``In silico'' study of the binding of two novel antagonists to the nociceptin receptor

    Science.gov (United States)

    Della Longa, Stefano; Arcovito, Alessandro

    2018-02-01

    Antagonists of the nociceptin receptor (NOP) are raising interest for their possible clinical use as antidepressant drugs. Recently, the structure of NOP in complex with some piperidine-based antagonists has been revealed by X-ray crystallography. In this study, a multi-flexible docking (MF-docking) procedure, i.e. docking to multiple receptor conformations extracted by preliminary molecular dynamics trajectories, together with hybrid quantum mechanics/molecular mechanics (QM/MM) simulations have been carried out to provide the binding mode of two novel NOP antagonists, one of them selective (BTRX-246040, formerly named LY-2940094) and one non selective (AT-076), i.e. able to inactivate NOP as well as the classical µ- k- and δ-opioid receptors (MOP KOP and DOP). According to our results, the pivotal role of residue D1303,32 (upper indexes are Ballesteros-Weinstein notations) is analogous to that enlighten by the already known X-ray structures of opioid receptors: binding of the molecules are predicted to require a slight readjustment of the hydrophobic pocket (residues Y1313,33, M1343,36, I2195,43, Q2806,52 and V2836,55) in the orthosteric site of NOP, accommodating either the pyridine-pyrazole (BTRX-246040) or the isoquinoline (AT-076) moiety of the ligand, in turn allowing the protonated piperidine nitrogen to maximize interaction (salt-bridge) with residue D1303,32 of the NOP, and the aromatic head to be sandwiched in optimal π-stacking between Y1313,33 and M1343,36. The QM/MM optimization after the MF-docking procedure has provided the more likely conformations for the binding to the NOP receptor of BTRX-246040 and AT-076, based on different pharmacophores and exhibiting different selectivity profiles. While the high selectivity for NOP of BTRX-246040 can be explained by interactions with NOP specific residues, the lack of selectivity of AT-076 could be associated to its ability to penetrate into the deep hydrophobic pocket of NOP, while retaining a

  8. An investigation into the receptor-regulating effects of the acute administration of opioid agonists and an antagonist on beta adrenergic receptors in the rat cerebral cortex

    International Nuclear Information System (INIS)

    Roper, I.

    1987-01-01

    Past and current research indicated that biochemical deviations which might be involved in the etiology and pathophysiology of depression, included abnormalities or imbalances in the noradrenergic, serotonergic, hormonal and possibly in the endogenous opioid, dopaminergic, histaminergic, cholinergic and trace amine systems. In order to investigate a possible link between the noradrenergic system and opioids, it was decided to test the acute effects of opioid administration on cortical beta adrenoceptor numbers and affinity. As these receptors have been most consistently downregulated by antidepressant treatment, they may be involved in the mechanism of antidepressant action of these agents. It was decided to investigate beta adrenoceptor-regulatory effects of opioid treatment. Naloxone was tested alone, with a view to suppressing any possible endogenous opioid influences upon beta receptor status and revealing an effect which would possibly be the opposite of that brought about by the administration of opioid agonists. Naloxone was administered together with morphine to demonstrate that any beta receptor up- or downregulation which might be measured, had indeed been opioid-receptor mediated. It was found that the acute administration of four different mu opioid agonists, naloxone and naloxone plus morphine, did not cause any statistically significant alterations in cortical beta adrenergic receptor numbers or affinity in the rat. A radioactive ligand, the beta adrenoceptor-labelling compound referred to as DHA (L-dihydroalprenolol HCI) was used in this study

  9. μ opioid receptor activation hyperpolarizes respiratory-controlling Kölliker-Fuse neurons and suppresses post-inspiratory drive.

    Science.gov (United States)

    Levitt, Erica S; Abdala, Ana P; Paton, Julian F R; Bissonnette, John M; Williams, John T

    2015-10-01

    In addition to reductions in respiratory rate, opioids also cause aspiration and difficulty swallowing, indicating impairment of the upper airways. The Kölliker-Fuse (KF) maintains upper airway patency and a normal respiratory pattern. In this study, activation of μ opioid receptors in the KF reduced respiratory frequency and tidal volume in anaesthetized rats. Nerve recordings in an in situ preparation showed that activation of μ opioid receptors in the KF eliminated the post-inspiration phase of the respiratory cycle. In brain slices, μ opioid agonists hyperpolarized a distinct population (61%) of KF neurons by activation of an inwardly rectifying potassium conductance. These results suggest that KF neurons that are hyperpolarized by opioids could contribute to opioid-induced respiratory disturbances, particularly the impairment of upper airways. Opioid-induced respiratory effects include aspiration and difficulty swallowing, suggesting impairment of the upper airways. The pontine Kölliker-Fuse nucleus (KF) controls upper airway patency and regulates respiration, in particular the inspiratory/expiratory phase transition. Given the importance of the KF in coordinating respiratory pattern, the mechanisms of μ opioid receptor activation in this nucleus were investigated at the systems and cellular level. In anaesthetized, vagi-intact rats, injection of opioid agonists DAMGO or [Met(5) ]enkephalin (ME) into the KF reduced respiratory frequency and amplitude. The μ opioid agonist DAMGO applied directly into the KF of the in situ arterially perfused working heart-brainstem preparation of rat resulted in robust apneusis (lengthened low amplitude inspiration due to loss of post-inspiratory drive) that was rapidly reversed by the opioid antagonist naloxone. In brain slice preparations, activation of μ opioid receptors on KF neurons hyperpolarized a distinct population (61%) of neurons. As expected, the opioid-induced hyperpolarization reduced the excitability of

  10. Distribution of kappa opioid receptors in the brain of young and old male rats

    International Nuclear Information System (INIS)

    Maggi, R.; Limonta, P.; Dondi, D.; Martini, L.; Piva, F.

    1989-01-01

    The experiments to be described have been designed in order to: (a) provide new information on the concentrations of opioid kappa receptors in different regions of the brain of the male rats; and (b) to analyze whether the density of brain kappa receptors might be modified by the process of aging. The concentration of kappa receptors was investigated in the hypothalamus, amygdala, mesencephalon, corpus striatum, hippocampus, thalamus, frontal poles, anterior and posterior cortex collected from male rats of 2 and 19 months of age. 3 H-bremazocine (BRZ) was used as the ligand of kappa receptors, after protection of mu and delta receptors respectively with dihydromorphine and d-ala-d-leu-enkephalin. The results obtained show that: (1) in young male rats, the number of kappa opioid receptors is different in the various brain areas examined. (2) Aging exerts little influence on the number of kappa receptors in the majority of the brain structures considered. However in the amygdala and in the thalamus the number of kappa receptors was increased in old animals

  11. Sex and age-dependent effects of a maternal junk food diet on the mu-opioid receptor in rat offspring.

    Science.gov (United States)

    Gugusheff, Jessica R; Bae, Sung Eun; Rao, Alexandra; Clarke, Iain J; Poston, Lucilla; Taylor, Paul D; Coen, Clive W; Muhlhausler, Beverly S

    2016-03-15

    Perinatal junk food exposure increases the preference for palatable diets in juvenile and adult rat offspring. Previous studies have implicated reduced sensitivity of the opioid pathway in the programming of food preferences; however it is not known when during development these changes in opioid signalling first emerge. This study aimed to determine the impact of a maternal junk food (JF) diet on mu-opioid receptor (MuR) expression and ligand binding in two key regions of the reward pathway, the nucleus accumbens (NAc) and the ventral tegmental area (VTA) in rats during the early suckling (postnatal day (PND) 1 and 7) and late suckling/early post-weaning (PND 21 and 28) periods. Female rats were fed either a JF or a control diet for two weeks prior to mating and throughout pregnancy and lactation. MuR expression in the VTA was significantly reduced in female JF offspring on PND 21 and 28 (by 32% and 57% respectively, Pjunk food exposure on MuR mRNA expression or binding were detected at these time points in male offspring. These findings provide evidence that the opioid signalling system is a target of developmental programming by the end of the third postnatal week in females, but not in males. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Interacting cannabinoid and opioid receptors in the nucleus accumbens core control adolescent social play

    Directory of Open Access Journals (Sweden)

    Antonia Manduca

    2016-11-01

    Full Text Available Social play behavior is a highly rewarding, developmentally important form of social interaction in young mammals. However, its neurobiological underpinnings remain incompletely understood. Previous work has suggested that opioid and endocannabinoid neurotransmission interact in the modulation of social play. Therefore, we combined behavioral, pharmacological, electrophysiological and genetic approaches to elucidate the role of the endocannabinoid 2-arachidonoylglycerol (2-AG in social play, and how cannabinoid and opioid neurotransmission interact to control social behavior in adolescent rodents. Systemic administration of the 2-AG hydrolysis inhibitor JZL184 or the opioid receptor agonist morphine increased social play behavior in adolescent rats. These effects were blocked by systemic pretreatment with either CB1 cannabinoid receptor (CB1R or mu-opioid receptor (MOR antagonists. The social play-enhancing effects of systemic morphine or JZL184 treatment were also prevented by direct infusion of the CB1R antagonist SR141716 and the MOR antagonist naloxone into the nucleus accumbens core (NAcC. Searching for synaptic correlates of these effects in adolescent NAcC excitatory synapses, we observed that CB1R antagonism blocked the effect of the MOR agonist DAMGO and, conversely, that naloxone reduced the effect of a cannabinoid agonist. These results were recapitulated in mice, and completely abolished in CB1R and MOR knockout mice, suggesting that the functional interaction between CB1R and MOR in the NAcC in the modulation of mediates social behavior is widespread in rodents. The data shed new light on the mechanism by which endocannabinoid lipids and opioid peptides interact to orchestrate rodent socioemotional behaviors.

  13. Ghrelin interacts with neuropeptide Y Y1 and opioid receptors to increase food reward.

    Science.gov (United States)

    Skibicka, Karolina P; Shirazi, Rozita H; Hansson, Caroline; Dickson, Suzanne L

    2012-03-01

    Ghrelin, a stomach-derived hormone, is an orexigenic peptide that was recently shown to potently increase food reward behavior. The neurochemical circuitry that links ghrelin to the mesolimbic system and food reward behavior remains unclear. Here we examined the contribution of neuropeptide Y (NPY) and opioids to ghrelin's effects on food motivation and intake. Both systems have well-established links to the mesolimbic ventral tegmental area (VTA) and reward/motivation control. NPY mediates the effect of ghrelin on food intake via activation of NPY-Y1 receptor (NPY-Y1R); their connection with respect to motivated behavior is unexplored. The role of opioids in any aspect of ghrelin's action on food-oriented behaviors is unknown. Rats were trained in a progressive ratio sucrose-induced operant schedule to measure food reward/motivation behavior. Chow intake was measured immediately after the operant test. In separate experiments, we explored the suppressive effects of a selective NPY-Y1R antagonist or opioid receptor antagonist naltrexone, injected either intracerebroventricularly or intra-VTA, on ghrelin-induced food reward behavior. The ventricular ghrelin-induced increase in sucrose-motivated behavior and chow intake were completely blocked by intracerebroventricular pretreatment with either an NPY-Y1R antagonist or naltrexone. The intra-VTA ghrelin-induced sucrose-motivated behavior was blocked only by intra-VTA naltrexone. In contrast, the intra-VTA ghrelin-stimulated chow intake was attenuated only by intra-VTA NPY-Y1 blockade. Finally, ghrelin infusion was associated with an elevated VTA μ-opioid receptor expression. Thus, we identify central NPY and opioid signaling as the necessary mediators of food intake and reward effects of ghrelin and localize these interactions to the mesolimbic VTA.

  14. The effect of opioid receptor blockade on the neural processing of thermal stimuli.

    Directory of Open Access Journals (Sweden)

    Eszter D Schoell

    Full Text Available The endogenous opioid system represents one of the principal systems in the modulation of pain. This has been demonstrated in studies of placebo analgesia and stress-induced analgesia, where anti-nociceptive activity triggered by pain itself or by cognitive states is blocked by opioid antagonists. The aim of this study was to characterize the effect of opioid receptor blockade on the physiological processing of painful thermal stimulation in the absence of cognitive manipulation. We therefore measured BOLD (blood oxygen level dependent signal responses and intensity ratings to non-painful and painful thermal stimuli in a double-blind, cross-over design using the opioid receptor antagonist naloxone. On the behavioral level, we observed an increase in intensity ratings under naloxone due mainly to a difference in the non-painful stimuli. On the neural level, painful thermal stimulation was associated with a negative BOLD signal within the pregenual anterior cingulate cortex, and this deactivation was abolished by naloxone.

  15. Analgesia produced by exposure to 2450-MHz radiofrequency radiation (RFR) is mediated by brain mu- and kappa-opioid receptors

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    Salomon, G.; Park, E.J.; Quock, R.M. (Univ. of Illinois, Rockford (United States))

    1992-02-26

    This study was conducted to identify the opioid receptor subtype(s) responsible for RFR-induced analgesia. Male Swiss Webster mice, 20-25 g, were exposed to 20 mW/cm{sup 2} RFR in a 2,450-MHz waveguide system for 10 min, then tested 15 min later in the abdominal constriction paradigm which detects {mu}- and {kappa}-opioid activity. Immediately following RFR exposure, different groups of mice were pretreated intracerebroventricularly with different opioid receptor blockers with selectivity for {mu}- or {kappa}-opioid receptors. Results show that RFR-induced analgesia was attenuated by higher but not lower doses of the non-selective antagonist naloxone, but the selective {mu}-opioid antagonist {beta}-funaltrexamine and by the selective {kappa}-opioid antagonist norbinaltorphimine. RFR-induced analgesia was also reduced by subcutaneous pretreatment with 5.0 mg/kg of the {mu}-/{kappa}-opioid antagonist({minus})-5,9-diethyl-{alpha}-5,9-dialkyl-2{prime}-hydroxy-6,7-benzomorphan(MR-2266). These findings suggest that RFR-induced analgesia may be mediated by both {mu}- and {kappa}-opioid mechanisms.

  16. Cell-Autonomous Regulation of Mu-Opioid Receptor Recycling by Substance P

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    Shanna L. Bowman

    2015-03-01

    Full Text Available How neurons coordinate and reprogram multiple neurotransmitter signals is an area of broad interest. Here, we show that substance P (SP, a neuropeptide associated with inflammatory pain, reprograms opioid receptor recycling and signaling. SP, through activation of the neurokinin 1 (NK1R receptor, increases the post-endocytic recycling of the mu-opioid receptor (MOR in trigeminal ganglion (TG neurons in an agonist-selective manner. SP-mediated protein kinase C (PKC activation is both required and sufficient for increasing recycling of exogenous and endogenous MOR in TG neurons. The target of this cross-regulation is MOR itself, given that mutation of either of two PKC phosphorylation sites on MOR abolishes the SP-induced increase in recycling and resensitization. Furthermore, SP enhances the resensitization of fentanyl-induced, but not morphine-induced, antinociception in mice. Our results define a physiological pathway that cross-regulates opioid receptor recycling via direct modification of MOR and suggest a mode of homeostatic interaction between the pain and analgesic systems.

  17. Modelling the PKPD of oxycodone in experimental pain - impact of opioid receptor polymorphisms

    DEFF Research Database (Denmark)

    Olsen, Rasmus; Foster, David J R; Upton, Richard N

    2016-01-01

    BACKGROUND: Polymorphisms in the opioid receptor genes may affect the pharmacodynamics (PD) of oxycodone and be part of the reason behind the diversity in clinical response. The aim of the analysis was to model the exposure-response profile of oxycodone for three different pain variables and search...... for genetic covariates. Model simulations were used to predict how population and effect-size impact the power to detect clinical significant SNPs. METHOD: The population pharmacokinetic-pharmacodynamic (PKPD) model of oral single-dosed oxycodone was based on pooled data from three published studies...... in healthy volunteers. Pain tolerance data from muscle pressure (n=36), visceral pressure (n=54) and skin pinch (n=34) were included. Genetic associations with 18 opioid-receptor SNPs were explored using a stepwise covariate approach. Model simulations were performed using the estimated model parameters...

  18. 2012 David W. Robertson Award for Excellence in Medicinal Chemistry: Neoclerodanes as Atypical Opioid Receptor Ligands⊥

    Science.gov (United States)

    Prisinzano, Thomas E.

    2013-01-01

    The neoclerodane diterpene salvinorin A is the major active component of the hallucinogenic mint plant Salvia divinorum Epling & Játiva (Lamiaceae). Since the finding that salvinorin A exerts its potent psychotropic actions through the activation of opioid receptors, the site of action of morphine and related analogues, there has been much interest in elucidating the underlying mechanisms behind its effects. These effects are particularly remarkable, because (1) salvinorin A is the first reported non-nitrogenous opioid receptor agonist, and (2) its effects are not mediated through the previously investigated targets of psychotomimetics. This perspective outlines our research program, illustrating a new direction to the development of tools to further elucidate the biological mechanisms of drug tolerance and dependence. The information gained from these efforts is expected to facilitate the design of novel agents to treat pain, drug abuse, and other CNS disorders. PMID:23548164

  19. On the Modularity of the Intrinsic Flexibility of the µ Opioid Receptor: A Computational Study

    Science.gov (United States)

    Fossépré, Mathieu; Leherte, Laurence; Laaksonen, Aatto; Vercauteren, Daniel P.

    2014-01-01

    The µ opioid receptor (µOR), the principal target to control pain, belongs to the G protein-coupled receptors (GPCRs) family, one of the most highlighted protein families due to their importance as therapeutic targets. The conformational flexibility of GPCRs is one of their essential characteristics as they take part in ligand recognition and subsequent activation or inactivation mechanisms. It is assessed that the intrinsic mechanical properties of the µOR, more specifically its particular flexibility behavior, would facilitate the accomplishment of specific biological functions, at least in their first steps, even in the absence of a ligand or any chemical species usually present in its biological environment. The study of the mechanical properties of the µOR would thus bring some indications regarding the highly efficient ability of the µOR to transduce cellular message. We therefore investigate the intrinsic flexibility of the µOR in its apo-form using all-atom Molecular Dynamics simulations at the sub-microsecond time scale. We particularly consider the µOR embedded in a simplified membrane model without specific ions, particular lipids, such as cholesterol moieties, or any other chemical species that could affect the flexibility of the µOR. Our analyses highlighted an important local effect due to the various bendability of the helices resulting in a diversity of shape and volume sizes adopted by the µOR binding site. Such property explains why the µOR can interact with ligands presenting highly diverse structural geometry. By investigating the topology of the µOR binding site, a conformational global effect is depicted: the correlation between the motional modes of the extra- and intracellular parts of µOR on one hand, along with a clear rigidity of the central µOR domain on the other hand. Our results show how the modularity of the µOR flexibility is related to its pre-ability to activate and to present a basal activity. PMID:25549261

  20. Ivy and neurogliaform interneurons are a major target of μ opioid receptor modulation

    OpenAIRE

    Krook-Magnuson, Esther; Luu, Lillian; Lee, Sang-Hun; Varga, Csaba; Soltesz, Ivan

    2011-01-01

    Mu opioid receptors (μORs) are selectively expressed on interneurons in area CA1 of the hippocampus. Fast-spiking, parvalbumin expressing, basket cells express μORs, but circumstantial evidence suggests that another major, unidentified, GABAergic cell class must also be modulated by μORs. Here we report that the abundant, dendritically targeting, neurogliaform family of cells (Ivy and neurogliaform cells) is a previously unrecognized target of direct modulation by μORs. Ivy and neurogliaform ...

  1. Sex Differences in Kappa Opioid Receptor Function and Their Potential Impact on Addiction

    OpenAIRE

    Chartoff, Elena H.; Mavrikaki, Maria

    2015-01-01

    Behavioral, biological, and social sequelae that lead to drug addiction differ between men and women. Our efforts to understand addiction on a mechanistic level must include studies in both males and females. Stress, anxiety, and depression are tightly linked to addiction, and whether they precede or result from compulsive drug use depends on many factors, including biological sex. The neuropeptide dynorphin (DYN), an endogenous ligand at kappa opioid receptors (KORs), is necessary for stress...

  2. Atypical Opioid Mechanisms of Control of Injury-Induced Cutaneous Pain by Delta Receptors

    Science.gov (United States)

    2017-07-01

    i.e. mu opioid receptor agonists such as morphine) cause unacceptable side effects including addiction . Injuries suffered most frequently by active...slides. The slides were then processed for fluorescent in situ hybridization with RNAscope technology (ACD Biosystems) to detect Oprd1 mRNA, as...tissue as done in Bardoni et al., Neuron, 2014) and negative controls (no probe). Controls indicated that the technology and reagents work as expected

  3. Toll-like receptor 9 is required for opioid-induced microglia apoptosis.

    Directory of Open Access Journals (Sweden)

    Lei He

    2011-04-01

    Full Text Available Opioids have been widely applied in clinics as one of the most potent pain relievers for centuries, but their abuse has deleterious physiological effects beyond addiction. However, the underlying mechanism by which microglia in response to opioids remains largely unknown. Here we show that morphine induces the expression of Toll-like receptor 9 (TLR9, a key mediator of innate immunity and inflammation. Interestingly, TLR9 deficiency significantly inhibited morphine-induced apoptosis in microglia. Similar results were obtained when endogenous TLR9 expression was suppressed by the TLR9 inhibitor CpGODN. Inhibition of p38 MAPK by its specific inhibitor SB203580 attenuated morphine-induced microglia apoptosis in wild type microglia. Morphine caused a dramatic decrease in Bcl-2 level but increase in Bax level in wild type microglia, but not in TLR9 deficient microglia. In addition, morphine treatment failed to induce an increased levels of phosphorylated p38 MAPK and MAP kinase kinase 3/6 (MKK3/6, the upstream MAPK kinase of p38 MAPK, in either TLR9 deficient or µ-opioid receptor (µOR deficient primary microglia, suggesting an involvement of MAPK and µOR in morphine-mediated TLR9 signaling. Moreover, morphine-induced TLR9 expression and microglia apoptosis appears to require μOR. Collectively, these results reveal that opioids prime microglia to undergo apoptosis through TLR9 and µOR as well. Taken together, our data suggest that inhibition of TLR9 and/or blockage of µOR is capable of preventing opioid-induced brain damage.

  4. Acute inflammation induces segmental, bilateral, supraspinally mediated opioid release in the rat spinal cord, as measured by μ-opioid receptor internalization

    Science.gov (United States)

    Chen, Wenling; Marvizón, Juan Carlos G.

    2009-01-01

    The objective of this study was to measure opioid release in the spinal cord during acute and long-term inflammation using μ-opioid receptor (MOR) internalization. In particular, we determined whether opioid release occurs in the segments receiving the noxious signals or in the entire spinal cord, and whether it involves supraspinal signals. Internalization of neurokinin 1 receptors (NK1Rs) was measured to track the intensity of the noxious stimulus. Rats received peptidase inhibitors intrathecally to protect opioids from degradation. Acute inflammation of the hindpaw with formalin induced moderate MOR internalization in the L5 segment bilaterally, whereas NK1R internalization occurred only ipsilaterally. MOR internalization was restricted to the lumbar spinal cord, regardless of whether the peptidase inhibitors were injected in a lumbar or thoracic site. Formalin-induced MOR internalization was substantially reduced by isoflurane anesthesia. It was also markedly reduced by a lidocaine block of the cervical-thoracic spinal cord (which did not affect the evoked NK1R internalization) indicating that spinal opioid release is mediated supraspinally. In the absence of peptidase inhibitors, formalin and hindpaw clamp induced a small amount of MOR internalization, which was significantly higher than in controls. To study spinal opioid release during chronic inflammation, we injected Complete Freund's Adjuvant (CFA) in the hindpaw and peptidase inhibitors intrathecally. Two days later, no MOR or NK1R internalization was detected. Furthermore, CFA inflammation decreased MOR internalization induced by clamping the inflamed hindpaw. These results show that acute inflammation, but not chronic inflammation, induce segmental opioid release in the spinal cord that involves supraspinal signals. PMID:19298846

  5. Acute inflammation induces segmental, bilateral, supraspinally mediated opioid release in the rat spinal cord, as measured by mu-opioid receptor internalization.

    Science.gov (United States)

    Chen, W; Marvizón, J C G

    2009-06-16

    The objective of this study was to measure opioid release in the spinal cord during acute and long-term inflammation using mu-opioid receptor (MOR) internalization. In particular, we determined whether opioid release occurs in the segments receiving the noxious signals or in the entire spinal cord, and whether it involves supraspinal signals. Internalization of neurokinin 1 receptors (NK1Rs) was measured to track the intensity of the noxious stimulus. Rats received peptidase inhibitors intrathecally to protect opioids from degradation. Acute inflammation of the hind paw with formalin induced moderate MOR internalization in the L5 segment bilaterally, whereas NK1R internalization occurred only ipsilaterally. MOR internalization was restricted to the lumbar spinal cord, regardless of whether the peptidase inhibitors were injected in a lumbar or thoracic site. Formalin-induced MOR internalization was substantially reduced by isoflurane anesthesia. It was also markedly reduced by a lidocaine block of the cervical-thoracic spinal cord (which did not affect the evoked NK1R internalization) indicating that spinal opioid release is mediated supraspinally. In the absence of peptidase inhibitors, formalin and hind paw clamp induced a small amount of MOR internalization, which was significantly higher than in controls. To study spinal opioid release during chronic inflammation, we injected complete Freund's adjuvant (CFA) in the hind paw and peptidase inhibitors intrathecally. Two days later, no MOR or NK1R internalization was detected. Furthermore, CFA inflammation decreased MOR internalization induced by clamping the inflamed hind paw. These results show that acute inflammation, but not chronic inflammation, induces segmental opioid release in the spinal cord that involves supraspinal signals.

  6. Opioid receptor subtypes mediating the noise-induced decreases in high-affinity choline uptake in the rat brain.

    Science.gov (United States)

    Lai, H; Carino, M A

    1992-07-01

    Acute (20 min) exposure to 100-dB white noise elicits a naltrexone-sensitive decrease in sodium-dependent high-affinity choline uptake in the frontal cortex and hippocampus of the rat. In the present study, the subtypes of opioid receptors involved were investigated by pretreating rats with microinjection of specific opioid-receptor antagonists into the lateral cerebroventricle before noise exposure. We found that the noise-induced decrease in high-affinity choline uptake in the hippocampus was blocked by pretreatment with either mu-, delta-, or kappa-opioid-receptor antagonists, whereas the effect of noise on frontal cortical high-affinity choline uptake was blocked by a mu- and delta- but not by a kappa-antagonist. These data further confirm the role of endogenous opioids in mediating the effects of noise on central cholinergic activity and indicate that different neural mechanisms are involved in the effects of noise on the frontal cortical and hippocampal cholinergic systems.

  7. The effect of benfotiamine on mu-opioid receptor mediated antinociception in experimental diabetes.

    Science.gov (United States)

    Nacitarhan, C; Minareci, E; Sadan, G

    2014-03-01

    Diabetic neuropathy is a prevalent, disabling disorder. Currently, the only treatments available to patients with diabetic neuropathy are glucose control and pain management. B vitamin present neuroprotective effects, which are suggested to be related to their analgesic action in various models of neuropathic pain. According to our literature knowledge there is no report about antinociceptive effects of thiamine as benfotiamine and opioids together in diabetic mice. The purpose of this study was to determine the effects of benfotiamine on the antinociception produced by mu-opioid receptor agonist fentanyl in diabetic mice. The effects of benfotiamine on antinociception produced by fentanyl in diabetic mice were studied in 4 groups. Antinociceptive effect was determined with tail flick, hot plate and formalin test. Our results showed that, mu-opioid agonist fentanyl in benfotiamine applied diabetic group caused more potent antinociceptive effect than in diabetic group without benfotiamine treatment. In brief benfotiamine supplement in diet did not bring out antinociceptive effect itself, but during development of STZ diabetes, benfotiamine replacement increased the antinociceptive effect of fentanyl in mice tail-flick test. This effect is probably due to the replacement of benfotiamine efficiency occurring in diabetes mellitus. Finally, we suppose that oral benfotiamine replacement therapy may be useful to ameliorate analgesic effect of mu-opioid agonists on neuropathic pain in diabetic case. © J. A. Barth Verlag in Georg Thieme Verlag KG Stuttgart · New York.

  8. Desensitization and Tolerance of Mu Opioid Receptors on Pontine Kölliker-Fuse Neurons.

    Science.gov (United States)

    Levitt, Erica S; Williams, John T

    2018-01-01

    Acute desensitization of mu opioid receptors is thought to be an initial step in the development of tolerance to opioids. Given the resistance of the respiratory system to develop tolerance, desensitization of neurons in the Kölliker-Fuse (KF), a key area in the respiratory circuit, was examined. The activation of G protein-coupled inwardly rectifying potassium current was measured using whole-cell voltage-clamp recordings from KF and locus coeruleus (LC) neurons contained in acute rat brain slices. A saturating concentration of the opioid agonist [Met 5 ]-enkephalin (ME) caused significantly less desensitization in KF neurons compared with LC neurons. In contrast to LC, desensitization in KF neurons was not enhanced by activation of protein kinase C or in slices from morphine-treated rats. Cellular tolerance to ME and morphine was also lacking in KF neurons from morphine-treated rats. The lack of cellular tolerance in KF neurons correlates with the relative lack of tolerance to the respiratory depressant effect of opioids. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

  9. Kappa opioid receptors in rat spinal cord vary across the estrous cycle.

    Science.gov (United States)

    Chang, P C; Aicher, S A; Drake, C T

    2000-04-07

    Kappa opioid receptors (KORs) were immunocytochemically localized in the lumbosacral spinal cord of female rats in different stages of the estrous cycle to examine the influence of hormonal status on receptor density. KOR labeling was primarily in fine processes and a few neuronal cell bodies in the superficial dorsal horn and the dorsolateral funiculus. Quantitative light microscopic densitometry of the superficial dorsal horn revealed that rats in diestrus had significantly lower KOR densities than those in proestrus or estrus. This suggests that female reproductive hormones regulate spinal KOR levels, which may contribute to variations in analgesic effectiveness of KOR agonists across the estrous cycle.

  10. Population pharmacokinetics of nalmefene in healthy subjects and its relation to μ-opioid receptor occupancy.

    Science.gov (United States)

    Kyhl, Lars-Erik Broksoe; Li, Shen; Faerch, Kirstine Ullitz; Soegaard, Birgitte; Larsen, Frank; Areberg, Johan

    2016-02-01

    The aims of this study were to develop a population pharmacokinetic (PK) model to describe the PK of nalmefene in healthy subjects and to relate the exposure of nalmefene to the μ-opioid receptor occupancy by simulations in the target population. Data from nine phase I studies (243 subjects) with extensive blood sampling were pooled and used for the population PK model building. Data from four other phase I studies (85 subjects) were pooled and used as an external validation dataset. Eight subjects from an imaging study contributed occupancy data and the pharmacokinetic/pharmacodynamic (PK/PD) relationship was modelled. Combining the population PK model and the PK/PD relationship enabled simulations to predict μ-opioid occupancy. A two compartment model with first order absorption best described the nalmefene PK data. The typical subject in the population was estimated to have a systemic clearance of 60.4 l h(-1) and a central volume of distribution of 266 l. Absolute oral bioavailability was estimated to 41% without food intake and with food about 53%. Simulation of the μ-opioid receptor occupancy shows that the 95% confidence bound is within or above 60-90% occupancy for up to 22-24 h after a single dose of 20 mg nalmefene. A robust population PK model for nalmefene was developed. Based on the concentration-occupancy model the μ-opioid receptor occupancy after a single 20 mg dose of nalmefene is predicted to be above the target therapeutic occupancy for about 24 h in about 95% of the target population. © 2015 The British Pharmacological Society.

  11. Deletion of the δ opioid receptor gene impairs place conditioning but preserves morphine reinforcement.

    Science.gov (United States)

    Le Merrer, Julie; Plaza-Zabala, Ainhoa; Del Boca, Carolina; Matifas, Audrey; Maldonado, Rafael; Kieffer, Brigitte L

    2011-04-01

    Converging experimental data indicate that δ opioid receptors contribute to mediate drug reinforcement processes. Whether their contribution reflects a role in the modulation of drug reward or an implication in conditioned learning, however, has not been explored. In the present study, we investigated the impact of δ receptor gene knockout on reinforced conditioned learning under several experimental paradigms. We assessed the ability of δ receptor knockout mice to form drug-context associations with either morphine (appetitive)- or lithium (aversive)-induced Pavlovian place conditioning. We also examined the efficiency of morphine to serve as a positive reinforcer in these mice and their motivation to gain drug injections, with operant intravenous self-administration under fixed and progressive ratio schedules and at two different doses. Mutant mice showed impaired place conditioning in both appetitive and aversive conditions, indicating disrupted context-drug association. In contrast, mutant animals displayed intact acquisition of morphine self-administration and reached breaking-points comparable to control subjects. Thus, reinforcing effects of morphine and motivation to obtain the drug were maintained. Collectively, the data suggest that δ receptor activity is not involved in morphine reinforcement but facilitates place conditioning. This study reveals a novel aspect of δ opioid receptor function in addiction-related behaviors. Copyright © 2011 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

  12. /sup 3/H)-(H-D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2) ((/sup 3/H)CTOP), a potent and highly selective peptide for mu opioid receptors in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Hawkins, K.N.; Knapp, R.J.; Lui, G.K.; Gulya, K.; Kazmierski, W.; Wan, Y.P.; Pelton, J.T.; Hruby, V.J.; Yamamura, H.I.

    1989-01-01

    The cyclic, conformationally restricted octapeptide (3H)-(H-D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2) ((3H)CTOP) was synthesized and its binding to mu opioid receptors was characterized in rat brain membrane preparations. Association rates (k+1) of 1.25 x 10(8) M-1 min-1 and 2.49 x 10(8) M-1 min-1 at 25 and 37 degrees C, respectively, were obtained, whereas dissociation rates (k-1) at the same temperatures were 1.93 x 10(-2) min-1 and 1.03 x 10(-1) min-1 at 25 and 37 degrees C, respectively. Saturation isotherms of (3H)CTOP binding to rat brain membranes gave apparent Kd values of 0.16 and 0.41 nM at 25 and 37 degrees C, respectively. Maximal number of binding sites in rat brain membranes were found to be 94 and 81 fmol/mg of protein at 25 and 37 degrees C, respectively. (3H)CTOP binding over a concentration range of 0.1 to 10 nM was best fit by a one site model consistent with binding to a single site. The general effect of different metal ions and guanyl-5'-yl-imidodiphosphate on (3H)CTOP binding was to reduce its affinity. High concentrations (100 mM) of sodium also produced a reduction of the apparent mu receptor density. Utilizing the delta opioid receptor specific peptide (3H)-(D-Pen2,D-Pen5)enkephalin, CTOP appeared to be about 2000-fold more specific for mu vs. delta opioid receptor than naloxone. Specific (3H)CTOP binding was inhibited by a large number of opioid or opiate ligands.

  13. The Mu opioid receptor promotes opioid and growth factor-induced proliferation, migration and Epithelial Mesenchymal Transition (EMT in human lung cancer.

    Directory of Open Access Journals (Sweden)

    Frances E Lennon

    Full Text Available Recent epidemiologic studies implying differences in cancer recurrence based on anesthetic regimens raise the possibility that the mu opioid receptor (MOR can influence cancer progression. Based on our previous observations that overexpression of MOR in human non-small cell lung cancer (NSCLC cells increased tumor growth and metastasis, this study examined whether MOR regulates growth factor receptor signaling and epithelial mesenchymal transition (EMT in human NSCLC cells. We utilized specific siRNA, shRNA, chemical inhibitors and overexpression vectors in human H358 NSCLC cells that were either untreated or treated with various concentrations of DAMGO, morphine, fentanyl, EGF or IGF. Cell function assays, immunoblot and immunoprecipitation assays were then performed. Our results indicate MOR regulates opioid and growth factor-induced EGF receptor signaling (Src, Gab-1, PI3K, Akt and STAT3 activation which is crucial for consequent human NSCLC cell proliferation and migration. In addition, human NSCLC cells treated with opioids, growth factors or MOR overexpression exhibited an increase in snail, slug and vimentin and decrease ZO-1 and claudin-1 protein levels, results consistent with an EMT phenotype. Further, these effects were reversed with silencing (shRNA or chemical inhibition of MOR, Src, Gab-1, PI3K, Akt and STAT3 (p<0.05. Our data suggest a possible direct effect of MOR on opioid and growth factor-signaling and consequent proliferation, migration and EMT transition during lung cancer progression. Such an effect provides a plausible explanation for the epidemiologic findings.

  14. Opioid Receptors Blockade Modulates Apoptosis in a Rat Model of ...

    African Journals Online (AJOL)

    inflammation characterized by replacement of liver tissue by ... Materials and Methods: Cirrhosis was induced in rats by bile duct ligation .... treated with deoxyribonucleic acid labeling solution containing ... binding was inhibited using non-immune serum. .... studies considering the increased level of tumor necrosis factor-α.

  15. Delta-opioid receptor analgesia is independent of microglial activation in a rat model of neuropathic pain.

    Directory of Open Access Journals (Sweden)

    Joanna Mika

    Full Text Available The analgesic effect of delta-opioid receptor (DOR ligands in neuropathic pain is not diminished in contrast to other opioid receptor ligands, which lose their effectiveness as analgesics. In this study, we examine whether this effect is related to nerve injury-induced microglial activation. We therefore investigated the influence of minocycline-induced inhibition of microglial activation on the analgesic effects of opioid receptor agonists: morphine, DAMGO, U50,488H, DPDPE, Deltorphin II and SNC80 after chronic constriction injury (CCI to the sciatic nerve in rats. Pre-emptive and repeated administration of minocycline (30 mg/kg, i.p. over 7 days significantly reduced allodynia and hyperalgesia as measured on day 7 after CCI. The antiallodynic and antihyperalgesic effects of intrathecally (i.t. administered morphine (10-20 µg, DAMGO (1-2 µg and U50,488H (25-50 µg were significantly potentiated in rats after minocycline, but no such changes were observed after DPDPE (10-20 µg, deltorphin II (1.5-15 µg and SNC80 (10-20 µg administration. Additionally, nerve injury-induced down-regulation of all types of opioid receptors in the spinal cord and dorsal root ganglia was not influenced by minocycline, which indicates that the effects of opioid ligands are dependent on other changes, presumably neuroimmune interactions. Our study of rat primary microglial cell culture using qRT-PCR, Western blotting and immunocytochemistry confirmed the presence of mu-opioid receptors (MOR and kappa-opioid receptors (KOR, further we provide the first evidence for the lack of DOR on microglial cells. In summary, DOR analgesia is different from analgesia induced by MOR and KOR receptors because it does not dependent on injury-induced microglial activation. DOR agonists appear to be the best candidates for new drugs to treat neuropathic pain.

  16. The μ-opioid receptor gene and smoking initiation and nicotine dependence

    Directory of Open Access Journals (Sweden)

    Kendler Kenneth S

    2006-08-01

    Full Text Available Abstract The gene encoding the mu-opioid receptor (OPRM1 is reported to be associated with a range of substance dependence. Experiments in knockout mice indicate that the mu-opioid receptor may mediate reinforcing effects of nicotine. In humans, opioid antagonist naltrexone may reduce the reinforcing effects of tobacco smoking. Additionally, the OPRM1 gene is located in a region showing linkage to nicotine dependence. The OPRM1 is thus a plausible candidate gene for smoking behavior. To investigate whether OPRM1 contributes to the susceptibility of smoking initiation and nicotine dependence, we genotyped 11 SNPs in the gene for 688 Caucasian subjects of lifetime smokers and nonsmokers. Three SNPs showed nominal significance for smoking initiation and one reached significance for nicotine dependence. The global test for three-marker (rs9479757-rs2075572-rs10485057 haplotypes was significant for smoking initiation (p = 0.0022. The same three-marker haplotype test was marginal (p = 0.0514 for nicotine dependence. These results suggest that OPRM1 may be involved in smoking initiation and nicotine dependence.

  17. TRV0109101, a G Protein-Biased Agonist of the µ-Opioid Receptor, Does Not Promote Opioid-Induced Mechanical Allodynia following Chronic Administration.

    Science.gov (United States)

    Koblish, Michael; Carr, Richard; Siuda, Edward R; Rominger, David H; Gowen-MacDonald, William; Cowan, Conrad L; Crombie, Aimee L; Violin, Jonathan D; Lark, Michael W

    2017-08-01

    Prescription opioids are a mainstay in the treatment of acute moderate to severe pain. However, chronic use leads to a host of adverse consequences including tolerance and opioid-induced hyperalgesia (OIH), leading to more complex treatment regimens and diminished patient compliance. Patients with OIH paradoxically experience exaggerated nociceptive responses instead of pain reduction after chronic opioid usage. The development of OIH and tolerance tend to occur simultaneously and, thus, present a challenge when studying the molecular mechanisms driving each phenomenon. We tested the hypothesis that a G protein-biased µ -opioid peptide receptor (MOPR) agonist would not induce symptoms of OIH, such as mechanical allodynia, following chronic administration. We observed that the development of opioid-induced mechanical allodynia (OIMA), a model of OIH, was absent in β -arrestin1 -/- and β -arrestin2 -/- mice in response to chronic administration of conventional opioids such as morphine, oxycodone and fentanyl, whereas tolerance developed independent of OIMA. In agreement with the β -arrestin knockout mouse studies, chronic administration of TRV0109101, a G protein-biased MOPR ligand and structural analog of oliceridine, did not promote the development of OIMA but did result in drug tolerance. Interestingly, following induction of OIMA by morphine or fentanyl, TRV0109101 was able to rapidly reverse allodynia. These observations establish a role for β -arrestins in the development of OIH, independent of tolerance, and suggest that the use of G protein-biased MOPR ligands, such as oliceridine and TRV0109101, may be an effective therapeutic avenue for managing chronic pain with reduced propensity for opioid-induced hyperalgesia. Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.

  18. Structural Analysis of Botulinum Neurotoxin Type G Receptor Binding

    Energy Technology Data Exchange (ETDEWEB)

    Schmitt, John; Karalewitz, Andrew; Benefield, Desire A.; Mushrush, Darren J.; Pruitt, Rory N.; Spiller, Benjamin W.; Barbieri, Joseph T.; Lacy, D. Borden (Vanderbilt); (MCW)

    2010-10-19

    Botulinum neurotoxin (BoNT) binds peripheral neurons at the neuromuscular junction through a dual-receptor mechanism that includes interactions with ganglioside and protein receptors. The receptor identities vary depending on BoNT serotype (A-G). BoNT/B and BoNT/G bind the luminal domains of synaptotagmin I and II, homologous synaptic vesicle proteins. We observe conditions under which BoNT/B binds both Syt isoforms, but BoNT/G binds only SytI. Both serotypes bind ganglioside G{sub T1b}. The BoNT/G receptor-binding domain crystal structure provides a context for examining these binding interactions and a platform for understanding the physiological relevance of different Syt receptor isoforms in vivo.

  19. Enantioselective kappa opioid binding sites on the macrophage cell line, P388d sub 1

    Energy Technology Data Exchange (ETDEWEB)

    Carr, D.J.J.; Blalock, J.E. (Univ. of Alabama, Birmingham (USA)); DeCosta, B.R.; Jacobson, A.E.; Rice, K.C. (NIDDK, NIH, Bethesda, MD (USA))

    1991-01-01

    A kappa opioid binding site has been characterized on the macrophage cell line, P388d{sub 1}, using the kappa selective affinity ligand, ({sup 3H}(1S,2S)-(-)-trans-2-isothiocyanato-N-methyl-N-(2-(1-phrrolidinyl) cyclohexyl) benzeneacetamide ((-)BD166). The kappa site has a relative molecular mass (Mr) of 38,000 under nonreducing conditions and 42,000 under reducing conditions. Moreover, it exhibits enantioselectivity in that 1S,2S-(-)-trans-3,4-dichloro-N-methyl-N-(2-(1-pyrrolidinyl)cyclohexyl) benzeneacetamide ((-)-U-50,488) blocks ({sup 3}H)95{alpha},7{alpha},8{beta})-(-)-N-methyl-N-(7-(1- pyrrolidinyl)-1-oxaspiro-(4,5)-dec-8-yl)benzeneacetamide (U-69,593) binding to P388d{sub 1} cells with an IC{sub 50} = 7.0 nM whereas 1R,2R-(+)-trans-3,4-dichloro-N-methyl-N-(2-(1-pyrrolidinyl)cyclohexyl) benzeneacetamide ((+)U-50,488) blocks ({sup 3}H)U-69,593 binding to P388d{sub 1} cells with an IC{sub 50} = 700 nM.

  20. Effects of the kappa opioid receptor antagonist MR-2266-BS on the acquisition of ethanol preference

    Energy Technology Data Exchange (ETDEWEB)

    Sandi, C.; Borrell, J.; Guaza, C. (Cajal Institute, Madrid (Spain))

    1990-01-01

    Using a paradigm by which rats forced to drink a weak ethanol solution develop ethanol preference in consecutive retention testing days, the effects of the administration of the kappa opioid antagonist MR-2266-BS, prior to or after the forced ethanol session, were studied. Pre-conditioning subcutaneous (s.c.) administration of 1 mg/kg of MR-2266-BS induced a decrease in subsequent ethanol consumption without significantly modifying the acquisition of ethanol preference. Post-conditioning administration of MR-2266-BS induced both a dose-dependent reduction in ethanol consumption and in preference throughout the three following days. The results of the present study provide further support of the involvement of kappa-type opioids on drinking behavior, and suggest that kappa receptors may be involved in the consumption and development of preference to ethanol.

  1. Gonadal cell surface receptor for plasma retinol-binding protein

    International Nuclear Information System (INIS)

    Krishna Bhat, M.; Cama, H.R.

    1979-01-01

    A specific membrane receptor for plasma retinol-binding protein has been demonstrated in testicular cells. Prealbumin-2 did not show any specific binding to the membrane. The affinity of retinol-binding protein for receptor drastically decreases upon delivery of retinol and the retinol-binding protein does not enter the cell. The mechanism of delivery of retinol to the target cell by plasma retinol-binding protein has been investigated. The process involves two steps; direct binding of retinol-binding protein to the receptor and uptake of retinol by the target cell with a concomitant drastic reduction in the affinity of the retinol-binding protein to the receptor. Probably the second step of the process needs a cytosolic factor, possibly the cellular retinol-binding protein or an enzyme. The binding of retinol-binding protein to the receptor is saturable and reversible. The interaction shows a Ksub(d) value of 2.1x10 -10 . The specific binding of a retinol-binding protein with great affinity has been employed in the development of a method for radioassay of the receptor. The receptor level of the gonadal cell has been found to vary with the stage of differentiation. The receptor concentrations in 11-week-old birds and adult birds are comparable. Testosterone treatment of 11-week-old birds produced a substantial increase in the receptor concentration over control, while the protein content increased marginally, indicating that, probably, synthesis of the receptor is specifcally induced by testosterone during spermatogenesis, and the concentration of receptor is relatively higher before the formation of the acrosome. (Auth.)

  2. Molecular Docking, Molecular Dynamics, and Structure-Activity Relationship Explorations of 14-Oxygenated N-Methylmorphinan-6-ones as Potent μ-Opioid Receptor Agonists.

    Science.gov (United States)

    Noha, Stefan M; Schmidhammer, Helmut; Spetea, Mariana

    2017-06-21

    Among opioids, morphinans are of major importance as the most effective analgesic drugs acting primarily via μ-opioid receptor (μ-OR) activation. Our long-standing efforts in the field of opioid analgesics from the class of morphinans led to N-methylmorphinan-6-ones differently substituted at positions 5 and 14 as μ-OR agonists inducing potent analgesia and fewer undesirable effects. Herein we present the first thorough molecular modeling study and structure-activity relationship (SAR) explorations aided by docking and molecular dynamics (MD) simulations of 14-oxygenated N-methylmorphinan-6-ones to gain insights into their mode of binding to the μ-OR and interaction mechanisms. The structure of activated μ-OR provides an essential model for how ligand/μ-OR binding is encoded within small chemical differences in otherwise structurally similar morphinans. We reveal important molecular interactions that these μ-agonists share and distinguish them. The molecular docking outcomes indicate the crucial role of the relative orientation of the ligand in the μ-OR binding site, influencing the propensity of critical non-covalent interactions that are required to facilitate ligand/μ-OR interactions and receptor activation. The MD simulations point out minor differences in the tendency to form hydrogen bonds by the 4,5α-epoxy group, along with the tendency to affect the 3-7 lock switch. The emerged SARs reveal the subtle interplay between the substituents at positions 5 and 14 in the morphinan scaffold by enabling the identification of key structural elements that determine the distinct pharmacological profiles. This study provides a significant structural basis for understanding ligand binding and μ-OR activation by the 14-oxygenated N-methylmorphinan-6-ones, which should be useful for guiding drug design.

  3. Cell-Autonomous Regulation of Mu-Opioid Receptor Recycling by Substance P

    Science.gov (United States)

    Bowman, Shanna L.; Soohoo, Amanda L.; Shiwarski, Daniel J.; Schulz, Stefan; Pradhan, Amynah A.; Puthenveedu, Manojkumar A.

    2015-01-01

    SUMMARY How neurons coordinate and reprogram multiple neurotransmitter signals is an area of broad interest. Here, we show that substance P (SP), a neuropep-tide associated with inflammatory pain, reprograms opioid receptor recycling and signaling. SP, through activation of the neurokinin 1 (NK1R) receptor, increases the post-endocytic recycling of the muopioid receptor (MOR) in trigeminal ganglion (TG) neurons in an agonist-selective manner. SP-mediated protein kinase C (PKC) activation is both required and sufficient for increasing recycling of exogenous and endogenous MOR in TG neurons. The target of this cross-regulation is MOR itself, given that mutation of either of two PKC phosphorylation sites on MOR abolishes the SP-induced increase in recycling and resensitization. Furthermore, SP enhances the resensitization of fentanyl-induced, but not morphine-induced, antinociception in mice. Our results define a physiological pathway that cross-regulates opioid receptor recycling via direct modification of MOR and suggest a mode of homeo-static interaction between the pain and analgesic systems. PMID:25801029

  4. Reduced emotional and corticosterone responses to stress in μ-opioid receptor knockout mice

    Science.gov (United States)

    Ide, Soichiro; Sora, Ichiro; Ikeda, Kazutaka; Minami, Masabumi; Uhl, George R.; Ishihara, Kumatoshi

    2014-01-01

    The detailed mechanisms of emotional modulation in the nervous system by opioids remain to be elucidated, although the opioid system is well known to play important roles in the mechanisms of analgesia and drug dependence. In the present study, we conducted behavioral tests of anxiety and depression and measured corticosterone concentrations in both male and female μ-opioid receptor knockout (MOP-KO) mice to reveal the involvement of μ-opioid receptors in stress-induced emotional responses. MOP-KO mice entered more and spent more time in the open arms of the elevated plus maze compared with wild-type mice. MOP-KO mice also displayed significantly decreased immobility in a 15 min tail-suspension test compared with wild-type mice. Similarly, MOP-KO mice exhibited significantly decreased immobility on days 2, 3, and 4 in a 6 min forced swim test conducted for 5 consecutive days. The increase in plasma corticosterone concentration induced by tail-suspension, repeated forced swim, or restraint stress was reduced in MOP-KO mice compared with wild-type mice. Corticosterone levels were not different between wild-type and MOP-KO mice before stress exposure. In contrast, although female mice tended to exhibit fewer anxiety-like responses in the tail-suspension test in both genotypes, no significant gender differences were observed in stress-induced emotional responses. These results suggest that MOPs play an important facilitatory role in emotional responses to stress, including anxiety- and depression-like behavior and corticosterone levels. PMID:19596019

  5. alpha-Adrenoceptor and opioid receptor modulation of clonidine-induced antinociception.

    Science.gov (United States)

    Sierralta, F; Naquira, D; Pinardi, G; Miranda, H F

    1996-10-01

    1. The antinociceptive action of clonidine (Clon) and the interactions with alpha 1, alpha 2 adrenoceptor and opioid receptor antagonists was evaluated in mice by use of chemical algesiometric test (acetic acid writhing test). 2. Clon produced a dose-dependent antinociceptive action and the ED50 for intracerebroventricular (i.c.v.) was lower than for intraperitoneal (i.p.) administration (1 ng kg-1 vs 300 ng kg-1). The parallelism of the dose-response curves indicates activation of a common receptor subtype. 3. Systemic administration of prazosin and terazosin displayed antinociceptive activity. Pretreatment with prazosin produced a dual action: i.c.v. Clon effect did not change, and i.p. Clon effect was enhanced. Yohimbine i.c.v. or i.p. did not induce antinonciception, but antagonized Clon-induced activity. These results suggest that alpha 1- and alpha 2-adrenoceptors, either located at the pre- and/or post-synaptic level, are involved in the control of spinal antinociception. 4. Naloxone (NX) and naltrexone (NTX) induced antinociceptive effects at low doses (microgram kg-1 range) and a lower antinociceptive effect at higher doses (mg kg-1 range). Low doses of NX or NTX antagonized Clon antinociception, possibly in relation to a preferential mu opioid receptor antagonism. In contrast, high doses of NX or NTX increased the antinociceptive activity of Clon, which could be due to an enhanced inhibition of the release of substance P. 5. The results obtained in the present work suggest the involvement of alpha 1-, alpha 2-adrenoceptor and opioid receptors in the modulation of the antinociceptive activity of clonidine, which seems to be exerted either at spinal and/or supraspinal level.

  6. Concomitant duplications of opioid peptide and receptor genes before the origin of jawed vertebrates.

    Directory of Open Access Journals (Sweden)

    Görel Sundström

    Full Text Available BACKGROUND: The opioid system is involved in reward and pain mechanisms and consists in mammals of four receptors and several peptides. The peptides are derived from four prepropeptide genes, PENK, PDYN, PNOC and POMC, encoding enkephalins, dynorphins, orphanin/nociceptin and beta-endorphin, respectively. Previously we have described how two rounds of genome doubling (2R before the origin of jawed vertebrates formed the receptor family. METHODOLOGY/PRINCIPAL FINDINGS: Opioid peptide gene family members were investigated using a combination of sequence-based phylogeny and chromosomal locations of the peptide genes in various vertebrates. Several adjacent gene families were investigated similarly. The results show that the ancestral peptide gene gave rise to two additional copies in the genome doublings. The fourth member was generated by a local gene duplication, as the genes encoding POMC and PNOC are located on the same chromosome in the chicken genome and all three teleost genomes that we have studied. A translocation has disrupted this synteny in mammals. The PDYN gene seems to have been lost in chicken, but not in zebra finch. Duplicates of some peptide genes have arisen in the teleost fishes. Within the prepropeptide precursors, peptides have been lost or gained in different lineages. CONCLUSIONS/SIGNIFICANCE: The ancestral peptide and receptor genes were located on the same chromosome and were thus duplicated concomitantly. However, subsequently genetic linkage has been lost. In conclusion, the system of opioid peptides and receptors was largely formed by the genome doublings that took place early in vertebrate evolution.

  7. Part I. Naltrexone-derived conjugate addition ligands for opioid receptors. Part II. Chemical and enantioselective aspects of the metabolism of verapamil

    International Nuclear Information System (INIS)

    Olsen, L.D.

    1987-01-01

    Selective chemoaffinity ligands to aid in identification and purification of opioid receptor subtypes were prepared from 6α- and 6β-naltrexol, obtained stereoselectively from the μ-receptor antagonist naltrexone. The targets were the 6α- and 6β-methacrylate ethers and 6α- and 6β-methacrylate esters prepared from reaction of 6α- and 6β-naltrexol with methyl α-(bromomethyl)acrylate or methacryloyl chloride. Of three methacrylate derivatives, the 6α-ether was the most potent in an opioid receptor binding assay with [ 3 H]-naltrexone. In the presence of sodium ion, preincubation of the 6α-ether resulted in recovery of about 60% of original [ 3 H]-naltrexone binding suggesting some irreversible effects. The methacrylate esters precipitated withdrawal in morphine dependent monkeys. The enantiomers of verapamil, a calcium channel antagonist, have different pharmacological and pharmacokinetic properties. The oxidative metabolism of verapamil was studied in rat and human liver microsomes and in man after a single oral dose

  8. Part I. Naltrexone-derived conjugate addition ligands for opioid receptors. Part II. Chemical and enantioselective aspects of the metabolism of verapamil

    Energy Technology Data Exchange (ETDEWEB)

    Olsen, L.D.

    1987-01-01

    Selective chemoaffinity ligands to aid in identification and purification of opioid receptor subtypes were prepared from 6..cap alpha..- and 6..beta..-naltrexol, obtained stereoselectively from the ..mu..-receptor antagonist naltrexone. The targets were the 6..cap alpha..- and 6..beta..-methacrylate ethers and 6..cap alpha..- and 6..beta..-methacrylate esters prepared from reaction of 6..cap alpha..- and 6..beta..-naltrexol with methyl ..cap alpha..-(bromomethyl)acrylate or methacryloyl chloride. Of three methacrylate derivatives, the 6..cap alpha..-ether was the most potent in an opioid receptor binding assay with (/sup 3/H)-naltrexone. In the presence of sodium ion, preincubation of the 6..cap alpha..-ether resulted in recovery of about 60% of original (/sup 3/H)-naltrexone binding suggesting some irreversible effects. The methacrylate esters precipitated withdrawal in morphine dependent monkeys. The enantiomers of verapamil, a calcium channel antagonist, have different pharmacological and pharmacokinetic properties. The oxidative metabolism of verapamil was studied in rat and human liver microsomes and in man after a single oral dose.

  9. Analgesic tone conferred by constitutively active mu opioid receptors in mice lacking β-arrestin 2

    Directory of Open Access Journals (Sweden)

    Hales Tim G

    2011-04-01

    Full Text Available Abstract Hedonic reward, dependence and addiction are unwanted effects of opioid analgesics, linked to the phasic cycle of μ opioid receptor activation, tolerance and withdrawal. In vitro studies of recombinant G protein coupled receptors (GPCRs over expressed in cell lines reveal an alternative tonic signaling mechanism that is independent of agonist. Such studies demonstrate that constitutive GPCR signaling can be inhibited by inverse agonists but not by neutral antagonists. However, ligand-independent activity has been difficult to examine in vivo, at the systems level, due to relatively low levels of constitutive activity of most GPCRs including μ receptors, often necessitating mutagenesis or pharmacological manipulation to enhance basal signaling. We previously demonstrated that the absence of β-arrestin 2 (β-arr2 augments the constitutive coupling of μ receptors to voltage-activated Ca2+ channels in primary afferent dorsal root ganglion neurons from β-arr2-/- mice. We used this in vitro approach to characterize neutral competitive antagonists and inverse agonists of the constitutively active wild type μ receptors in neurons. We administered these agents to β-arr2-/- mice to explore the role of constitutive μ receptor activity in nociception and hedonic tone. This study demonstrates that the induction of constitutive μ receptor activity in vivo in β-arr2-/- mice prolongs tail withdrawal from noxious heat, a phenomenon that was reversed by inverse agonists, but not by antagonists that lack negative efficacy. By contrast, the aversive effects of inverse agonists were similar in β-arr2-/- and β-arr2+/+ mice, suggesting that hedonic tone was unaffected.

  10. Preparation and biodistribution in mice of ( sup 11 C)carfentanil; A radiopharmaceutical for studying brain. mu. -opioid receptors by positron emission tomography

    Energy Technology Data Exchange (ETDEWEB)

    Saji, Hideo; Tsutsumi, Daisuke; Iida, Yasuhiko; Yokoyama, Akira (Kyoto Univ. (Japan). Faculty of Pharmaceutical Science); Magata, Yasuhiro; Konishi, Junji

    1992-02-01

    A potent {mu}-opioid agonist, ({sup 11}C)carfentanil, was prepared by the methylation of carfentanil carboxylic acid with ({sup 11}C)methyl iodide in order to study brain {mu}-opioid receptors by positron emission tomography. Synthesis (including purification) was completed within 25 min and the radiochemical yield was approximately 40%. The radiochemical purity of the product was more than 99% and its specific activity was 3.7-7.4 GBq/{mu}mol. Biodistribution studies performed in mice after intravenous injection showed a high brain uptake and rapid blood clearance, so a high brain/blood ratio of 1.5-1.8 was found from 5 to 30 min. Regional cerebral distribution studies in the mouse showed a significantly higher uptake of ({sup 11}C)carfentanil by the thalamus and striatum than by the cerebellum, with the radioactivity in the striatum disappearing more rapidly than that in the thalamus. Treatment with naloxone significantly reduced the uptake of ({sup 11}C)carfentanil by the thalamus and striatum. These results indicate that ({sup 11}C)carfentanil binds specifically to brain {mu}-opioid receptors. (author).

  11. β-Arrestin-2 knockout prevents development of cellular μ-opioid receptor tolerance but does not affect opioid-withdrawal-related adaptations in single PAG neurons.

    Science.gov (United States)

    Connor, M; Bagley, E E; Chieng, B C; Christie, M J

    2015-01-01

    Tolerance to the behavioural effects of morphine is blunted in β-arrestin-2 knockout mice, but opioid withdrawal is largely unaffected. The cellular mechanisms of tolerance have been studied in some neurons from β-arrestin-2 knockouts, but tolerance and withdrawal mechanisms have not been examined at the cellular level in periaqueductal grey (PAG) neurons, which are crucial for central tolerance and withdrawal phenomena. μ-Opioid receptor (MOPr) inhibition of voltage-gated calcium channel currents (ICa ) was examined by patch-clamp recordings from acutely dissociated PAG neurons from wild-type and β-arrestin-2 knockout mice treated chronically with morphine (CMT) or vehicle. Opioid withdrawal-induced activation of GABA transporter type 1 (GAT-1) currents was determined using perforated patch recordings from PAG neurons in brain slices. MOPr inhibition of ICa in PAG neurons was unaffected by β-arrestin-2 deletion. CMT impaired coupling of MOPrs to ICa in PAG neurons from wild-type mice, but this cellular tolerance was not observed in neurons from CMT β-arrestin-2 knockouts. However, β-arrestin-2 knockouts displayed similar opioid-withdrawal-induced activation of GAT-1 currents as wild-type PAG neurons. In β-arrestin-2 knockout mice, the central neurons involved in the anti-nociceptive actions of opioids also fail to develop cellular tolerance to opioids following chronic morphine. The results also provide the first cellular physiological evidence that opioid withdrawal is not disrupted by β-arrestin-2 deletion. However, the unaffected basal sensitivity to opioids in PAG neurons provides further evidence that changes in basal MOPr sensitivity cannot account for the enhanced acute nociceptive response to morphine reported in β-arrestin-2 knockouts. This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2. © 2014 The British

  12. Light microscopic autoradiographic localization of mu and delta opioid binding sites in the mouse central nervous system

    International Nuclear Information System (INIS)

    Moskowitz, A.S.; Goodman, R.R.

    1984-01-01

    Much work has been done on opioid systems in the rat CNS. Although the mouse is widely used in pharmacological studies of opioid action, little has been done to characterize opioid systems in this species. In the present study the distribution of mu and delta opioid binding sites in the mouse CNS was examined using a quantitative in vitro autoradiography procedure. Tritiated dihydromorphine was used to visualize mu sites and [3H-d-Ala2-d-Leu5]enkephalin with a low concentration of morphine was used to visualize delta sites. Mu and delta site localizations in the mouse are very similar to those previously described in the rat (Goodman, R.R., S.H. Snyder, M.J. Kuhar, and W.S. Young, 3d (1980) Proc. Natl. Acad. Sci. U.S.A. 77:6239-6243), with certain exceptions and additions. Mu and delta sites were observed in sensory processing areas, limbic system, extrapyramidal motor system, and cranial parasympathetic system. Differential distributions of mu and delta sites were noted in many areas. Mu sites were prominent in laminae I, IV, and VI of the neocortex, in patches in the striatum, and in the ventral pallidum, nucleus accumbens, medial and midline thalamic nuclei, medial habenular nucleus, interpeduncular nucleus, and laminae I and II of the spinal cord. In contrast, delta sites were prominent in all laminae of the neocortex, olfactory tubercle, diffusely throughout the striatum, and in the basal, lateral, and cortical nuclei of the amygdala. The determination of the differential distributions of opioid binding sites should prove useful in suggesting anatomical substrates for the actions of opiates and opioids

  13. Opioid transport by ATP-binding cassette transporters at the blood-brain barrier: implications for neuropsychopharmacology.

    Science.gov (United States)

    Tournier, Nicolas; Declèves, Xavier; Saubaméa, Bruno; Scherrmann, Jean-Michel; Cisternino, Salvatore

    2011-01-01

    Some of the ATP-binding cassette (ABC) transporters like P-glycoprotein (P-gp; ABCB1, MDR1), BCRP (ABCG2) and MRPs (ABCCs) that are present at the blood-brain barrier (BBB) influence the brain pharmacokinetics (PK) of their substrates by restricting their uptake or enhancing their clearance from the brain into the blood, which has consequences for their CNS pharmacodynamics (PD). Opioid drugs have been invaluable tools for understanding the PK-PD relationships of these ABC-transporters. The effects of morphine, methadone and loperamide on the CNS are modulated by P-gp. This review examines the ways in which other opioid drugs and some of their active metabolites interact with ABC transporters and suggests new mechanisms that may be involved in the variability of the response of the CNS to these drugs like carrier-mediated system belonging to the solute carrier (SLC) superfamily. Exposure to opioids may also alter the expression of ABC transporters. P-gp can be overproduced during morphine treatment, suggesting that the drug has a direct or, more likely, an indirect action. Variations in cerebral neurotransmitters during exposure to opioids and the release of cytokines during pain could be new endogenous stimuli affecting transporter synthesis. This review concludes with an analysis of the pharmacotherapeutic and clinical impacts of the interactions between ABC transporters and opioids.

  14. High Efficacy but Low Potency of δ-Opioid Receptor-G Protein Coupling in Brij-58-Treated, Low-Density Plasma Membrane Fragments.

    Science.gov (United States)

    Roubalova, Lenka; Vosahlikova, Miroslava; Brejchova, Jana; Sykora, Jan; Rudajev, Vladimir; Svoboda, Petr

    2015-01-01

    HEK293 cells stably expressing PTX-insensitive δ-opioid receptor-Gi1α (C351I) fusion protein were homogenized, treated with low concentrations of non-ionic detergent Brij-58 at 0°C and fractionated by flotation in sucrose density gradient. In optimum range of detergent concentrations (0.025-0.05% w/v), Brij-58-treated, low-density membranes exhibited 2-3-fold higher efficacy of DADLE-stimulated, high-affinity [32P]GTPase and [35S]GTPγS binding than membranes of the same density prepared in the absence of detergent. The potency of agonist DADLE response was significantly decreased. At high detergent concentrations (>0.1%), the functional coupling between δ-opioid receptors and G proteins was completely diminished. The same detergent effects were measured in plasma membranes isolated from PTX-treated cells. Therefore, the effect of Brij-58 on δ-opioid receptor-G protein coupling was not restricted to the covalently bound Gi1α within δ-opioid receptor-Gi1α fusion protein, but it was also valid for PTX-sensitive G proteins of Gi/Go family endogenously expressed in HEK293 cells. Characterization of the direct effect of Brij-58 on the hydrophobic interior of isolated plasma membranes by steady-state anisotropy of diphenylhexatriene (DPH) fluorescence indicated a marked increase of membrane fluidity. The time-resolved analysis of decay of DPH fluorescence by the "wobble in cone" model of DPH motion in the membrane indicated that the exposure to the increasing concentrations of Brij-58 led to a decreased order and higher motional freedom of the dye. Limited perturbation of plasma membrane integrity by low concentrations of non-ionic detergent Brij-58 results in alteration of δ-OR-G protein coupling. Maximum G protein-response to agonist stimulation (efficacy) is increased; affinity of response (potency) is decreased. The total degradation plasma membrane structure at high detergent concentrations results in diminution of functional coupling between δ-opioid

  15. Morphine withdrawal enhances constitutive μ-opioid receptor activity in the ventral tegmental area.

    Science.gov (United States)

    Meye, Frank J; van Zessen, Ruud; Smidt, Marten P; Adan, Roger A H; Ramakers, Geert M J

    2012-11-14

    μ-Opioid receptors (MORs) in the ventral tegmental area (VTA) are pivotally involved in addictive behavior. While MORs are typically activated by opioids, they can also become constitutively active in the absence of any agonist. In the current study, we present evidence that MOR constitutive activity is highly relevant in the mouse VTA, as it regulates GABAergic input to dopamine neurons. Specifically, suppression of MOR constitutive activity with the inverse agonist KC-2-009 enhanced GABAergic neurotransmission onto VTA dopamine neurons. This inverse agonistic effect was fully blocked by the specific MOR neutral antagonist CTOP, which had no effect on GABAergic transmission itself. We next show that withdrawal from chronic morphine further increases the magnitude of inverse agonistic effects at the MOR, suggesting enhanced MOR constitutive activity. We demonstrate that this increase can be an adaptive response to the detrimental elevation in cAMP levels known to occur during morphine withdrawal. These findings offer important insights in the physiological occurrence and function of MOR constitutive activity, and have important implications for therapeutic strategies aimed at normalizing MOR signaling during addiction and opioid overdose.

  16. Role of kappa-opioid receptors in the effects of salvinorin A and ketamine on attention in rats.

    Science.gov (United States)

    Nemeth, Christina L; Paine, Tracie A; Rittiner, Joseph E; Béguin, Cécile; Carroll, F Ivy; Roth, Bryan L; Cohen, Bruce M; Carlezon, William A

    2010-06-01

    Disruptions in perception and cognition are characteristic of psychiatric conditions such as schizophrenia. Studies of pharmacological agents that alter perception and cognition in humans might provide a better understanding of the brain substrates of these complex processes. One way to study these states in rodents is with tests that require attention and visual perception for correct performance. We examined the effects of two drugs that cause disruptions in perception and cognition in humans-the kappa-opioid receptor (KOR) agonist salvinorin A (salvA; 0.125-4.0 mg/kg) and the non-competitive NMDA receptor antagonist ketamine (0.63-20 mg/kg)-on behavior in rats using the 5-choice serial reaction time task (5CSRTT), a food-motivated test that quantifies attention. We also compared the binding profiles of salvA and ketamine at KORs and NMDA receptors. SalvA and ketamine produced the same pattern of disruptive effects in the 5CSRTT, characterized by increases in signs often associated with reduced motivation (omission errors) and deficits in processing (elevated latencies to respond correctly). Sessions in which rats were fed before testing suggest that reduced motivation produces a subtly different pattern of behavior. Pretreatment with the KOR antagonist JDTic (10 mg/kg) blocked all salvA effects and some ketamine effects. Binding and function studies revealed that ketamine is a full agonist at KORs, although not as potent or selective as salvA. SalvA and ketamine have previously under-appreciated similarities in their behavioral effects and pharmacological profiles. By implication, KORs might be involved in some of the cognitive abnormalities observed in psychiatric disorders such as schizophrenia.

  17. Synthesis of triated N1'-alkyl derivatives of the delta opioid receptor ligand naltrindole

    International Nuclear Information System (INIS)

    Lever, J.R.; Johnson, S.M.

    1997-01-01

    Tritiated N1'-methyl and N1'-ethyl analogues of naltrindole (NTI) have been synthesized for evaluation as radioligands for studies of delta opioid receptors. The two N1'-alkyl-5',7'-dibromoNTI precursors for radiolabeling were prepared by base-promoted alkylation of 2,4-dibromophenylhydrazine with either iodomethane or iodoethane followed by condensation with naltrexone using the Fischer indole synthesis. Catalytic debromotritiation followed by HPLC purification afforded [ 3 H]MeNTI (17.3 Ci/mmol) and [ 3 H]EtNTI (22.5 Ci/mmol) with high chemical and radiochemical purities (≥ 99.8%). (author)

  18. N-(3-( sup 18 F)fluoropropyl)-N-nordiprenorphine: Synthesis and characterization of a new agent for imaging opioid receptors with positron emission tomography

    Energy Technology Data Exchange (ETDEWEB)

    Chesis, P.L.; Hwang, D.R.; Welch, M.J. (Washington Univ. School of Medicine, St. Louis, MO (USA))

    1990-05-01

    A series of N-fluoroalkyl (1-5) and N-alkyl (6-8) analogues of the high-affinity opioid receptor antagonist diprenorphine (9) has been synthesized and evaluated with in vitro binding assays. Three of the N-fluoroalkyl compounds were prepared with the positron-emitting radionuclide {sup 18}F (1a, 2a, 5a), and their biodistribution was determined in rats. Compounds 2a and 5a were made by using a two-step labeling procedure, ({sup 18}F)fluoride displacement of an iodoalkyl triflate followed by N-alkylation, that required 2 h and proceeded in 4-6% overall radiochemical yield at the end of synthesis. The effective specific activity of compounds 2a and 5a, determined by competitive receptor binding assay, was 840-1820 Ci/mmol. Compound 1a was made by the same two-step procedure, with the bromoalkyl triflate, in 0.3-0.6% radiochemical yield at an effective specific activity of 106-264 Ci/mmol. Specificity of binding in vivo was measured as the percent injected dose/gram of striatal tissue divided by the percent injected dose/gram of cerebellar tissue. The best striatum to cerebellum ratio (3.32 +/- 0.74 at 30 min) was achieved with N-(3-({sup 18}F)-fluoropropyl)-N-nordiprenorphine (2a, ({sup 18}F)FPND). The high specific binding demonstrated by this compound indicates that it may be useful for in vivo imaging of opioid receptors with positron emission tomography.

  19. Cholecystokinin-8 suppressed /sup 3/H-etorphine binding to rat brain opiate receptors

    Energy Technology Data Exchange (ETDEWEB)

    Wang, X.J.; Fan, S.G.; Ren, M.F.; Han, J.S.

    1989-01-01

    Radioreceptor assay (RRA) was adopted to analyze the influence of CCK-8 on /sup 3/H-etorphine binding to opiate receptors in rat brain synaptosomal membranes (P2). In the competition experiment CCK-8 suppressed the binding of /sup 3/H-etorphine. This effect was completely reversed by proglumide at 1/mu/M. Rosenthal analysis for saturation revealed two populations of /sup 3/H-etorphine binding sites. CCK-8 inhibited /sup 3/H-etorphine binding to the high affinity sites by an increase in Kd and decrease in Bmax without significant changes in the Kd and Bmax of the low affinity sites. This effect of CCK-8 was also completely reversed by proglumide at 1/mu/M. Unsulfated CCK-8 produced only a slight increase in Kd of the high affinity sites without affecting Bmax. The results suggest that CCK-8 might be capable of suppressing the high affinity opioid binding sites via the activation of CCK receptor.

  20. Lateralized kappa opioid receptor signaling from the amygdala central nucleus promotes stress-induced functional pain.

    Science.gov (United States)

    Nation, Kelsey M; De Felice, Milena; Hernandez, Pablo I; Dodick, David W; Neugebauer, Volker; Navratilova, Edita; Porreca, Frank

    2018-05-01

    The response of diffuse noxious inhibitory controls (DNIC) is often decreased, or lost, in stress-related functional pain syndromes. Because the dynorphin/kappa opioid receptor (KOR) pathway is activated by stress, we determined its role in DNIC using a model of stress-induced functional pain. Male, Sprague-Dawley rats were primed for 7 days with systemic morphine resulting in opioid-induced hyperalgesia. Fourteen days after priming, when hyperalgesia was resolved, rats were exposed to environmental stress and DNIC was evaluated by measuring hind paw response threshold to noxious pressure (test stimulus) after capsaicin injection in the forepaw (conditioning stimulus). Morphine priming without stress did not alter DNIC. However, stress produced a loss of DNIC in morphine-primed rats in both hind paws that was abolished by systemic administration of the KOR antagonist, nor-binaltorphimine (nor-BNI). Microinjection of nor-BNI into the right, but not left, central nucleus of the amygdala (CeA) prevented the loss of DNIC in morphine-primed rats. Diffuse noxious inhibitory controls were not modulated by bilateral nor-BNI in the rostral ventromedial medulla. Stress increased dynorphin content in both the left and right CeA of primed rats, reaching significance only in the right CeA; no change was observed in the rostral ventromedial medulla or hypothalamus. Although morphine priming alone is not sufficient to influence DNIC, it establishes a state of latent sensitization that amplifies the consequences of stress. After priming, stress-induced dynorphin/KOR signaling from the right CeA inhibits DNIC in both hind paws, likely reflecting enhanced descending facilitation that masks descending inhibition. Kappa opioid receptor antagonists may provide a new therapeutic strategy for stress-related functional pain disorders.

  1. Activation of Peripheral κ-Opioid Receptors Normalizes Caffeine Effects Modified in Nicotine-Dependent Rats during Nicotine Withdrawal.

    Science.gov (United States)

    Sudakov, S K; Bogdanova, N G

    2016-10-01

    The study examined the effect of peripheral (intragastric) ICI-204,448, an agonist of gastric κ-opioid receptors, on the psychostimulating and anxiolytic effects of caffeine in nicotinedependent rats at the stage of nicotine withdrawal. In these rats, the effects of caffeine (10 mg/kg) were perverted. In nicotine-dependent rats, caffeine produced an anxiolytic effect accompanied by pronounced stimulation of motor activity, in contrast to anxiogenic effect induced by caffeine in intact rats without nicotine dependence. During nicotine withdrawal, nicotine-dependent rats demonstrated enhanced sensitivity to nicotine. Intragastric administration of κ-opioid receptor agonist ICI-204,448 normalized the effect of caffeine in nicotinedependent rats. We have previously demonstrated that activation of peripheral κ-opioid receptors inhibited central κ-opioid activity and eliminated manifestations of nicotine withdrawal syndrome in nicotine-dependent rats, e.g. metabolism activation, stimulation of motor activity, and enhancement of food consumption. In its turn, inhibition of central κ-opioid structures activates the brain adenosine system, which can attenuate the caffeine-induced effects in nicotine-dependent rats.

  2. Cutaneous nociceptors lack sensitisation, but reveal μ-opioid receptor-mediated reduction in excitability to mechanical stimulation in neuropathy

    Directory of Open Access Journals (Sweden)

    Schmidt Yvonne

    2012-11-01

    Full Text Available Abstract Background Peripheral nerve injuries often trigger a hypersensitivity to tactile stimulation. Behavioural studies demonstrated efficient and side effect-free analgesia mediated by opioid receptors on peripheral sensory neurons. However, mechanistic approaches addressing such opioid properties in painful neuropathies are lacking. Here we investigated whether opioids can directly inhibit primary afferent neuron transmission of mechanical stimuli in neuropathy. We analysed the mechanical thresholds, the firing rates and response latencies of sensory fibres to mechanical stimulation of their cutaneous receptive fields. Results Two weeks following a chronic constriction injury of the saphenous nerve, mice developed a profound mechanical hypersensitivity in the paw innervated by the damaged nerve. Using an in vitro skin-nerve preparation we found no changes in the mechanical thresholds and latencies of sensory fibres from injured nerves. The firing rates to mechanical stimulation were unchanged or reduced following injury. Importantly, μ-opioid receptor agonist [D-Ala2,N-Me-Phe4,Gly5]-ol-enkephalin (DAMGO significantly elevated the mechanical thresholds of nociceptive Aδ and C fibres. Furthermore, DAMGO substantially diminished the mechanically evoked discharges of C nociceptors in injured nerves. These effects were blocked by DAMGO washout and pre-treatment with the selective μ-opioid receptor antagonist Cys2-Tyr3-Orn5-Pen7-amide. DAMGO did not alter the responses of sensory fibres in uninjured nerves. Conclusions Our findings suggest that behaviourally manifested neuropathy-induced mechanosensitivity does not require a sensitised state of cutaneous nociceptors in damaged nerves. Yet, nerve injury renders nociceptors sensitive to opioids. Prevention of action potential generation or propagation in nociceptors might represent a cellular mechanism underlying peripheral opioid-mediated alleviation of mechanical hypersensitivity in neuropathy.

  3. Heterologous activation of protein kinase C stimulates phosphorylation of delta-opioid receptor at serine 344, resulting in beta-arrestin- and clathrin-mediated receptor internalization

    DEFF Research Database (Denmark)

    Xiang, B; Yu, G H; Guo, J

    2001-01-01

    The purpose of the current study is to investigate the effect of opioid-independent, heterologous activation of protein kinase C (PKC) on the responsiveness of opioid receptor and the underlying molecular mechanisms. Our result showed that removing the C terminus of delta opioid receptor (DOR......) containing six Ser/Thr residues abolished both DPDPE- and phorbol 12-myristate 13-acetate (PMA)-induced DOR phosphorylation. The phosphorylation levels of DOR mutants T352A, T353A, and T358A/T361A/S363S were comparable to that of the wild-type DOR, whereas S344G substitution blocked PMA-induced receptor......, and ionomycin resulted in DOR internalization that required phosphorylation of Ser-344. Expression of dominant negative beta-arrestin and hypertonic sucrose treatment blocked PMA-induced DOR internalization, suggesting that PKC mediates DOR internalization via a beta-arrestin- and clathrin-dependent mechanism...

  4. (D-Pen2,4 prime -125I-Phe4,D-Pen5)enkephalin: A selective high affinity radioligand for delta opioid receptors with exceptional specific activity

    Energy Technology Data Exchange (ETDEWEB)

    Knapp, R.J.; Sharma, S.D.; Toth, G.; Duong, M.T.; Fang, L.; Bogert, C.L.; Weber, S.J.; Hunt, M.; Davis, T.P.; Wamsley, J.K. (Department of Pharmacology, University of Arizona, College of Medicine, Tucson (United States))

    1991-09-01

    (D-Pen2,4{prime}-125I-Phe4,D-Pen5)enkephalin ((125I)DPDPE) is a highly selective radioligand for the delta opioid receptor with a specific activity (2200 Ci/mmol) that is over 50-fold greater than that of tritium-labeled DPDPE analogs. (125I)DPDPE binds to a single site in rat brain membranes with an equilibrium dissociation constant (Kd) value of 421 {plus minus} 67 pM and a receptor density (Bmax) value of 36.4 {plus minus} 2.7 fmol/mg protein. The high affinity of this site for delta opioid receptor ligands and its low affinity for mu or kappa receptor-selective ligands are consistent with its being a delta opioid receptor. The distribution of these sites in rat brain, observed by receptor autoradiography, is also consistent with that of delta opioid receptors. Association and dissociation binding kinetics of 1.0 nM (125I) DPDPE are monophasic at 25 degrees C. The association rate (k + 1 = 5.80 {plus minus} 0.88 {times} 10(7) M-1 min-1) is about 20- and 7-fold greater than that measured for 1.0 nM (3H) DPDPE and 0.8 nM (3H) (D-Pen2,4{prime}-Cl-Phe4, D-Pen5)enkephalin, respectively. The dissociation rate of (125I)DPDPE (0.917 {plus minus} 0.117 {times} 10(-2) min-1) measured at 1.0 nM is about 3-fold faster than is observed for either of the other DPDPE analogs. The rapid binding kinetics of (125I)DPDPE is advantageous because binding equilibrium is achieved with much shorter incubation times than are required for other cyclic enkephalin analogs. This, in addition to its much higher specific activity, makes (125I)DPDPE a valuable new radioligand for studies of delta opioid receptors.

  5. Orexin Receptor Multimerization versus Functional Interactions: Neuropharmacological Implications for Opioid and Cannabinoid Signalling and Pharmacogenetics

    Directory of Open Access Journals (Sweden)

    Miles D. Thompson

    2017-10-01

    Full Text Available Orexins/hypocretins are neuropeptides formed by proteolytic cleavage of a precursor peptide, which are produced by neurons found in the lateral hypothalamus. The G protein-coupled receptors (GPCRs for these ligands, the OX1 and OX2 orexin receptors, are more widely expressed throughout the central nervous system. The orexin/hypocretin system has been implicated in many pathways, and its dysregulation is under investigation in a number of diseases. Disorders in which orexinergic mechanisms are being investigated include narcolepsy, idiopathic sleep disorders, cluster headache and migraine. Human narcolepsy has been associated with orexin deficiency; however, it has only rarely been attributed to mutations in the gene encoding the precursor peptide. While gene variations within the canine OX2 gene hcrtr2 have been directly linked with narcolepsy, the majority of human orexin receptor variants are weakly associated with diseases (the idiopathic sleep disorders, cluster headache and polydipsia-hyponatremia in schizophrenia or are of potential pharmacogenetic significance. Evidence for functional and/or heterodimerization between wild-type variant orexin receptors and opioid and cannabinoid receptors is discussed in the context of its relevance to depression and epilepsy.

  6. Induction of synaptic long-term potentiation after opioid withdrawal.

    Science.gov (United States)

    Drdla, Ruth; Gassner, Matthias; Gingl, Ewald; Sandkühler, Jürgen

    2009-07-10

    mu-Opioid receptor (MOR) agonists represent the gold standard for the treatment of severe pain but may paradoxically also enhance pain sensitivity, that is, lead to opioid-induced hyperalgesia (OIH). We show that abrupt withdrawal from MOR agonists induces long-term potentiation (LTP) at the first synapse in pain pathways. Induction of opioid withdrawal LTP requires postsynaptic activation of heterotrimeric guanine nucleotide-binding proteins and N-methyl-d-aspartate receptors and a rise of postsynaptic calcium concentrations. In contrast, the acute depression by opioids is induced presynaptically at these synapses. Withdrawal LTP can be prevented by tapered withdrawal and shares pharmacology and signal transduction pathways with OIH. These findings provide a previously unrecognized target to selectively combat pro-nociceptive effects of opioids without compromising opioid analgesia.

  7. Flavonoids with M1 Muscarinic Acetylcholine Receptor Binding Activity

    Directory of Open Access Journals (Sweden)

    Meyyammai Swaminathan

    2014-06-01

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

  8. Effects of opioid drugs on dopamine mediated locomotor activity in rats

    Energy Technology Data Exchange (ETDEWEB)

    Leathern, L L

    1986-01-01

    Opioid drugs influence various behavioural parameters including locomotor activity in experimental animals. The interaction between the opioid and dopaminergic systems is one possible explanation for the effect of opioid drugs on locomotor activity. In this study behavioural and biochemical assays were done to investigate the interaction between the opioid and dopaminergic systems. Behavioural studies were done by measurement of locomotor activity (LA) of rats after acute or chronic pretreatment with opioid andor dopaminergic drugs. Biochemical studies were in the form of radioligand binding assays, the effect on the number (Bmax) and affinity (K/sub D/) of receptors was measured after chronic pretreatment with opioid andor dopaminergic drugs. The opioid drugs used are morphine, nalbuphine and naloxone. Dopaminergic drugs used included: agonists-apomorphine and piribedil; antagonists-pimozide, haloperidol, chlorpromazine. In the acute situation increased LA was obtained with morphine and the DA agonists. A correlation between the behavioural and biochemical assays was found. Chronic pretreatment with morphine enhanced apomorphine induced LA, this supersensitivity was also measured as an increased receptor density (Bmax) of D2 receptors in the striatum. Chronic morphine pretreatment caused a decrease in morphine induced LA, while this subsensitivity was not apparent in the ligand binding assays - where no change in receptor number was observed. Chronic naloxone pretreatment enhanced morphine induced LA, as well as increased the Bmax of opioid receptors in the whole brain. It is concluded that an interaction between the opioid and dopaminergic systems does exist, and may account for the mechanism of action of the opioids.

  9. Modulation of melanocortin- induced changes in spinal nociception by µ-opioid receptor agonist and antagonist in neuropathic rats

    NARCIS (Netherlands)

    Gispen, W.H.; Starowitcz, K.; Przewlocki, R.; Przewlocka, B.

    2002-01-01

    Co-localization of opioid and melanocortin receptor expression, especially at the spinal cord level in the dorsal horn and in the gray matter surrounding the central canal led to the suggestion that melanocortins might play a role in nociceptive processes. In the present studies, we aimed to

  10. Immunohistochemical observations of methionine-enkephalin and delta opioid receptor in the digestive system of Octopus ocellatus.

    Science.gov (United States)

    Sha, Ailong; Sun, Hushan; Wang, Yiyan

    2013-02-01

    The study was designed to determine whether methionine-enkephalin (met-Enk) or delta opioid receptor was present in the digestive system of Octopus ocellatus. The results showed that they were both in the bulbus oris, esophagus, crop, stomach, gastric cecum, intestine, posterior salivary glands of O. ocellatus, one of them, met-Enk in the rectum, anterior salivary glands, digestive gland. And the distributions were extensive in the digestive system. Strong or general met-Enk immunoreactivity was observed in the inner epithelial cells of the bulbus oris, esophagus, stomach, gastric cecum, intestine, anterior salivary glands and the adventitia of the intestine and rectum, and so was the delta opioid receptor immunoreactivity in the inner epithelial cells of the bulbus oris, esophagus, and crop, however, they were weak in other parts. Combining with delta opioid receptor, met-Enk may be involved in the regulations of food intake, absorption, movement of gastrointestinal smooth muscle and secretion of digestive gland. The different densities of met-Enk and delta opioid receptor may be related to the different functions in the digestive system of O. ocellatus. Copyright © 2012 Elsevier Ltd. All rights reserved.

  11. Preserved cardiac mitochondrial function and reduced ischaemia/reperfusion injury afforded by chronic continuous hypoxia: Role of opioid receptors

    Czech Academy of Sciences Publication Activity Database

    Maslov, L. N.; Naryzhnaya, N. V.; Prokudina, E. S.; Kolář, František; Gorbunov, A. S.; Zhang, Y.; Wang, H.; Tsibulnikov, S.Yu.; Portnichenko, A. G.; Lasukova, T. V.; Lishmanov, Yu. B.

    2015-01-01

    Roč. 42, č. 5 (2015), s. 496-501 ISSN 1440-1681 R&D Projects: GA ČR(CZ) GAP303/12/1162 Institutional support: RVO:67985823 Keywords : cardioprotection * chronic hypoxia * ischaemia/reperfusion * mitochondrial function * opioid receptors Subject RIV: ED - Physiology Impact factor: 2.004, year: 2015

  12. Polymorphisms of the Kappa Opioid Receptor and Prodynorphin Genes: HIV risk and HIV Natural History

    Science.gov (United States)

    Proudnikov, Dmitri; Randesi, Matthew; Levran, Orna; Yuferov, Vadim; Crystal, Howard; Ho, Ann; Ott, Jurg; Kreek, Mary Jeanne

    2013-01-01

    Objective Studies indicate cross-desensitization between opioid receptors (e.g., kappa opioid receptor, OPRK1), and chemokine receptors (e.g., CXCR4) involved in HIV infection. We tested whether gene variants of OPRK1 and its ligand, prodynorphin (PDYN), influence the outcome of HIV therapy. Methods Three study points, admission to the Women’s Interagency HIV Study (WIHS), initiation of highly active antiretroviral therapy (HAART) and the most recent visit were chosen for analysis as crucial events in the clinical history of the HIV patients. Regression analyses of 17 variants of OPRK1, and 11 variants of PDYN with change of viral load (VL) and CD4 count between admission and initiation of HAART, and initiation of HAART to the most recent visit to WIHS were performed in 598 HIV+ subjects including African Americans, Hispanics and Caucasians. Association with HIV status was done in 1009 subjects. Results Before HAART, greater VL decline (improvement) in carriers of PDYN IVS3+189C>T, and greater increase of CD4 count (improvement) in carriers of OPRK1 −72C>T, were found in African Americans. Also, greater increase of CD4 count in carriers of OPRK1 IVS2+7886A>G, and greater decline of CD4 count (deterioration) in carriers of OPRK1 −1205G>A, were found in Caucasians. After HAART, greater decline of VL in carriers of OPRK1 IVS2+2225G>A, and greater increase of VL in carriers of OPRK1 IVS2+10658G>T and IVS2+10963A>G, were found in Caucasians. Also, a lesser increase of CD4 count was found in Hispanic carriers of OPRK1 IVS2+2225G>A. Conclusion OPRK1 and PDYN polymorphisms may alter severity of HIV infection and response to treatment. PMID:23392455

  13. Ligand recognition by RAR and RXR receptors: binding and selectivity.

    Science.gov (United States)

    Sussman, Fredy; de Lera, Angel R

    2005-10-06

    Fundamental biological functions, most notably embriogenesis, cell growth, cell differentiation, and cell apoptosis, are in part regulated by a complex genomic network that starts with the binding (and activation) of retinoids to their cognate receptors, members of the superfamily of nuclear receptors. We have studied ligand recognition of retinoic receptors (RXRalpha and RARgamma) using a molecular-mechanics-based docking method. The protocol used in this work is able to rank the affinity of pairs of ligands for a single retinoid receptor, the highest values corresponding to those that adapt better to the shape of the binding site and generate the optimal set of electrostatic and apolar interactions with the receptor. Moreover, our studies shed light onto some of the energetic contributions to retinoid receptor ligand selectivity. In this regard we show that there is a difference in polarity between the binding site regions that anchor the carboxylate in RAR and RXR, which translates itself into large differences in the energy of interaction of both receptors with the same ligand. We observe that the latter energy change is canceled off by the solvation energy penalty upon binding. This energy compensation is borne out as well by experiments that address the effect of site-directed mutagenesis on ligand binding to RARgamma. The hypothesis that the difference in binding site polarity might be exploited to build RXR-selective ligands is tested with some compounds having a thiazolidinedione anchoring group.

  14. 5-Hydroxytryptamine 1A/7 and 4alpha receptors differentially prevent opioid-induced inhibition of brain stem cardiorespiratory function.

    Science.gov (United States)

    Wang, Xin; Dergacheva, Olga; Kamendi, Harriet; Gorini, Christopher; Mendelowitz, David

    2007-08-01

    Opioids evoke respiratory depression, bradycardia, and reduced respiratory sinus arrhythmia, whereas serotonin (5-HT) agonists stimulate respiration and cardiorespiratory interactions. This study tested whether serotonin agonists can prevent the inhibitory effects of opioids on cardiorespiratory function. Spontaneous and rhythmic inspiratory-related activity and gamma-aminobutyric acid (GABA) neurotransmission to premotor parasympathetic cardioinhibitory neurons in the nucleus ambiguus were recorded simultaneously in an in vitro thick slice preparation. The mu-opioid agonist fentanyl inhibited respiratory frequency. The 5-hydroxytryptamine 1A/7 receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin increased respiratory frequency by itself and also prevented the fentanyl-induced respiratory depression. The 5-hydroxytryptamine 4alpha agonist BIMU-8 did not by itself change inspiratory activity but prevented the mu-opioid-mediated respiratory depression. Both spontaneous and inspiratory-evoked GABAergic neurotransmission to cardiac vagal neurons were inhibited by fentanyl. 8-Hydroxy-2-(di-n-propylamino)tetralin inhibited spontaneous but not inspiratory-evoked GABAergic activity to parasympathetic cardiac neurons. However, 8-hydroxy-2-(di-n-propylamino)tetralin differentially altered the opioid-mediated depression of inspiratory-evoked GABAergic activity but did not change the opioid-induced reduction in spontaneous GABAergic neurotransmission. In contrast, BIMU-8 did not alter GABAergic neurotransmission to cardiac vagal neurons by itself but prevented the fentanyl depression of both spontaneous and inspiratory-elicited GABAergic neurotransmission to cardiac vagal neurons. In the presence of tetrodotoxin, the inhibition of GABAergic inhibitory postsynaptic currents with fentanyl is prevented by coapplication of BIMU-8, indicating that BIMU-8 acts at presynaptic GABAergic terminals to prevent fentanyl-induced depression. These results suggest that activation of 5

  15. Gender dependent rate of metabolism of the opioid receptor-PET ligand [18F]fluoroethyl-diprenorphine

    International Nuclear Information System (INIS)

    Henriksen, G.; Spilker, M.E.; Hauser, A.I.; Boecker, H.; Schwaiger, M.; Wester, H.J.; Sprenger, T.; Platzer, S.; Toelle, T.R.

    2006-01-01

    Aim: The morphinane-derivate 6-O-(2-[ 18 F]fluoroethyl)-6-O-desmethyldiprenorphine ([ 18 F]FDPN) is a non-selective opioid receptor ligand currently used in positron emission tomography (PET). Correction for plasma metabolites of the arterial input function is necessary for quantitative measurements of [ 18 ]FDPN binding. A study was undertaken to investigate if there are gender dependent differences in the rate of metabolism of [ 18 F]FDPN. Methods: The rate of metabolism of [ 18 F]FDPN was mathematically quantified by fitting a bi-exponential function to each individual's dynamic metabolite data. Results: No statistically significant gender differences were found for age, weight, body mass index or dose. However, significant differences (p 18 F]FDPN faster than men. These differences were found in the contribution of the fast and slow kinetic components of the model describing the distribution of radioactive species in plasma, indicating a higher rate of enzyme-dependent degradation of [ 18 F]FDPN in women than in men. Conclusion: The findings reinforce the need for individualized metabolite correction during [ 18 F]FDPN-PET scans and also indicate that in certain cases, grouping according to gender could be performed in order to minimize methodological errors of the input function prior to kinetic analyses. (orig.)

  16. Endogenous opioid peptides as neurotransmitters in the rat hippocampus

    International Nuclear Information System (INIS)

    Neumaier, J.F.

    1989-01-01

    The role of endogenous opioid peptides as neurotransmitters in the rat hippocampus was investigated by using extracellular recording and radioligand binding techniques in the hippocampal slice preparation. Synaptic conductances from endogenously released opioid peptides have been difficult to detect. This problem was approach by designing a novel assay of opioid peptide release, in which release was detected by measuring binding competition between endogenous opioids and added radioligand. Membrane depolarization displaced [ 3 H]-diprenorphine binding in a transient, calcium-dependent, and peptidase-sensitive manner. Autoradiographic localization of the sites of [ 3 H]-diprenorphine binding displacement showed that significant opioid peptide release and receptor occupancy occurred in each major subregion of the hippocampal slices. This assay method can not be used to define optimal electrical stimulation conditions for releasing endogenous opioids. The binding displacement method was extended to the study of the sigma receptor. Depolarization of hippocampal slices was found to reduce the binding of the sigma-selective radioligand [ 3 H]-ditolylguanidine in a transient and calcium-dependent manner with no apparent direct effects on sigma receptor affinity

  17. Functional role of peripheral opioid receptors in the regulation of cardiac spinal afferent nerve activity during myocardial ischemia

    Science.gov (United States)

    Longhurst, John C.

    2013-01-01

    Thinly myelinated Aδ-fiber and unmyelinated C-fiber cardiac sympathetic (spinal) sensory nerve fibers are activated during myocardial ischemia to transmit the sensation of angina pectoris. Although recent observations showed that myocardial ischemia increases the concentrations of opioid peptides and that the stimulation of peripheral opioid receptors inhibits chemically induced visceral and somatic nociception, the role of opioids in cardiac spinal afferent signaling during myocardial ischemia has not been studied. The present study tested the hypothesis that peripheral opioid receptors modulate cardiac spinal afferent nerve activity during myocardial ischemia by suppressing the responses of cardiac afferent nerve to ischemic mediators like bradykinin and extracellular ATP. The nerve activity of single unit cardiac afferents was recorded from the left sympathetic chain (T2–T5) in anesthetized cats. Forty-three ischemically sensitive afferent nerves (conduction velocity: 0.32–3.90 m/s) with receptive fields in the left and right ventricles were identified. The responses of these afferent nerves to repeat ischemia or ischemic mediators were further studied in the following protocols. First, epicardial administration of naloxone (8 μmol), a nonselective opioid receptor antagonist, enhanced the responses of eight cardiac afferent nerves to recurrent myocardial ischemia by 62%, whereas epicardial application of vehicle (PBS) did not alter the responses of seven other cardiac afferent nerves to ischemia. Second, naloxone applied to the epicardial surface facilitated the responses of seven cardiac afferent nerves to epicardial ATP by 76%. Third, administration of naloxone enhanced the responses of seven other afferent nerves to bradykinin by 85%. In contrast, in the absence of naloxone, cardiac afferent nerves consistently responded to repeated application of ATP (n = 7) or bradykinin (n = 7). These data suggest that peripheral opioid peptides suppress the

  18. Sex Differences in Serotonin 1 Receptor Binding in Rat Brain

    Science.gov (United States)

    Fischette, Christine T.; Biegon, Anat; McEwen, Bruce S.

    1983-10-01

    Male and female rats exhibit sex differences in binding by serotonin 1 receptors in discrete areas of the brain, some of which have been implicated in the control of ovulation and of gonadotropin release. The sex-specific changes in binding, which occur in response to the same hormonal (estrogenic) stimulus, are due to changes in the number of binding sites. Castration alone also affects the number of binding sites in certain areas. The results lead to the conclusion that peripheral hormones modulate binding by serotonin 1 receptors. The status of the serotonin receptor system may affect the reproductive capacity of an organism and may be related to sex-linked emotional disturbances in humans.

  19. Satiety and the role of μ-opioid receptors in the portal vein.

    Science.gov (United States)

    De Vadder, Filipe; Gautier-Stein, Amandine; Mithieux, Gilles

    2013-12-01

    Mu-opioid receptors (MORs) are known to influence food intake at the brain level, through their involvement in the food reward system. MOR agonists stimulate food intake. On the other hand, MOR antagonists suppress food intake. MORs are also active in peripheral organs, especially in the small intestine where they control the gut motility. Recently, an indirect role in the control of food intake was ascribed to MORs in the extrinsic gastrointestinal neural system. MORs present in the neurons of the portal vein walls sense blood peptides released from the digestion of dietary protein. These peptides behave as MOR antagonists. Their MOR antagonist action initiates a gut-brain circuitry resulting in the induction of intestinal gluconeogenesis, a function controlling food intake. Thus, periportal MORs are a key mechanistic link in the satiety effect of protein-enriched diets. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. μ Opioid Receptor Expression after Morphine Administration Is Regulated by miR-212/132 Cluster.

    Directory of Open Access Journals (Sweden)

    Adrian Garcia-Concejo

    Full Text Available Since their discovery, miRNAs have emerged as a promising therapeutical approach in the treatment of several diseases, as demonstrated by miR-212 and its relation to addiction. Here we prove that the miR-212/132 cluster can be regulated by morphine, through the activation of mu opioid receptor (Oprm1. The molecular pathways triggered after morphine administration also induce changes in the levels of expression of oprm1. In addition, miR-212/132 cluster is actively repressing the expression of mu opioid receptor by targeting a sequence in the 3' UTR of its mRNA. These findings suggest that this cluster is closely related to opioid signaling, and function as a post-transcriptional regulator, modulating morphine response in a dose dependent manner. The regulation of miR-212/132 cluster expression is mediated by MAP kinase pathway, CaMKII-CaMKIV and PKA, through the phosphorylation of CREB. Moreover, the regulation of both oprm1 and of the cluster promoter is mediated by MeCP2, acting as a transcriptional repressor on methylated DNA after prolonged morphine administration. This mechanism explains the molecular signaling triggered by morphine as well as the regulation of the expression of the mu opioid receptor mediated by morphine and the implication of miR-212/132 in these processes.

  1. Study of plasma binding of receptor-specific peptides

    OpenAIRE

    Gregor, David

    2008-01-01

    The binding ability of two receptor specific peptides namely 90Y-DOTA-TATE and 111In-DOTA-TATE was studied in therm of interspecies comparison by the method of equilibrium dialysis. This plasma protein binding was different for the chosen animal species (human, rat, rabbit, bovine eventually pork) whereas binding of 90Y-DOTA- TATE was higher than binding of 111In-DOTA-TATE. KEYWORDS: Protein binding, radiofarmaceuticals, equilibrium dialysis, 90Y-DOTA-TATE, 111In- DOTA-TATE

  2. Inhibition of GABAergic Neurotransmission by HIV-1 Tat and Opioid Treatment in the Striatum Involves ?-Opioid Receptors

    OpenAIRE

    Xu, Changqing; Fitting, Sylvia

    2016-01-01

    Due to combined antiretroviral therapy (cART), human immunodeficiency virus type 1 (HIV-1) is considered a chronic disease with high prevalence of mild forms of neurocognitive impairments, also referred to as HIV-associated neurocognitive disorders (HAND). Although opiate drug use can exacerbate HIV-1 Tat-induced neuronal damage, it remains unknown how and to what extent opioids interact with Tat on the GABAergic system. We conducted whole-cell recordings in mouse striatal slices and examined...

  3. Mechanism Governing Human Kappa-Opioid Receptor Expression under Desferrioxamine-Induced Hypoxic Mimic Condition in Neuronal NMB Cells

    Directory of Open Access Journals (Sweden)

    Jennifer Babcock

    2017-01-01

    Full Text Available Cellular adaptation to hypoxia is a protective mechanism for neurons and relevant to cancer. Treatment with desferrioxamine (DFO to induce hypoxia reduced the viability of human neuronal NMB cells. Surviving/attached cells exhibited profound increases of expression of the human kappa-opioid receptor (hKOR and hypoxia inducible factor-1α (HIF-1α. The functional relationship between hKOR and HIF-1α was investigated using RT-PCR, Western blot, luciferase reporter, mutagenesis, siRNA and receptor-ligand binding assays. In surviving neurons, DFO increased HIF-1α expression and its amount in the nucleus. DFO also dramatically increased hKOR expression. Two (designated as HIFC and D out of four potential HIF response elements of the hKOR gene (HIFA–D synergistically mediated the DFO response. Mutation of both elements completely abolished the DFO-induced effect. The CD11 plasmid (containing HIFC and D with an 11 bp spacing produced greater augmentation than that of the CD17 plasmid (HIFC and D with a 17 bp-spacing, suggesting that a proper topological interaction of these elements synergistically enhanced the promoter activity. HIF-1α siRNA knocked down the increase of endogenous HIF-1α messages and diminished the DFO-induced increase of hKOR expression. Increased hKOR expression resulted in the up-regulation of hKOR protein. In conclusion, the adaptation of neuronal hKOR under hypoxia was governed by HIF-1, revealing a new mechanism of hKOR regulation.

  4. Morphine-induced internalization of the L83I mutant of the rat μ-opioid receptor.

    Science.gov (United States)

    Cooke, A E; Oldfield, S; Krasel, C; Mundell, S J; Henderson, G; Kelly, E

    2015-01-01

    Naturally occurring single-nucleotide polymorphisms (SNPs) within GPCRs can result in alterations in various pharmacological parameters. Understanding the regulation and function of endocytic trafficking of the μ-opioid receptor (MOP receptor) is of great importance given its implication in the development of opioid tolerance. This study has compared the agonist-dependent trafficking and signalling of L83I, the rat orthologue of a naturally occurring variant of the MOP receptor. Cell surface elisa, confocal microscopy and immunoprecipitation assays were used to characterize the trafficking properties of the MOP-L83I variant in comparison with the wild-type receptor in HEK 293 cells. Functional assays were used to compare the ability of the L83I variant to signal to several downstream pathways. Morphine-induced internalization of the L83I MOP receptor was markedly increased in comparison with the wild-type receptor. The altered trafficking of this variant was found to be specific to morphine and was both G-protein receptor kinase- and dynamin-dependent. The enhanced internalization of L83I variant in response to morphine was not due to increased phosphorylation of serine 375, arrestin association or an increased ability to signal. These results suggest that morphine promotes a specific conformation of the L83I variant that makes it more liable to internalize in response to morphine, unlike the wild-type receptor that undergoes significantly less morphine-stimulated internalization, providing an example of a ligand-selective biased receptor. The presence of this SNP within an individual may consequently affect the development of tolerance and analgesic responses. This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2. © 2014 The British Pharmacological Society.

  5. Internalisation of the mu-opioid receptor by endomorphin-1 and leu-enkephalin is dependant on aromatic amino acid residues.

    Science.gov (United States)

    Del Borgo, Mark P; Blanchfield, Joanne T; Toth, Istvan

    2008-04-15

    The opioid receptor system in the central nervous system controls a number of physiological processes, most notably pain. However, most opioids currently available have a variety of side-effects as well as exhibiting tolerance. Tolerance is most likely to be a complex phenomenon, however, the role of receptor internalisation is thought to play a crucial role. In this study, we examined the role of aromaticity in ligand-mediated receptor internalisation of the mu-opioid receptor (MOPR). These studies show that the amount of receptor internalisation may be dependant on the amphiphilicity of the ligand. Specifically, deletion of the C-terminus aromatic residues of endomorphin 1, particularly tryptophan reduces receptor-mediated internalisation whilst the addition of tryptophan within the enkephalin sequence increases receptor internalisation and decreases tolerance.

  6. Deltorphins: a family of naturally occurring peptides with high affinity and selectivity for delta opioid binding sites.

    OpenAIRE

    Erspamer, V; Melchiorri, P; Falconieri-Erspamer, G; Negri, L; Corsi, R; Severini, C; Barra, D; Simmaco, M; Kreil, G

    1989-01-01

    Deltorphins are endogenous linear heptapeptides, isolated from skin extracts of frogs belonging to the genus Phyllomedusa, that have a higher affinity and selectivity for delta opioid binding sites than any other natural compound known. Two deltorphins with the sequence Tyr-Ala-Phe-Asp(or Glu)-Val-Val-Gly-NH2 have been isolated from skin extracts of Phyllomedusa bicolor. The alanine in position 2 is in the D configuration. These peptides, [D-Ala2]deltorphins I and II, show an even higher affi...

  7. Methodological aspects on drug receptor binding analysis

    International Nuclear Information System (INIS)

    Wahlstroem, A.

    1978-01-01

    Although drug receptors occur in relatively low concentrations, they can be visualized by the use of appropriate radioindicators. In most cases the procedure is rapid and can reach a high degree of accuracy. Specificity of the interaction is studied by competition analysis. The necessity of using several radioindicators to define a receptor population is emphasized. It may be possible to define isoreceptors and drugs with selectivity for one isoreceptor. (Author)

  8. Endomorphin-2: a biased agonist at the μ-opioid receptor.

    Science.gov (United States)

    Rivero, Guadalupe; Llorente, Javier; McPherson, Jamie; Cooke, Alex; Mundell, Stuart J; McArdle, Craig A; Rosethorne, Elizabeth M; Charlton, Steven J; Krasel, Cornelius; Bailey, Christopher P; Henderson, Graeme; Kelly, Eamonn

    2012-08-01

    Previously we correlated the efficacy for G protein activation with that for arrestin recruitment for a number of agonists at the μ-opioid receptor (MOPr) stably expressed in HEK293 cells. We suggested that the endomorphins (endomorphin-1 and -2) might be biased toward arrestin recruitment. In the present study, we investigated this phenomenon in more detail for endomorphin-2, using endogenous MOPr in rat brain as well as MOPr stably expressed in HEK293 cells. For MOPr in neurons in brainstem locus ceruleus slices, the peptide agonists [d-Ala(2),N-Me-Phe(4),Gly(5)-ol]-enkephalin (DAMGO) and endomorphin-2 activated inwardly rectifying K(+) current in a concentration-dependent manner. Analysis of these responses with the operational model of pharmacological agonism confirmed that endomorphin-2 had a much lower operational efficacy for G protein-mediated responses than did DAMGO at native MOPr in mature neurons. However, endomorphin-2 induced faster desensitization of the K(+) current than did DAMGO. In addition, in HEK293 cells stably expressing MOPr, the ability of endomorphin-2 to induce phosphorylation of Ser375 in the COOH terminus of the receptor, to induce association of arrestin with the receptor, and to induce cell surface loss of receptors was much more efficient than would be predicted from its efficacy for G protein-mediated signaling. Together, these results indicate that endomorphin-2 is an arrestin-biased agonist at MOPr and the reason for this is likely to be the ability of endomorphin-2 to induce greater phosphorylation of MOPr than would be expected from its ability to activate MOPr and to induce activation of G proteins.

  9. Tachykinin NK₁ receptor antagonist co-administration attenuates opioid withdrawal-mediated spinal microglia and astrocyte activation.

    Science.gov (United States)

    Tumati, Suneeta; Largent-Milnes, Tally M; Keresztes, Attila I; Yamamoto, Takashi; Vanderah, Todd W; Roeske, William R; Hruby, Victor J; Varga, Eva V

    2012-06-05

    Prolonged morphine treatment increases pain sensitivity in many patients. Enhanced spinal Substance P release is one of the adaptive changes associated with sustained opioid exposure. In addition to pain transmitting second order neurons, spinal microglia and astrocytes also express functionally active Tachykinin NK₁ (Substance P) receptors. In the present work we investigated the role of glial Tachykinin NK₁ receptors in morphine withdrawal-mediated spinal microglia and astrocyte activation. Our data indicate that intrathecal co-administration (6 days, twice daily) of a selective Tachykinin NK₁ receptor antagonist (N-acetyl-L-tryptophan 3,5-bis(trifluoromethyl)benzylester (L-732,138; 20 μg/injection)) attenuates spinal microglia and astrocyte marker and pro-inflammatory mediator immunoreactivity as well as hyperalgesia in withdrawn rats. Furthermore, covalent linkage of the opioid agonist with a Tachykinin NK₁ antagonist pharmacophore yielded a bivalent compound that did not augment spinal microglia or astrocyte marker or pro-inflammatory mediator immunoreactivity and did not cause paradoxical pain sensitization upon drug withdrawal. Thus, bivalent opioid/Tachykinin NK₁ receptor antagonists may provide a novel paradigm for long-term pain management.

  10. Sex Differences in Kappa Opioid Receptor Function and Their Potential Impact on Addiction

    Science.gov (United States)

    Chartoff, Elena H.; Mavrikaki, Maria

    2015-01-01

    Behavioral, biological, and social sequelae that lead to drug addiction differ between men and women. Our efforts to understand addiction on a mechanistic level must include studies in both males and females. Stress, anxiety, and depression are tightly linked to addiction, and whether they precede or result from compulsive drug use depends on many factors, including biological sex. The neuropeptide dynorphin (DYN), an endogenous ligand at kappa opioid receptors (KORs), is necessary for stress-induced aversive states and is upregulated in the brain after chronic exposure to drugs of abuse. KOR agonists produce signs of anxiety, fear, and depression in laboratory animals and humans, findings that have led to the hypothesis that drug withdrawal-induced DYN release is instrumental in negative reinforcement processes that drive addiction. However, these studies were almost exclusively conducted in males. Only recently is evidence available that there are sex differences in the effects of KOR activation on affective state. This review focuses on sex differences in DYN and KOR systems and how these might contribute to sex differences in addictive behavior. Much of what is known about how biological sex influences KOR systems is from research on pain systems. The basic molecular and genetic mechanisms that have been discovered to underlie sex differences in KOR function in pain systems may apply to sex differences in KOR function in reward systems. Our goals are to discuss the current state of knowledge on how biological sex contributes to KOR function in the context of pain, mood, and addiction and to explore potential mechanisms for sex differences in KOR function. We will highlight evidence that the function of DYN-KOR systems is influenced in a sex-dependent manner by: polymorphisms in the prodynorphin (pDYN) gene, genetic linkage with the melanocortin-1 receptor (MC1R), heterodimerization of KORs and mu opioid receptors (MORs), and gonadal hormones. Finally, we

  11. Sex differences in kappa opioid receptor function and their potential impact on addiction

    Directory of Open Access Journals (Sweden)

    Elena eChartoff

    2015-12-01

    Full Text Available Behavioral, biological and social sequelae that lead to drug addiction differ between men and women. Our efforts to understand addiction on a mechanistic level must include studies in both males and females. Stress, anxiety, and depression are tightly linked to addiction, and whether they precede or result from compulsive drug use depends on many factors, including biological sex. The neuropeptide dynorphin (DYN, an endogenous ligand at kappa opioid receptors (KORs, is necessary for stress-induced aversive states and is upregulated in the brain after chronic exposure to drugs of abuse. KOR agonists produce signs of anxiety, fear, and depression in laboratory animals and humans, findings that have led to the hypothesis that drug withdrawal-induced DYN release is instrumental in negative reinforcement processes that drive addiction. However, these studies were almost exclusively conducted in males. Only recently is evidence available that there are sex differences in the effects of KOR activation on affective state. This review focuses on sex differences in DYN and KOR systems and how these might contribute to sex differences in addictive behavior. Much of what is known about how biological sex influences KOR systems is from research on pain systems. The basic molecular and genetic mechanisms that have been discovered to underlie sex differences in KOR function in pain systems may apply to sex differences in KOR function in reward systems. Our goals are to discuss the current state of knowledge on how biological sex contributes to KOR function in the context of pain,mood and addiction and to explore potential mechanisms for sex differences in KOR function. We will highlight evidence that the function of DYN-KOR systems is influenced in a sex-dependent manner by: polymorphisms in the prodynorphin (pDYN gene, genetic linkage with the melanocortin-1 receptor (MC1R, heterodimerization of KORs and mu opioid receptors (MORs, and gonadal hormones

  12. Sex Differences in Kappa Opioid Receptor Function and Their Potential Impact on Addiction.

    Science.gov (United States)

    Chartoff, Elena H; Mavrikaki, Maria

    2015-01-01

    Behavioral, biological, and social sequelae that lead to drug addiction differ between men and women. Our efforts to understand addiction on a mechanistic level must include studies in both males and females. Stress, anxiety, and depression are tightly linked to addiction, and whether they precede or result from compulsive drug use depends on many factors, including biological sex. The neuropeptide dynorphin (DYN), an endogenous ligand at kappa opioid receptors (KORs), is necessary for stress-induced aversive states and is upregulated in the brain after chronic exposure to drugs of abuse. KOR agonists produce signs of anxiety, fear, and depression in laboratory animals and humans, findings that have led to the hypothesis that drug withdrawal-induced DYN release is instrumental in negative reinforcement processes that drive addiction. However, these studies were almost exclusively conducted in males. Only recently is evidence available that there are sex differences in the effects of KOR activation on affective state. This review focuses on sex differences in DYN and KOR systems and how these might contribute to sex differences in addictive behavior. Much of what is known about how biological sex influences KOR systems is from research on pain systems. The basic molecular and genetic mechanisms that have been discovered to underlie sex differences in KOR function in pain systems may apply to sex differences in KOR function in reward systems. Our goals are to discuss the current state of knowledge on how biological sex contributes to KOR function in the context of pain, mood, and addiction and to explore potential mechanisms for sex differences in KOR function. We will highlight evidence that the function of DYN-KOR systems is influenced in a sex-dependent manner by: polymorphisms in the prodynorphin (pDYN) gene, genetic linkage with the melanocortin-1 receptor (MC1R), heterodimerization of KORs and mu opioid receptors (MORs), and gonadal hormones. Finally, we

  13. Evidence of morphine like substance and μ-opioid receptor expression in Toxacara canis (Nematoda: Ascaridae).

    Science.gov (United States)

    Golabi, Mostafa; Naem, Soraya; Imani, Mehdi; Dalirezh, Nowruz

    2016-01-01

    Toxocara canis (Nematoda: Ascaridae) is an intestinal nematode parasite of dogs, which can also cause disease in humans. Transmission to humans usually occurs because of direct contact with T. canis eggs present in soil contaminated with the feces of infected dogs. This nematode has extraordinary abilities to survive for many years in different tissues of vertebrates, and develop to maturity in the intestinal tract of its definitive host. Survival of parasitic nematodes within a host requires immune evasion using complicated pathways. Morphine-like substance, as well as opioids, which are known as down regulating agents, can modulate both innate and acquired immune responses, and let the parasite survives in their hosts. In the present study, we aimed to find evidences of morphine-like substance and µ-opiate receptor expression in T. canis , using high performance liquid chromatography (HPLC) and reverse transcription polymerase chain reaction (RT-PCR). The results indicated that T. canis produced morphine-like substances at the level of 2.31± 0.26 ng g -1 wet weight, and expressed µ-opiate receptor as in expected size of 441 bp. According to our findings, it was concluded that T. canis , benefits using morphine-like substance to modulate host immunity.

  14. Ivy and neurogliaform interneurons are a major target of μ opioid receptor modulation

    Science.gov (United States)

    Krook-Magnuson, Esther; Luu, Lillian; Lee, Sang-Hun; Varga, Csaba; Soltesz, Ivan

    2011-01-01

    Mu opioid receptors (μORs) are selectively expressed on interneurons in area CA1 of the hippocampus. Fast-spiking, parvalbumin expressing, basket cells express μORs, but circumstantial evidence suggests that another major, unidentified, GABAergic cell class must also be modulated by μORs. Here we report that the abundant, dendritically targeting, neurogliaform family of cells (Ivy and neurogliaform cells) is a previously unrecognized target of direct modulation by μORs. Ivy and neurogliaform cells are not only numerous, but also have unique properties, including promiscuous gap junctions formed with various interneuronal subtypes, volume transmission, and the ability to produce a postsynaptic GABAB response after a single presynaptic spike. Using a mouse line expressing green fluorescent protein under the neuropeptide Y promoter, we find that across all layers of CA1, activation of μORs hyperpolarizes Ivy and neurogliaform cells. Further, paired recordings between synaptically coupled Ivy and pyramidal cells show that Ivy cell terminals are dramatically inhibited by μOR-activation. Effects in Ivy and neurogliaform cells are seen at similar concentrations of agonist as those producing inhibition in fast-spiking PV basket cells. We also report that Ivy cells display the recently described phenomenon of persistent firing, a state of continued firing in the absence of continued input, and that induction of persistent firing is inhibited by μOR-activation. Together these findings identify a major, previously unrecognized, target of μOR-modulation. Given the prominence of this cell type in and beyond CA1, as well as its unique role in microcircuitry, opioid modulation of neurogliaform cells has wide implications. PMID:22016519

  15. Ivy and neurogliaform interneurons are a major target of μ-opioid receptor modulation.

    Science.gov (United States)

    Krook-Magnuson, Esther; Luu, Lillian; Lee, Sang-Hun; Varga, Csaba; Soltesz, Ivan

    2011-10-19

    μ-Opioid receptors (μORs) are selectively expressed on interneurons in area CA1 of the hippocampus. Fast-spiking, parvalbumin-expressing, basket cells express μORs, but circumstantial evidence suggests that another major, unidentified, GABAergic cell class must also be modulated by μORs. Here we report that the abundant, dendritically targeting, neurogliaform family of cells (Ivy and neurogliaform cells) is a previously unrecognized target of direct modulation by μORs. Ivy and neurogliaform cells are not only numerous but also have unique properties, including promiscuous gap junctions formed with various interneuronal subtypes, volume transmission, and the ability to produce a postsynaptic GABA(B) response after a single presynaptic spike. Using a mouse line expressing green fluorescent protein under the neuropeptide Y promoter, we find that, across all layers of CA1, activation of μORs hyperpolarizes Ivy and neurogliaform cells. Furthermore, paired recordings between synaptically coupled Ivy and pyramidal cells show that Ivy cell terminals are dramatically inhibited by μOR activation. Effects in Ivy and neurogliaform cells are seen at similar concentrations of agonist as those producing inhibition in fast-spiking parvalbumin basket cells. We also report that Ivy cells display the recently described phenomenon of persistent firing, a state of continued firing in the absence of continued input, and that induction of persistent firing is inhibited by μOR activation. Together, these findings identify a major, previously unrecognized, target of μOR modulation. Given the prominence of this cell type in and beyond CA1, as well as its unique role in microcircuitry, opioid modulation of neurogliaform cells has wide implications.

  16. Role and psychological dependenci arrangement of opioid by type of reseptor opioid

    OpenAIRE

    Arif Nurrochmad, Arif Nurrochmad

    2015-01-01

    Opioid receptor can be classified as p., 8, and K-opioid receptor that widely expressed in the CNS. The development of selective receptor agonist and cloning of each receptor have contributed greatly to our increasing knowledge of the neuropharmacological profile of each opioid receptor type. This review focuses on the functional interaction among these opioid receptor types that contribute to opioid dependence especially in psychological dependence. Several lines of evidence provide argument...

  17. δ-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    International Nuclear Information System (INIS)

    Heiss, Anika; Ammer, Hermann; Eisinger, Daniela A.

    2009-01-01

    δ-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen 2,5 ]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory G i/o proteins, because pre-treatment with pertussis toxin, but not over-expression of the G q/11 scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the Gβγ scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  18. {delta}-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    Energy Technology Data Exchange (ETDEWEB)

    Heiss, Anika; Ammer, Hermann [Institute of Pharmacology, Toxicology and Pharmacy Ludwig-Maximilians-University of Munich Koeniginstrasse 16 80539 Muenchen Federal Republic of Germany (Germany); Eisinger, Daniela A., E-mail: eisinger@pharmtox.vetmed.uni-muenchen.de [Institute of Pharmacology, Toxicology and Pharmacy Ludwig-Maximilians-University of Munich Koeniginstrasse 16 80539 Muenchen Federal Republic of Germany (Germany)

    2009-07-15

    {delta}-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen{sup 2,5}]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory G{sub i/o} proteins, because pre-treatment with pertussis toxin, but not over-expression of the G{sub q/11} scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the G{beta}{gamma} scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  19. Morphine reduces the threshold of helium preconditioning against myocardial infarction: the role of opioid receptors in rabbits

    Science.gov (United States)

    Pagel, Paul S.; Krolikowski, John G.; Amour, Julien; Warltier, David C.; Weihrauch, Dorothee

    2015-01-01

    Objectives Brief, repetitive administration of helium before prolonged coronary artery occlusion and reperfusion protects myocardium against infarction. Opioid receptors mediate the cardioprotective effects of ischemic pre- and postconditioning, but whether these receptors also play a role in helium preconditioning is unknown. We tested the hypotheses that opioid receptors mediate helium preconditioning and that morphine (a μ1-opioid receptor agonist with δ1-opioid agonist properties) lowers the threshold of cardioprotection produced by helium in vivo. Design Randomized, prospective study. Setting University research laboratory. Participants Male New Zealand white rabbits. Interventions Rabbits (n=56) were instrumented for measurement of systemic hemodynamics and subjected to a 30 min left anterior descending coronary artery (LAD) occlusion and 3 h reperfusion. In separate experimental groups, rabbits (n=6 or 7 per group) received 0.9% saline (control), one or three cycles of 70% helium-30% oxygen administered for 5 min interspersed with 5 min of an air-oxygen mixture, morphine (0.1 mg/kg, i.v.), or the nonselective opioid antagonist naloxone (6 mg/kg, i.v.) before LAD occlusion. Other groups of rabbits received three cycles of helium or one cycle of helium plus morphine (0.1 mg/kg) in the absence or presence of naloxone (6 mg/kg) before ischemia and reperfusion. Statistical analysis of data was performed with analysis of variance for repeated measures followed by Bonferroni’s modification of Student’s t test. Measurements and Main Results Myocardial infarct size was determined using triphenyltetrazolium chloride staining and presented as a percentage of the left ventricular area at risk. Helium reduced myocardial infarct size in an exposure-related manner [36±6 (P>0.05) and 25±4% (P<0.05 versus control) for one and three cycles of helium, respectively; data are mean±SD] compared with control (44±7%). Morphine and naloxone alone did not affect infarct

  20. Growth inhibition of thyroid follicular cell-derived cancers by the opioid growth factor (OGF) - opioid growth factor receptor (OGFr) axis

    International Nuclear Information System (INIS)

    McLaughlin, Patricia J; Zagon, Ian S; Park, Sunny S; Conway, Andrea; Donahue, Renee N; Goldenberg, David

    2009-01-01

    Carcinoma of the thyroid gland is an uncommon cancer, but the most frequent malignancy of the endocrine system. Most thyroid cancers are derived from the follicular cell. Follicular carcinoma (FTC) is considered more malignant than papillary thyroid carcinoma (PTC), and anaplastic thyroid cancer (ATC) is one of the most lethal human cancers. Opioid Growth Factor (OGF; chemical term - [Met 5 ]-enkephalin) and its receptor, OGFr, form an inhibitory axis regulating cell proliferation. Both the peptide and receptor have been detected in a wide variety of cancers, and OGF is currently used clinically as a biotherapy for some non-thyroid neoplasias. This study addressed the question of whether the OGF-OGFr axis is present and functional in human thyroid follicular cell - derived cancer. Utilizing human ATC (KAT-18), PTC (KTC-1), and FTC (WRO 82-1) cell lines, immunohistochemistry was employed to ascertain the presence and location of OGF and OGFr. The growth characteristics in the presence of OGF or the opioid antagonist naltrexone (NTX), and the specificity of opioid peptides for proliferation of ATC, were established in KAT-18 cells. Dependence on peptide and receptor were investigated using neutralization studies with antibodies and siRNA experiments, respectively. The mechanism of peptide action on DNA synthesis and cell survival was ascertained. The ubiquity of the OGF-OGFr axis in thyroid follicular cell-derived cancer was assessed in KTC-1 (PTC) and WRO 82-1 (FTC) tumor cells. OGF and OGFr were present in KAT-18 cells. Concentrations of 10 -6 M OGF inhibited cell replication up to 30%, whereas NTX increased cell growth up to 35% relative to cultures treated with sterile water. OGF treatment reduced cell number by as much as 38% in KAT-18 ATC in a dose-dependent and receptor-mediated manner. OGF antibodies neutralized the inhibitory effects of OGF, and siRNA knockdown of OGFr negated growth inhibition by OGF. Cell survival was not altered by OGF, but DNA synthesis

  1. Local analgesic effect of tramadol is mediated by opioid receptors in late postoperative pain after plantar incision in rats.

    Science.gov (United States)

    de Oliveira Junior, José Oswaldo; de Freitas, Milena Fernandes; Bullara de Andrade, Carolina; Chacur, Marucia; Ashmawi, Hazem Adel

    2016-01-01

    Tramadol is a drug used to treat moderate to severe pain. It is known to present a peripheral effect, but the local mechanisms underlying its actions remain unclear. The role of peripheral opioid receptors in postoperative pain is not well understood. In the present study, we examined the peripheral opioid receptors to determine the local effect of tramadol in a plantar incision pain model. Rats were subjected to plantar incision and divided into four groups on postoperative day (POD) 1: SF_SF, 0.9% NaCl injected into the right hindpaw; SF_TraI, 0.9% NaCl and tramadol injected into the right hindpaw; SF_TraC, 0.9% NaCl and tramadol injected into the contralateral hindpaw; and Nal_Tra, naloxone and tramadol injected into the ipsilateral hindpaw. To determine the animals' nociceptive threshold, mechanical hyperalgesia was measured before incision, on POD1 before treatment and at 15, 30, 45, and 60 minutes after the incision. The same procedure was repeated on the POD2. The expression levels of μ-opioid receptor (MOR) and δ-opioid receptor (DOR) were obtained through immunoblotting assays in the lumbar dorsal root ganglia (L3-L6) in naïve rats and 1, 2, 3, and 7 days after the incision. Our results showed that the plantar incision was able to cause an increase in mechanical hyperalgesia and that tramadol reversed this hyperalgesia on POD1 and POD2. Tramadol injections in the contralateral paw did not affect the animals' nociceptive threshold. Naloxone was able to antagonize the tramadol effect partially on POD1 and completely on POD2. The DOR expression increased on POD2, POD3, and POD7, whereas the MOR expression did not change. Together, our results show that tramadol promoted a local analgesic effect in the postoperative pain model that was antagonized by naloxone in POD2, alongside the increase of DOR expression.

  2. Local analgesic effect of tramadol is not mediated by opioid receptors in early postoperative pain in rats

    Directory of Open Access Journals (Sweden)

    Angela Maria Sousa

    2015-06-01

    Full Text Available BACKGROUND AND OBJECTIVES: Tramadol is known as a central acting analgesic drug, used for the treatment of moderate to severe pain. Local analgesic effect has been demonstrated, in part due to local anesthetic-like effect, but other mechanisms remain unclear. The role of peripheral opioid receptors in the local analgesic effect is not known. In this study, we examined role of peripheral opioid receptors in the local analgesic effect of tramadol in the plantar incision model. METHODS: Young male Wistar rats were divided into seven groups: control, intraplantar tramadol, intravenous tramadol, intravenous naloxone-intraplantar tramadol, intraplantar naloxone-intraplantar tramadol, intravenous naloxone-intravenous tramadol, and intravenous naloxone. After receiving the assigned drugs (tramadol 5 mg, naloxone 200 µg or 0.9% NaCl, rats were submitted to plantar incision, and withdrawal thresholds after mechanical stimuli with von Frey filaments were assessed at baseline, 10, 15, 30, 45 and 60 min after incision. RESULTS: Plantar incision led to marked mechanical hyperalgesia during the whole period of observation in the control group, no mechanical hyperalgesia were observed in intraplantar tramadol group, intraplantar naloxone-intraplantar tramadol group and intravenous naloxone-intraplantar tramadol. In the intravenous tramadol group a late increase in withdrawal thresholds (after 45 min was observed, the intravenous naloxone-intravenous tramadol group and intravenous naloxone remained hyperalgesic during the whole period. CONCLUSIONS: Tramadol presented an early local analgesic effect decreasing mechanical hyperalgesia induced by plantar incision. This analgesic effect was not mediated by peripheral opioid receptors.

  3. Nalfurafine hydrochloride, a selective κ opioid receptor agonist, has no reinforcing effect on intravenous self-administration in rhesus monkeys

    Directory of Open Access Journals (Sweden)

    Kaoru Nakao

    2016-01-01

    Full Text Available Nalfurafine hydrochloride [(E-N-[17-(cyclopropylmethyl-4,5α-epoxy-3,14-dihydroxymorphinan-6β-yl]-3-(furan-3-yl-N-methylprop-2-enamide monohydrochloride; nalfurafine] is used in Japan as an antipruritic for the treatment of intractable pruritus in patients undergoing hemodialysis or with chronic liver disease. It is a potent and selective agonist at the κ opioid receptor, but also has weak and partial agonist activity at μ opioid receptors. Opioids, especially those acting at μ receptors, carry a risk of abuse. This is an important factor in the consideration of therapeutic risk vs. benefit in clinical use and the potential for misuse as a public health problem. It is therefore necessary to carefully evaluate the reinforcing effects of nalfurafine. To this end, we investigated intravenous self-administration of nalfurafine in rhesus monkeys. The number of self-administration of nalfurafine at doses of 0.0625, 0.125 and 0.25 μg/kg/infusion was not higher than that of saline in rhesus monkeys that frequently self-administered pentazocine (0.25 mg/kg/infusion. These results indicate that nalfurafine has no reinforcing effect in rhesus monkeys in the intravenous self-administration paradigm.

  4. Kappa opioid receptor antagonism and chronic antidepressant treatment have beneficial activities on social interactions and grooming deficits during heroin abstinence.

    Science.gov (United States)

    Lalanne, L; Ayranci, G; Filliol, D; Gavériaux-Ruff, C; Befort, K; Kieffer, B L; Lutz, P-E

    2017-07-01

    Addiction is a chronic brain disorder that progressively invades all aspects of personal life. Accordingly, addiction to opiates severely impairs interpersonal relationships, and the resulting social isolation strongly contributes to the severity and chronicity of the disease. Uncovering new therapeutic strategies that address this aspect of addiction is therefore of great clinical relevance. We recently established a mouse model of heroin addiction in which, following chronic heroin exposure, 'abstinent' mice progressively develop a strong and long-lasting social avoidance phenotype. Here, we explored and compared the efficacy of two pharmacological interventions in this mouse model. Because clinical studies indicate some efficacy of antidepressants on emotional dysfunction associated with addiction, we first used a chronic 4-week treatment with the serotonergic antidepressant fluoxetine, as a reference. In addition, considering prodepressant effects recently associated with kappa opioid receptor signaling, we also investigated the kappa opioid receptor antagonist norbinaltorphimine (norBNI). Finally, we assessed whether fluoxetine and norBNI could reverse abstinence-induced social avoidance after it has established. Altogether, our results show that two interspaced norBNI administrations are sufficient both to prevent and to reverse social impairment in heroin abstinent animals. Therefore, kappa opioid receptor antagonism may represent a useful approach to alleviate social dysfunction in addicted individuals. © 2016 Society for the Study of Addiction.

  5. Pain, opioids, and sleep: implications for restless legs syndrome treatment.

    Science.gov (United States)

    Trenkwalder, Claudia; Zieglgänsberger, Walter; Ahmedzai, Sam H; Högl, Birgit

    2017-03-01

    Opioid receptor agonists are known to relieve restless legs syndrome (RLS) symptoms, including both sensory and motor events, as well as improving sleep. The mechanisms of action of opioids in RLS are still a matter of speculation. The mechanisms by which endogenous opioids contribute to the pathophysiology of this polygenetic disorder, in which there are a number of variants, including developmental factors, remains unknown. A summary of the cellular mode of action of morphine and its (partial) antagonist naloxone via α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and the involvement of dendritic spine activation is described. By targeting pain and its consequences, opioids are the first-line treatment in many diseases and conditions with both acute and chronic pain and have thus been used in both acute and chronic pain conditions over the last 40 years. Addiction, dependence, and tolerability of opioids show a wide variability interindividually, as the response to opioids is influenced by a complex combination of genetic, molecular, and phenotypic factors. Although several trials have now addressed opioid treatment in RLS, hyperalgesia as a complication of long-term opioid treatment, or opioid-opioid interaction have not received much attention so far. Therapeutic opioids may act not only on opioid receptors but also via histamine or N-methyl-d-aspartate (NMDA) receptors. In patients with RLS, one of the few studies investigating opioid bindings found that possible brain regions involved in the severity of RLS symptoms are similar to those known to be involved in chronic pain, such as the medial pain system (medial thalamus, amygdala, caudate nucleus, anterior cingulate gyrus, insular cortex, and orbitofrontal cortex). The results of this diprenorphine positron emission tomography study suggested that the more severe the RLS, the greater the release of endogenous opioids. Since 1993, when the first small controlled study was performed with

  6. Novel Drosophila receptor that binds multiple growth factors

    International Nuclear Information System (INIS)

    Rosner, M.R.; Thompson, K.L.; Garcia, V.; Decker, S.J.

    1986-01-01

    The authors have recently reported the identification of a novel growth factor receptor from Drosophila cell cultures that has dual binding specificity for both insulin and epidermal growth factor (EGF). This 100 kDa protein is also antigenically related to the cytoplasmic region of the mammalian EGF receptor-tyrosine kinase. They now report that this protein binds to mammalian nerve growth factor and human transforming growth factor alpha as well as insulin and EGF with apparent dissociation constants ranging from 10 -6 to 10 -8 M. The 100 kDa protein can be affinity-labeled with these 125 I-labeled growth factors after immunoprecipitation with anti-EGF receptor antiserum. These four growth factors appear to share a common binding site, as evidenced by their ability to block affinity labelling by 125 I-insulin. No significant binding to the 100 kDa protein was observed with platelet-derived growth factor, transforming growth factor beta, or glucagon. The 100 kDa Drosophila protein has a unique ligand-binding spectrum with no direct counterpart in mammalian cells and may represent an evolutionary precursor of the mammalian receptors for these growth factors

  7. Discovery of a novel site of opioid action at the innate immune pattern-recognition receptor TLR4 and its role in addiction.

    Science.gov (United States)

    Jacobsen, Jonathan Henry W; Watkins, Linda R; Hutchinson, Mark R

    2014-01-01

    Opioids have historically, and continue to be, an integral component of pain management. However, despite pharmacokinetic and dynamic optimization over the past 100 years, opioids continue to produce many undesirable side effects such as tolerance, reward, and dependence. As such, opioids are liable for addiction. Traditionally, opioid addiction was viewed as a solely neuronal process, and while substantial headway has been made into understanding the molecular and cellular mechanisms mediating this process, research has however, been relatively ambivalent to how the rest of the central nervous system (CNS) responds to opioids. Evidence over the past 20 years has clearly demonstrated the importance of the immunocompetent cells of the CNS (glia) in many aspects of opioid pharmacology. Particular focus has been placed on microglia and astrocytes, who in response to opioids, become activated and release inflammatory mediators. Importantly, the mechanism underlying immune activation is beginning to be elucidated. Evidence suggests an innate immune pattern-recognition receptor (toll-like receptor 4) as an integral component underlying opioid-induced glial activation. The subsequent proinflammatory response may be viewed akin to neurotransmission creating a process termed central immune signaling. Translationally, we are beginning to appreciate the importance of central immune signaling as it contributes to many behavioral actions of addiction including reward, withdrawal, and craving. As such, the aim of this chapter is to review and integrate the neuronal and central immune signaling perspective of addiction. © 2014 Elsevier Inc. All rights reserved.

  8. Protection of adult rat cardiac myocytes from ischemic cell death: role of caveolar microdomains and delta-opioid receptors.

    Science.gov (United States)

    Patel, Hemal H; Head, Brian P; Petersen, Heidi N; Niesman, Ingrid R; Huang, Diane; Gross, Garrett J; Insel, Paul A; Roth, David M

    2006-07-01

    The role of caveolae, membrane microenvironments enriched in signaling molecules, in myocardial ischemia is poorly defined. In the current study, we used cardiac myocytes prepared from adult rats to test the hypothesis that opioid receptors (OR), which are capable of producing cardiac protection in vivo, promote cardiac protection in cardiac myocytes in a caveolae-dependent manner. We determined protein expression and localization of delta-OR (DOR) using coimmunohistochemistry, caveolar fractionation, and immunoprecipitations. DOR colocalized in fractions with caveolin-3 (Cav-3), a structural component of caveolae in muscle cells, and could be immunoprecipitated by a Cav-3 antibody. Immunohistochemistry confirmed plasma membrane colocalization of DOR with Cav-3. Cardiac myocytes were subjected to simulated ischemia (2 h) or an ischemic preconditioning (IPC) protocol (10 min ischemia, 30 min recovery, 2 h ischemia) in the presence and absence of methyl-beta-cyclodextrin (MbetaCD, 2 mM), which binds cholesterol and disrupts caveolae. We also assessed the cardiac protective effects of SNC-121 (SNC), a selective DOR agonist, on cardiac myocytes with or without MbetaCD and MbetaCD preloaded with cholesterol. Ischemia, simulated by mineral oil layering to inhibit gas exchange, promoted cardiac myocyte cell death (trypan blue staining), a response blunted by SNC (37 +/- 3 vs. 59 +/- 3% dead cells in the presence and absence of 1 muM SNC, respectively, P protective effects of IPC or SNC, resulting in cell death comparable to that of the ischemic group. By contrast, SNC-induced protection was not abrogated in cells incubated with cholesterol-saturated MbetaCD, which maintained caveolae structure and function. These findings suggest a key role for caveolae, perhaps through enrichment of signaling molecules, in contributing to protection of cardiac myocytes from ischemic damage.

  9. Antinociceptive action of DBO 17 and DBO 11 in mice: two 3,8 diazabicyclo (3.2.1.) octane derivates with selective mu opioid receptor affinity.

    Science.gov (United States)

    Fadda, P; Barlocco, D; Tronci, S; Cignarella, G; Fratta, W

    1997-11-01

    Two 3,8 diazabicyclo (3.2.1.) octane derivates, namely DBO 17 and DBO 11, were studied for the opioid-like activity. In the rat brain membrane preparation binding studies, DBO 17 and DBO 11 showed a high affinity and selectivity for the mu opioid receptor (Ki's: 5.1 and 25 nM, respectively). DBO 17 and DBO 11 inhibited the nociceptive response in the hot-plate test of mice with ED50 values of 0.16 mg/kg and 0.44 mg/kg, respectively. The antinociceptive action of both DBO 17 and DBO 11 was blocked by naloxone. Tolerance to the antinociceptive action of DBO 17 and DBO 11 was present after 13 and 7 days of repeated treatment, respectively. Both DBO 17 and DBO 11 were ineffective in morphine-tolerant mice and vice versa. Chronic treatments (three times daily for seven consecutive days) of DBO 17 and DBO 11 induced a naloxone-precipitated withdrawal syndrome in DBO 17 treated mice similar to that in morphine treated mice, whereas in DBO 11 treated mice abstinence signs were virtually absent. These results indicate an interesting pharmacological profile that suggests these compounds as possible new candidates for the clinical treatment of pain.

  10. Opioid adjuvant strategy: improving opioid effectiveness.

    Science.gov (United States)

    Bihel, Frédéric

    2016-01-01

    Opioid analgesics continue to be the mainstay of pharmacologic treatment of moderate to severe pain. Many patients, particularly those suffering from chronic pain, require chronic high-dose analgesic therapy. Achieving clinical efficacy and tolerability of such treatment regimens is hampered by the appearance of opioid-induced side effects such as tolerance, hyperalgesia and withdrawal syndrome. Among the therapeutic options to improve the opioid effectiveness, this current review focuses on strategies combining opioids to other drugs that can modulate opioid-mediated effects. We will discuss about experimental evidences reported for several potential opioid adjuvants, including N-methyl-D-aspartate receptor antagonists, 5-HT7 agonists, sigma-1 antagonists, I2-R ligands, cholecystokinin antagonists, neuropeptide FF-R antagonists and toll-like receptor 4 antagonists.

  11. Fluoxetine reverses the behavioral despair induced by neurogenic stress in mice: role of N-methyl-d-aspartate and opioid receptors.

    Science.gov (United States)

    Haj-Mirzaian, Arya; Kordjazy, Nastaran; Ostadhadi, Sattar; Amiri, Shayan; Haj-Mirzaian, Arvin; Dehpour, AhmadReza

    2016-06-01

    Opioid and N-methyl-d-aspartate (NMDA) receptors mediate different effects of fluoxetine. We investigated whether opioid and NMDA receptors are involved in the protective effect of fluoxetine against the behavioral despair induced by acute physical stress in male mice. We used the forced swimming test (FST), tail suspension test (TST), and open-field test (OFT) for behavioral evaluation. We used fluoxetine, naltrexone (opioid receptor antagonist), MK-801 (NMDA receptor antagonist), morphine (opioid receptor agonist), and NMDA (NMDA receptor agonist). Acute foot-shock stress (FSS) significantly induced behavioral despair (depressive-like) and anxiety-like behaviors in tests. Fluoxetine (5 mg/kg) reversed the depressant-like effect of FSS, but it did not alter the locomotion and anxiety-like behavior in animals. Acute administration of subeffective doses of naltrexone (0.3 mg/kg) or MK-801 (0.01 mg/kg) potentiated the antidepressant-like effect of fluoxetine, while subeffective doses of morphine (1 mg/kg) and NMDA (75 mg/kg) abolished this effect of fluoxetine. Also, co-administration of subeffective doses of naltrexone (0.05 mg/kg) and MK-801 (0.003 mg/kg) with fluoxetine (1 mg/kg) induced a significant decrease in the immobility time in FST and TST. Our results showed that opioid and NMDA receptors (alone or in combination) are involved in the antidepressant-like effect of fluoxetine against physical stress.

  12. Risperidone treatment increases CB1 receptor binding in rat brain

    DEFF Research Database (Denmark)

    Secher, Anna; Husum, Henriette; Holst, Birgitte

    2010-01-01

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

  13. β-Endorphin via the Delta Opioid Receptor is a Major Factor in the Incubation of Cocaine Craving

    Science.gov (United States)

    Dikshtein, Yahav; Barnea, Royi; Kronfeld, Noam; Lax, Elad; Roth-Deri, Ilana; Friedman, Alexander; Gispan, Iris; Elharrar, Einat; Levy, Sarit; Ben-Tzion, Moshe; Yadid, Gal

    2013-01-01

    Cue-induced cocaine craving intensifies, or ‘incubates', during the first few weeks of abstinence and persists over extended periods of time. One important factor implicated in cocaine addiction is the endogenous opioid β-endorphin. In the present study, we examined the possible involvement of β-endorphin in the incubation of cocaine craving. Rats were trained to self-administer cocaine (0.75 mg/kg, 10 days, 6 h/day), followed by either a 1-day or a 30-day period of forced abstinence. Subsequent testing for cue-induced cocaine-seeking behavior (without cocaine reinforcement) was performed. Rats exposed to the drug-associated cue on day 1 of forced abstinence demonstrated minimal cue-induced cocaine-seeking behavior concurrently with a significant increase in β-endorphin release in the nucleus accumbens (NAc). Conversely, exposure to the cue on day 30 increased cocaine seeking, while β-endorphin levels remained unchanged. Intra-NAc infusion of an anti-β-endorphin antibody (4 μg) on day 1 increased cue-induced cocaine seeking, whereas infusion of a synthetic β-endorphin peptide (100 ng) on day 30 significantly decreased cue response. Both intra-NAc infusions of the δ opioid receptor antagonist naltrindole (1 μg) on day 1 and naltrindole together with β-endorphin on day 30 increased cue-induced cocaine-seeking behavior. Intra-NAc infusion of the μ opioid receptor antagonist CTAP (30 ng and 3 μg) had no behavioral effect. Altogether, these results demonstrate a novel role for β-endorphin and the δ opioid receptor in the development of the incubation of cocaine craving. PMID:23800967

  14. Regional differences in mu and kappa opioid receptor G-protein activation in brain in male and female prairie voles.

    Science.gov (United States)

    Martin, T J; Sexton, T; Kim, S A; Severino, A L; Peters, C M; Young, L J; Childers, S R

    2015-12-17

    Prairie voles are unusual mammals in that, like humans, they are capable of forming socially monogamous pair bonds, display biparental care, and engage in alloparental behaviors. Both mu and kappa opioid receptors are involved in behaviors that either establish and maintain, or result from pair bond formation in these animals. Mu and kappa opioid receptors both utilize inhibitory G-proteins in signal transduction mechanisms, however the efficacy by which these receptor subtypes stimulate G-protein signaling across the prairie vole neuraxis is not known. Utilizing [(35)S]GTPγS autoradiography, we characterized the efficacy of G-protein stimulation in coronal sections throughout male and female prairie vole brains by [D-Ala2,NMe-Phe4,Gly-ol5]-enkephalin (DAMGO) and U50,488H, selective mu and kappa opioid agonists, respectively. DAMGO stimulation was highest in the forebrain, similar to that found with other rodent species. U-50,488H produced greater stimulation in prairie voles than is typically seen in mice and rats, particularly in select forebrain areas. DAMGO produced higher stimulation in the core versus the shell of the nucleus accumbens (NAc) in females, while the distribution of U-50,488H stimulation was the opposite. There were no gender differences for U50,488H stimulation of G-protein activity across the regions examined, while DAMGO stimulation was greater in sections from females compared to those from males for NAc core, entopeduncular nucleus, and hippocampus. These data suggest that the kappa opioid system may be more sensitive to manipulation in prairie voles compared to mice and rats, and that female prairie voles may be more sensitive to mu agonists in select brain regions than males. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

  15. Analgesic Effects of Diluted Bee Venom Acupuncture Mediated by δ-Opioid and α2-Adrenergic Receptors in Osteoarthritic Rats.

    Science.gov (United States)

    Huh, Jeong-Eun; Seo, Byung-Kwan; Lee, Jung-Woo; Kim, Chanyoung; Park, Yeon-Cheol; Lee, Jae-Dong; Baek, Yong-Hyeon

    2017-06-23

    Context • Pain from osteoarthritis is associated with peripheral nociception and central pain processing. Given the unmet need for innovative, effective, and well-tolerated therapies, many patients, after looking for more satisfactory alternatives, decide to use complementary and alternative modalities. The analgesic mechanism of subcutaneous injections of diluted bee venom into an acupoint is thought to be part of an anti-inflammatory effect and the central modulation of pain processing. Objectives • Using the rat model of collagenase-induced osteoarthritis (CIOA), the study intended to investigate the analgesic effects of bee venom acupuncture (BVA) as they are related to the acupuncture points and dosage used and to determine whether the analgesic mechanisms of BVA for pain were mediated by opioid or adrenergic receptors. Design • Male Sprague-Dawley rats were randomly assigned to one of 19 groups, with n = 10 for each group. Setting • The study was conducted at the East-West Bone and Joint Research Institute at Kyung Hee University (Seoul, South Korea). Intervention • All rats were intra-articularly injected with collagenase solution in the left knee, followed by a booster injection performed 4 d after the first injection. For the groups receiving BVA treatments, the treatment was administered into the ST-36 acupoint, except for 1 group that received the treatment into a nonacupoint. Three BVA intervention groups received no pretreatment with agonists or antagonists; 1 of them received a dose of 1 mg/kg of bee venom into acupoint ST-36, 1 received a dose of 2 mg/kg into acupoint ST-36, and 1 received a dose of 1 mg/kg into a nonacupoint location. For the intervention groups receiving pretreatments, the opioid-receptor or adrenergic-receptor agonists or antagonists were injected 20 min before the 1-mg/kg BVA treatments. Outcome Measures • Changes in the rats' pain thresholds were assessed by evaluation of pain-related behavior, using a tail flick

  16. Involvement of Opioid System, TRPM8, and ASIC Receptors in Antinociceptive Effect of Arrabidaea brachypoda (DC Bureau

    Directory of Open Access Journals (Sweden)

    Vinícius Peixoto Rodrigues

    2017-11-01

    Full Text Available Arrabidaea brachypoda (DC Bureau is a medicinal plant found in Brazil. Known as “cipó-una”, it is popularly used as a natural therapeutic agent against pain and inflammation. This study evaluated the chemical composition and antinociceptive activity of the dichloromethane fraction from the roots of A. brachypoda (DEAB and its mechanism of action. The chemical composition was characterized by high-performance liquid chromatography, and this fraction is composed only of dimeric flavonoids. The antinociceptive effect was evaluated in formalin and hot plate tests after oral administration (10–100 mg/kg in male Swiss mice. We also investigated the involvement of TRPV1 (transient receptor potential vanilloid 1, TRPA1 (transient receptor potential ankyrin 1, TRPM8 (transient receptor potential melastatin 8, and ASIC (acid-sensing ion channel, as well as the opioidergic, glutamatergic, and supraspinal pathways. Moreover, the nociceptive response was reduced (30 mg/kg in the early and late phase of the formalin test. DEAB activity appears to involve the opioid system, TRPM8, and ASIC receptors, clearly showing that the DEAB alleviates acute pain in mice and suggesting the involvement of the TRPM8 and ASIC receptors and the opioid system in acute pain relief.

  17. Involvement of Opioid System, TRPM8, and ASIC Receptors in Antinociceptive Effect of Arrabidaea brachypoda (DC) Bureau.

    Science.gov (United States)

    Rodrigues, Vinícius Peixoto; Rocha, Cláudia Quintino da; Périco, Larissa Lucena; Santos, Raquel de Cássia Dos; Ohara, Rie; Nishijima, Catarine Massucato; Ferreira Queiroz, Emerson; Wolfender, Jean-Luc; Rocha, Lúcia Regina Machado da; Santos, Adair Roberto Soares; Vilegas, Wagner; Hiruma-Lima, Clélia Akiko

    2017-11-02

    Arrabidaea brachypoda (DC) Bureau is a medicinal plant found in Brazil. Known as "cipó-una", it is popularly used as a natural therapeutic agent against pain and inflammation. This study evaluated the chemical composition and antinociceptive activity of the dichloromethane fraction from the roots of A. brachypoda (DEAB) and its mechanism of action. The chemical composition was characterized by high-performance liquid chromatography, and this fraction is composed only of dimeric flavonoids. The antinociceptive effect was evaluated in formalin and hot plate tests after oral administration (10-100 mg/kg) in male Swiss mice. We also investigated the involvement of TRPV1 (transient receptor potential vanilloid 1), TRPA1 (transient receptor potential ankyrin 1), TRPM8 (transient receptor potential melastatin 8), and ASIC (acid-sensing ion channel), as well as the opioidergic, glutamatergic, and supraspinal pathways. Moreover, the nociceptive response was reduced (30 mg/kg) in the early and late phase of the formalin test. DEAB activity appears to involve the opioid system, TRPM8, and ASIC receptors, clearly showing that the DEAB alleviates acute pain in mice and suggesting the involvement of the TRPM8 and ASIC receptors and the opioid system in acute pain relief.

  18. Differential activation of G-proteins by μ-opioid receptor agonists

    Science.gov (United States)

    Saidak, Zuzana; Blake-Palmer, Katherine; Hay, Debbie L; Northup, John K; Glass, Michelle

    2006-01-01

    We investigated the ability of the activated μ-opioid receptor (MOR) to differentiate between myristoylated Gαi1 and GαoA type Gα proteins, and the maximal activity of a range of synthetic and endogenous agonists to activate each Gα protein. Membranes from HEK293 cells stably expressing transfected MOR were chaotrope extracted to denature endogenous G-proteins and reconstituted with specific purified G-proteins. The Gα subunits were generated in bacteria and were demonstrated to be recognised equivalently to bovine brain purified Gα protein by CB1 cannabinoid receptors. The ability of agonists to catalyse the MOR-dependent GDP/[35S]GTPγS exchange was then compared for Gαi1 and GαoA. Activation of MOR by DAMGO produced a high-affinity saturable interaction for GαoA (Km=20±1 nM) but a low-affinity interaction with Gαi1 (Km=116±12 nM). DAMGO, met-enkephalin and leucine-enkephalin displayed maximal Gα activation among the agonists evaluated. Endomorphins 1 and 2, methadone and β-endorphin activated both Gα to more than 75% of the maximal response, whereas fentanyl partially activated both G-proteins. Buprenorphine and morphine demonstrated a statistically significant difference between the maximal activities between Gαi1 and GαoA. Interestingly, DAMGO, morphine, endomorphins 1 and 2, displayed significant differences in the potencies for the activation of the two Gα. Differences in maximal activity and potency, for Gαi1 versus GαoA, are both indicative of agonist selective activation of G-proteins in response to MOR activation. These findings may provide a starting point for the design of drugs that demonstrate greater selectivity between these two G-proteins and therefore produce a more limited range of effects. PMID:16415903

  19. Oxytocin and Opioid Receptor Gene Polymorphisms Associated with Greeting Behavior in Dogs

    Directory of Open Access Journals (Sweden)

    Enikő Kubinyi

    2017-09-01

    Full Text Available Meeting humans is an everyday experience for most companion dogs, and their behavior in these situations and its genetic background is of major interest. Previous research in our laboratory reported that in German shepherd dogs the lack of G allele, and in Border collies the lack of A allele, of the oxytocin receptor gene (OXTR 19208A/G single nucleotide polymorphism (SNP was linked to increased friendliness, which suggests that although broad traits are affected by genetic variability, the specific links between alleles and behavioral variables might be breed-specific. In the current study, we found that Siberian huskies with the A allele approached a friendly unfamiliar woman less frequently in a greeting test, which indicates that certain polymorphisms are related to human directed behavior, but that the relationship patterns between polymorphisms and behavioral phenotypes differ between populations. This finding was further supported by our next investigation. According to primate studies, endogenous opioid peptide (e.g., endorphins receptor genes have also been implicated in social relationships. Therefore, we examined the rs21912990 of the OPRM1 gene. Firstly, we found that the allele frequencies of Siberian huskies and gray wolves were similar, but differed from that of Border collies and German shepherd dogs, which might reflect their genetic relationship. Secondly, we detected significant associations between the OPRM1 SNP and greeting behavior among German shepherd dogs and a trend in Border collies, but we could not detect an association in Siberian huskies. Although our results with OXTR and OPRM1 gene variants should be regarded as preliminary due to the relatively low sample size, they suggest that (1 OXTR and OPRM1 gene variants in dogs affect human-directed social behavior and (2 their effects differ between breeds.

  20. Oxytocin and Opioid Receptor Gene Polymorphisms Associated with Greeting Behavior in Dogs.

    Science.gov (United States)

    Kubinyi, Enikő; Bence, Melinda; Koller, Dora; Wan, Michele; Pergel, Eniko; Ronai, Zsolt; Sasvari-Szekely, Maria; Miklósi, Ádám

    2017-01-01

    Meeting humans is an everyday experience for most companion dogs, and their behavior in these situations and its genetic background is of major interest. Previous research in our laboratory reported that in German shepherd dogs the lack of G allele, and in Border collies the lack of A allele, of the oxytocin receptor gene (OXTR) 19208A/G single nucleotide polymorphism (SNP) was linked to increased friendliness, which suggests that although broad traits are affected by genetic variability, the specific links between alleles and behavioral variables might be breed-specific. In the current study, we found that Siberian huskies with the A allele approached a friendly unfamiliar woman less frequently in a greeting test, which indicates that certain polymorphisms are related to human directed behavior, but that the relationship patterns between polymorphisms and behavioral phenotypes differ between populations. This finding was further supported by our next investigation. According to primate studies, endogenous opioid peptide (e.g., endorphins) receptor genes have also been implicated in social relationships. Therefore, we examined the rs21912990 of the OPRM1 gene. Firstly, we found that the allele frequencies of Siberian huskies and gray wolves were similar, but differed from that of Border collies and German shepherd dogs, which might reflect their genetic relationship. Secondly, we detected significant associations between the OPRM1 SNP and greeting behavior among German shepherd dogs and a trend in Border collies, but we could not detect an association in Siberian huskies. Although our results with OXTR and OPRM1 gene variants should be regarded as preliminary due to the relatively low sample size, they suggest that (1) OXTR and OPRM1 gene variants in dogs affect human-directed social behavior and (2) their effects differ between breeds.

  1. Regulation of ventilation and oxygen consumption by delta- and mu-opioid receptor agonists.

    Science.gov (United States)

    Schaeffer, J I; Haddad, G G

    1985-09-01

    To study the effect of endorphins on metabolic rate and on the relationship between O2 consumption (VO2) and ventilation, we administered enkephalin analogues (relatively selective delta-receptor agonists) and a morphiceptin analogue (a highly selective mu-receptor agonist) intracisternally in nine unanesthetized chronically instrumented adult dogs. Both delta- and mu-agonists decreased VO2 by 40-60%. delta-Agonists induced a dose-dependent decrease in mean instantaneous minute ventilation (VT/TT) associated with periodic breathing. The decrease in VT/TT started and resolved prior to the decrease and returned to baseline of VO2, respectively. In contrast, the mu-agonists induced an increase in VT/TT associated with rapid shallow breathing. Arterial PCO2 increased and arterial PO2 decreased after both delta- and mu-agonists. Low doses of intracisternal naloxone (0.002-2.0 micrograms/kg) reversed the opioid effect on VT/TT but not on VO2; higher doses of naloxone (5-25 micrograms/kg) reversed both. Naloxone administered alone had no effect on VT/TT or VO2. These data suggest that 1) both delta- and mu-agonists induce alveolar hypoventilation despite a decrease in VO2, 2) this hypoventilation results from a decrease in VT/TT after delta-agonists but an increase in dead space ventilation after mu-agonists, and 3) endorphins do not modulate ventilation and metabolic rate tonically, but we speculate that they may do so in response to stressful stimulation.

  2. Dopamine D4 receptor counteracts morphine-induced changes in M opioid receptor signaling in the striosomes of the rat caudate putamen.

    OpenAIRE

    Rivera, Alicia; Valderrama-Carvajal, Alejandra; Roales-Buján, Ruth; Suárez-Boomgaard, Diana; Medina-Luque, José; Shumilov, Kirill; De-la-Calle-Martin, Adelaida

    2015-01-01

    Morphine is one of the most potent analgesic drugs used to relieve moderate to severe pain. After long-term use of morphine, neuroadaptive changes in the brain promotes tolerance, which result in a reduced sensitivity to most of its effects with attenuation of analgesic efficacy, and dependence, revealed by drug craving and physical or psychological manifestations of drug withdrawal. The mu opioid receptor (MOR) is critical, not only in mediating morphine analgesia, but also in addictive beha...

  3. Role of neurotensin and opioid receptors in the cardiorespiratory effects of [Ile⁹]PK20, a novel antinociceptive chimeric peptide.

    Science.gov (United States)

    Kaczyńska, Katarzyna; Szereda-Przestaszewska, Małgorzata; Kleczkowska, Patrycja; Lipkowski, Andrzej W

    2014-10-15

    Ile(9)PK20 is a novel hybrid of opioid-neurotensin peptides synthesized from the C-terminal hexapeptide of neurotensin and endomorphin-2 pharmacophore. This chimeric compound shows potent central and peripheral antinociceptive activity in experimental animals, however nothing is known about its influence on the respiratory and cardiovascular parameters. The present study was designed to determine the cardiorespiratory effects exerted by an intravenous injection (i.v.) of [Ile(9)]PK20. Share of the vagal afferentation and the contribution of NTS1 neurotensin and opioid receptors were tested. Intravenous injection of the hybrid at a dose of 100 μg/kg in the intact, anaesthetized rats provoked an increase in tidal volume preceded by a prompt short-lived decrease. Immediately after the end of injection brief acceleration of the respiratory rhythm appeared, and was ensued by the slowing down of breathing. Changes in respiration were concomitant with a bi-phasic response of the blood pressure: an immediate increase was followed by a sustained hypotension. Midcervical vagotomy eliminated the increase in tidal volume and respiratory rate responses. Antagonist of opioid receptors - naloxone hydrochloride eliminated only [Ile(9)]PK20-evoked decline in tidal volume response. Blockade of NTS1 receptors with an intravenous dose of SR 142,948, lessened the remaining cardiorespiratory effects. This study depicts that [Ile(9)]PK20 acting through neurotensin NTS1 receptors augments the tidal component of the breathing pattern and activates respiratory timing response through the vagal pathway. Blood pressure effects occur outside vagal afferentation and might result from activation of the central and peripheral vascular NTS1 receptors. In summary the respiratory effects of the hybrid appeared not to be profound, but they were accompanied with unfavourable prolonged hypotension. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Association between Opioid Receptor mu 1 (OPRM1 Gene Polymorphisms and Tobacco and Alcohol Consumption in a Spanish Population

    Directory of Open Access Journals (Sweden)

    Francesc Francès

    2015-04-01

    Full Text Available Evidence gained from animals and humans suggests that the encephalic opioid system might be involved in the development of drug addiction through its role in reward. Our aim is to assess the influence of genetic variations in the opioid receptor mu 1 on alcohol and tobacco consumption in a Spanish population. 763 unrelated individuals (465 women, 298 men aged 18-85 years were recruited between October 2011 and April 2012. Participants were requested to answer a 35-item questionnaire on tobacco and alcohol consumption, as well as to complete the AUDIT and Fagerström tests. Individuals were genotyped for three polymorphisms in the opioid receptor mu 1 (OPRM1 gene, using a TaqMan® protocol. In males, the rs10485057 polymorphism was associated with total pure ethanol intake and with the risk of being an alcohol consumer. Also, this polymorphism was significantly associated with higher Fagerström scores. Rs1799971 had a different influence on adaptive and maladaptive patterns of alcohol use. Despite the limited sample size, our study might enrich current knowledge on patterns of alcohol use, because it encompasses both extreme and adaptive phenotypes, providing thus a wider perspective on this subject.

  5. Crystal structure of mouse coronavirus receptor-binding domain complexed with its murine receptor

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Guiqing; Sun, Dawei; Rajashankar, Kanagalaghatta R.; Qian, Zhaohui; Holmes, Kathryn V.; Li, Fang (Cornell); (UMM-MED); (Colorado)

    2011-09-28

    Coronaviruses have evolved diverse mechanisms to recognize different receptors for their cross-species transmission and host-range expansion. Mouse hepatitis coronavirus (MHV) uses the N-terminal domain (NTD) of its spike protein as its receptor-binding domain. Here we present the crystal structure of MHV NTD complexed with its receptor murine carcinoembryonic antigen-related cell adhesion molecule 1a (mCEACAM1a). Unexpectedly, MHV NTD contains a core structure that has the same {beta}-sandwich fold as human galectins (S-lectins) and additional structural motifs that bind to the N-terminal Ig-like domain of mCEACAM1a. Despite its galectin fold, MHV NTD does not bind sugars, but instead binds mCEACAM1a through exclusive protein-protein interactions. Critical contacts at the interface have been confirmed by mutagenesis, providing a structural basis for viral and host specificities of coronavirus/CEACAM1 interactions. Sugar-binding assays reveal that galectin-like NTDs of some coronaviruses such as human coronavirus OC43 and bovine coronavirus bind sugars. Structural analysis and mutagenesis localize the sugar-binding site in coronavirus NTDs to be above the {beta}-sandwich core. We propose that coronavirus NTDs originated from a host galectin and retained sugar-binding functions in some contemporary coronaviruses, but evolved new structural features in MHV for mCEACAM1a binding.

  6. Agonist Binding to Chemosensory Receptors: A Systematic Bioinformatics Analysis

    Directory of Open Access Journals (Sweden)

    Fabrizio Fierro

    2017-09-01

    Full Text Available Human G-protein coupled receptors (hGPCRs constitute a large and highly pharmaceutically relevant membrane receptor superfamily. About half of the hGPCRs' family members are chemosensory receptors, involved in bitter taste and olfaction, along with a variety of other physiological processes. Hence these receptors constitute promising targets for pharmaceutical intervention. Molecular modeling has been so far the most important tool to get insights on agonist binding and receptor activation. Here we investigate both aspects by bioinformatics-based predictions across all bitter taste and odorant receptors for which site-directed mutagenesis data are available. First, we observe that state-of-the-art homology modeling combined with previously used docking procedures turned out to reproduce only a limited fraction of ligand/receptor interactions inferred by experiments. This is most probably caused by the low sequence identity with available structural templates, which limits the accuracy of the protein model and in particular of the side-chains' orientations. Methods which transcend the limited sampling of the conformational space of docking may improve the predictions. As an example corroborating this, we review here multi-scale simulations from our lab and show that, for the three complexes studied so far, they significantly enhance the predictive power of the computational approach. Second, our bioinformatics analysis provides support to previous claims that several residues, including those at positions 1.50, 2.50, and 7.52, are involved in receptor activation.

  7. Crystal structure of NL63 respiratory coronavirus receptor-binding domain complexed with its human receptor

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Kailang; Li, Weikai; Peng, Guiqing; Li, Fang; (Harvard-Med); (UMM-MED)

    2010-03-04

    NL63 coronavirus (NL63-CoV), a prevalent human respiratory virus, is the only group I coronavirus known to use angiotensin-converting enzyme 2 (ACE2) as its receptor. Incidentally, ACE2 is also used by group II SARS coronavirus (SARS-CoV). We investigated how different groups of coronaviruses recognize the same receptor, whereas homologous group I coronaviruses recognize different receptors. We determined the crystal structure of NL63-CoV spike protein receptor-binding domain (RBD) complexed with human ACE2. NL63-CoV RBD has a novel {beta}-sandwich core structure consisting of 2 layers of {beta}-sheets, presenting 3 discontinuous receptor-binding motifs (RBMs) to bind ACE2. NL63-CoV and SARS-CoV have no structural homology in RBD cores or RBMs; yet the 2 viruses recognize common ACE2 regions, largely because of a 'virus-binding hotspot' on ACE2. Among group I coronaviruses, RBD cores are conserved but RBMs are variable, explaining how these viruses recognize different receptors. These results provide a structural basis for understanding viral evolution and virus-receptor interactions.

  8. Interaction of chemokines with their receptors--from initial chemokine binding to receptor activating steps

    DEFF Research Database (Denmark)

    Thiele, Stefanie; Rosenkilde, Mette Marie

    2014-01-01

    and surveillance. Chemokines are a group of 8-12 kDa large peptides with a secondary structure consisting of a flexible N-terminus and a core-domain usually stabilized by two conserved disulfide bridges. They mainly interact with the extracellular domains of their cognate 7TM receptors. Affinityand activity......-contributing interactions are attributed to different domains and known to occur in two steps. Here, knowledge on chemokine and receptor domains involved in the first binding-step and the second activation-step is reviewed. A mechanism comprising at least two steps seems consistent; however, several intermediate...... interactions possibly occur, resulting in a multi-step process, as recently proposed for other 7TM receptors. Overall, the N-terminus of chemokine receptors is pivotal for binding of all chemokines. During receptor activation, differences between the two major chemokine subgroups occur, as CC-chemokines mainly...

  9. Basal μ-opioid receptor availability in the amygdala predicts the inhibition of pain-related brain activity during heterotopic noxious counter-stimulation.

    Science.gov (United States)

    Piché, Mathieu; Watanabe, Nobuhiro; Sakata, Muneyuki; Oda, Keiichi; Toyohara, Jun; Ishii, Kenji; Ishiwata, Kiichi; Hotta, Harumi

    2014-01-01

    The aim of this study was to investigate the association between the magnitude of anti-nociceptive effects induced by heterotopic noxious counter-stimulation (HNCS) and the basal μ-opioid receptor availability in the amygdala. In 8 healthy volunteers (4 females and 4 males), transcutaneous electrical stimulation was applied to the right sural nerve to produce the nociceptive flexion reflex (RIII-reflex), moderate pain, and scalp somatosensory evoked potentials (SEPs). Immersion of the left hand in cold water for 20min was used as HNCS. In a separate session, basal μ-opioid receptor availability was measured using positron emission tomography with the radiotracer [(11)C]carfentanil. HNCS produced a reduction of the P260 amplitude (pbasal μ-opioid receptor availability in the amygdala on the right (R(2)=0.55, p=0.03) with a similar trend on the left (R(2)=0.24, p=0.22). Besides, HNCS did not induce significant changes in pain and RIII-reflex amplitude (p>0.05). These results suggest that activation of μ-opioid receptors in the amygdala may contribute to the anti-nociceptive effects of HNCS. The lack of RIII-reflex modulation further suggests that μ-opioid receptor activation in the amygdala contributes to decrease pain-related brain activity through a cerebral mechanism independent of descending modulation. Copyright © 2014 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.

  10. Characterization of [3H] oxymorphone binding sites in mouse brain

    DEFF Research Database (Denmark)

    Yoo, Ji Hoon; Borsodi, Anna; Tóth, Géza

    2017-01-01

    Oxymorphone, one of oxycodone's metabolic products, is a potent opioid receptor agonist which is thought to contribute to the analgesic effect of its parent compound and may have high potential abuse liability. Nonetheless, the in vivo pharmacological binding profile of this drug is still unclear....... This study uses mice lacking mu (MOP), kappa (KOP) or delta (DOP) opioid receptors as well as mice lacking all three opioid receptors to provide full characterisation of oxymorphone binding sites in the brain. Saturation binding studies using [3H]oxymorphone revealed high affinity binding sites in mouse......]Oxymorphone binding was completely abolished across the majority of the brain regions in mice lacking MOP as well as in mice lacking all three opioid receptors. DOP and KOP knockout mice retained [3H]oxymorphone binding sites suggesting oxymorphone may not target DOP or KOP. These results confirm that the MOP...

  11. Receptor binding kinetics equations: Derivation using the Laplace transform method.

    Science.gov (United States)

    Hoare, Sam R J

    Measuring unlabeled ligand receptor binding kinetics is valuable in optimizing and understanding drug action. Unfortunately, deriving equations for estimating kinetic parameters is challenging because it involves calculus; integration can be a frustrating barrier to the pharmacologist seeking to measure simple rate parameters. Here, a well-known tool for simplifying the derivation, the Laplace transform, is applied to models of receptor-ligand interaction. The method transforms differential equations to a form in which simple algebra can be applied to solve for the variable of interest, for example the concentration of ligand-bound receptor. The goal is to provide instruction using familiar examples, to enable investigators familiar with handling equilibrium binding equations to derive kinetic equations for receptor-ligand interaction. First, the Laplace transform is used to derive the equations for association and dissociation of labeled ligand binding. Next, its use for unlabeled ligand kinetic equations is exemplified by a full derivation of the kinetics of competitive binding equation. Finally, new unlabeled ligand equations are derived using the Laplace transform. These equations incorporate a pre-incubation step with unlabeled or labeled ligand. Four equations for measuring unlabeled ligand kinetics were compared and the two new equations verified by comparison with numerical solution. Importantly, the equations have not been verified with experimental data because no such experiments are evident in the literature. Equations were formatted for use in the curve-fitting program GraphPad Prism 6.0 and fitted to simulated data. This description of the Laplace transform method will enable pharmacologists to derive kinetic equations for their model or experimental paradigm under study. Application of the transform will expand the set of equations available for the pharmacologist to measure unlabeled ligand binding kinetics, and for other time

  12. Dynorphin/Kappa Opioid Receptor Signaling in Preclinical Models of Alcohol, Drug, and Food Addiction.

    Science.gov (United States)

    Karkhanis, Anushree; Holleran, Katherine M; Jones, Sara R

    2017-01-01

    The dynorphin/kappa opioid receptor (KOR) system is implicated in the "dark side" of addiction, in which stress exacerbates maladaptive responses to drug and alcohol exposure. For example, acute stress and acute ethanol exposure result in an elevation in dynorphin, the KOR endogenous ligand. Activation of KORs results in modulation of several neurotransmitters; however, this chapter will focus on its regulatory effects on dopamine in mesolimbic areas. Specifically, KOR activation has an inhibitory effect on dopamine release, thereby influencing reward processing. Repeated stimulation of KORs, for example, via chronic drug and/or stress exposure, results in increased function of the dynorphin/KOR system. This augmentation in KOR function shifts the homeostatic balance in favor of an overall reduction in dopamine signaling via either by reducing dopamine release or by increasing dopamine transporter function. This chapter examines the effects of chronic ethanol exposure on KOR function and the downstream effects on dopamine transmission. Additionally, the impact of chronic cocaine exposure and its effects on KOR function will be explored. Further, KORs may also be involved in driving excessive consumption of food, contributing to the risk of developing obesity. While some studies have shown that KOR agonists reduce drug intake, other studies have shown that antagonists reduce addiction-like behaviors, demonstrating therapeutic potential. For example, KOR inhibition reduces ethanol intake in dependent animals, motivation to self-administer cocaine in chronic stress-exposed animals, and food consumption in obese animals. This chapter will delve into the mechanisms by which modulation of the dynorphin/KOR system may be therapeutic. © 2017 Elsevier Inc. All rights reserved.

  13. Effects of kappa opioid receptors on conditioned place aversion and social interaction in males and females

    Science.gov (United States)

    Robles, Cindee F.; McMackin, Marissa Z.; Campi, Katharine L.; Doig, Ian E.; Takahashi, Elizabeth Y.; Pride, Michael; Trainor, Brian C.

    2014-01-01

    The effects of kappa opioid receptors (KOR) on motivated behavior are well established based on studies in male rodents, but relatively little is known about the effects of KOR in females. We examined the effects of KOR activation on conditioned place aversion and social interaction in the California mouse (Peromyscus californicus). Important differences were observed in long-term (place aversion) and short-term (social interaction) effects. Females but not males treated with a 2.5mg/kg dose of U50,488 formed a place aversion, whereas males but not females formed a place aversion at the 10 mg/kg dose. In contrast the short term effects of different doses of U50,488 on social interaction behavior were similar in males and females. Acute injection with 10 mg/kg of U50,488 (but not lower doses) reduced social interaction behavior in both males and females. The effects of U50,488 on phosphorylated extracellular signal regulated kinase (pERK) and p38 MAP kinase were cell type and region specific. Higher doses of U50,488 increased the number of pERK neurons in the ventrolateral bed nucleus of the stria terminals in males but not females, a nucleus implicated in male aggressive behavior. In contrast, both males and females treated with U50,488 had more activated p38 cells in the nucleus accumbens shell. Unexpectedly, cells expressing activated p38 co-expressed Iba-1, a widely used microglia marker. In summary we found strong sex differences in the effects of U50,488 on place aversion whereas the acute effects on U50,488 induced similar behavioral effects in males and females. PMID:24445073

  14. Effects of pharmacological manipulation of the kappa opioid receptors on the aversive effects of nicotine.

    Science.gov (United States)

    Ward, Melissa; Norman, Haval; D'Souza, Manoranjan S

    2018-02-15

    Nicotine, an addictive component of tobacco smoke, produces both rewarding and aversive effects. Increasing the aversive effects of nicotine may help in promoting smoking cessation. However, neural targets mediating the aversive effects of nicotine have not been fully identified. In this study, we evaluated the role of kappa opioid receptors (KORs) in the aversive effects of nicotine (0.4 mg/kg, base; s.c.) using the nicotine-induced conditioned taste aversion (CTA) model in Wistar rats. The KORs were activated using the selective KOR agonist (±)U-50,488H (0, 0.03, 0.15 & 0.3mg/kg; s.c.) and inhibited using the KOR antagonist nor-binaltorphimine (nor-BNI; 0, 15 & 30mg/kg; s.c.) in separate groups of rats using a between-subjects design. Pretreatment with the KOR agonist (±)U-50,488H (0.3mg/kg) significantly increased aversion for the nicotine-associated solution. Additionally, (±)U-50,488H (0.3mg/kg) on its own induced aversion to the flavored solution associated with it even in the absence of nicotine, suggesting that the KOR agonist induced increase in nicotine-induced aversion was an additive effect. Interestingly, administration of the KOR antagonist nor-BNI (30mg/kg) prior to conditioning with nicotine/saline, but not after conditioning with nicotine/saline, attenuated nicotine-induced aversive effects compared to saline controls. Taken together, these data suggest a role for KORs in the aversive effects of nicotine. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Voluntary ethanol intake predicts κ-opioid receptor supersensitivity and regionally distinct dopaminergic adaptations in macaques.

    Science.gov (United States)

    Siciliano, Cody A; Calipari, Erin S; Cuzon Carlson, Verginia C; Helms, Christa M; Lovinger, David M; Grant, Kathleen A; Jones, Sara R

    2015-04-15

    The dopaminergic projections from the ventral midbrain to the striatum have long been implicated in mediating motivated behaviors and addiction. Previously it was demonstrated that κ-opioid receptor (KOR) signaling in the striatum plays a critical role in the increased reinforcing efficacy of ethanol following ethanol vapor exposure in rodent models. Although rodents have been used extensively to determine the neurochemical consequences of chronic ethanol exposure, establishing high levels of voluntary drinking in these models has proven difficult. Conversely, nonhuman primates exhibit similar intake and pattern to humans in regard to drinking. Here we examine the effects of chronic voluntary ethanol self-administration on dopamine neurotransmission and the ability of KORs to regulate dopamine release in the dorsolateral caudate (DLC) and nucleus accumbens (NAc) core. Using voltammetry in brain slices from cynomolgus macaques after 6 months of ad libitum ethanol drinking, we found increased KOR sensitivity in both the DLC and NAc. The magnitude of ethanol intake predicted increases in KOR sensitivity in the NAc core, but not the DLC. Additionally, ethanol drinking increased dopamine release and uptake in the NAc, but decreased both of these measures in the DLC. These data suggest that chronic daily drinking may result in regionally distinct disruptions of striatal outputs. In concert with previous reports showing increased KOR regulation of drinking behaviors induced by ethanol exposure, the strong relationship between KOR activity and voluntary ethanol intake observed here gives further support to the hypothesis that KORs may provide a promising pharmacotherapeutic target in the treatment of alcoholism. Copyright © 2015 the authors 0270-6474/15/355959-10$15.00/0.

  16. Cortical delta-opioid receptors potentiate K+ homeostasis during anoxia and oxygen-glucose deprivation.

    Science.gov (United States)

    Chao, Dongman; Donnelly, David F; Feng, Yin; Bazzy-Asaad, Alia; Xia, Ying

    2007-02-01

    Central neurons are extremely vulnerable to hypoxic/ischemic insult, which is a major cause of neurologic morbidity and mortality as a consequence of neuronal dysfunction and death. Our recent work has shown that delta-opioid receptor (DOR) is neuroprotective against hypoxic and excitotoxic stress, although the underlying mechanisms remain unclear. Because hypoxia/ischemia disrupts ionic homeostasis with an increase in extracellular K(+), which plays a role in neuronal death, we asked whether DOR activation preserves K(+) homeostasis during hypoxic/ischemic stress. To test this hypothesis, extracellular recordings with K(+)-sensitive microelectrodes were performed in mouse cortical slices under anoxia or oxygen-glucose deprivation (OGD). The main findings in this study are that (1) DOR activation with [D-Ala(2), D-Leu(5)]-enkephalinamide attenuated the anoxia- and OGD-induced increase in extracellular K(+) and decrease in DC potential in cortical slices; (2) DOR inhibition with naltrindole, a DOR antagonist, completely abolished the DOR-mediated prevention of increase in extracellular K(+) and decrease in DC potential; (3) inhibition of protein kinase A (PKA) with N-(2-[p-bromocinnamylamino]-ethyl)-5-isoquinolinesulfonamide dihydrochloride had no effect on the DOR protection; and (4) inhibition of protein kinase C (PKC) with chelerythrine chloride reduced the DOR protection, whereas the PKC activator (phorbol 12-myristate 13-acetate) mimicked the effect of DOR activation on K(+) homeostasis. These data suggest that activation of DOR protects the cortex against anoxia- or ODG-induced derangement of potassium homeostasis, and this protection occurs via a PKC-dependent and PKA-independent pathway. We conclude that an important aspect of DOR-mediated neuroprotection is its early action against derangement of K(+) homeostasis during anoxia or ischemia.

  17. Are peripheral opioid antagonists the solution to opioid side effects?

    LENUS (Irish Health Repository)

    Bates, John J

    2012-02-03

    Opioid medication is the mainstay of therapy for severe acute and chronic pain. Unfortunately, the side effects of these medications can affect patient comfort and safety, thus limiting their proven therapeutic potential. Whereas the main analgesic effects of opioids are centrally mediated, many of the common side effects are mediated via peripheral receptors. Novel peripheral opioid antagonists have been recently introduced that can block the peripheral actions of opioids without affecting centrally mediated analgesia. We review the clinical and experimental evidence of their efficacy in ameliorating opioid side effects and consider what further information might be useful in defining their role. IMPLICATIONS: The major analgesic effects of opioid medication are mediated within the brain and spinal cord. Many of the side effects of opioids are caused by activation of receptors outside these areas. Recently developed peripherally restricted opioid antagonists have the ability to block many opioid side effects without affecting analgesia.

  18. The development of [18F]cyclofoxy as a ligand for imaging opioid receptors in the CNS of conscious humans

    International Nuclear Information System (INIS)

    Rice, K.C.; Newman, A.H.; Ostrowski, N.L.; Cohen, R.M.; Pert, A.; Pert, C.B.; Burke, T.R. Jr.; McLellan, C.A.; Channing, M.A.; Finn, R.D.; Dunn, B.; Simpson, N.; Carson, R.W.; Larson, S.M.; Eckelman, W.C.; Bennett, J.M.; Kawai, R.; Sawada, Y.; Herscovitch, P.; Yolles, P.S.; Nordhal, T.; Gross, M.; Blasberg, R.

    1989-01-01

    Positron emission tomography (PET) is a unique, noninvasive technique applicable to real time visualization and quantitation of drug receptor occupancy in the brain of conscious humans. Such studies with the normal and abnormal human CNS can potentially provide insight into the biochemical basis of disease states and the effects of drug therapy. The (-)-enantiomer of cyclofoxy, a fluorinated, potent narcotic antagonist derived from naltrexone, has been developed at NIH as an agent for study of the opioid receptor-endorphin system using PET. The development and current status of this program is described, including application of the NIH Opiate Total Synthesis for production the pharmacologically inert (+)-[ 18 F]cyclofoxy required for quantitation of receptor occupancy

  19. Allelic association of the D2 dopamine receptor gene with receptor-binding characteristics in alcoholism

    International Nuclear Information System (INIS)

    Noble, E.P.; Blum, K.; Ritchie, T.; Montgomery, A.; Sheridan, P.J.

    1991-01-01

    The allelic association of the human D2 dopamine receptor gene with the binding characteristics of the D2 dopamine receptor was determined in 66 brains of alcoholic and non-alcoholic subjects. In a blinded experiment, DNA from the cerebral cortex was treated with the restriction endonuclease Taql and probed with a 1.5-kilobase (kb) digest of a clone (lambda hD2G1) of the human D2 dopamine receptor gene. The binding characteristics (Kd [binding affinity] and Bmax [number of binding sites]) of the D2 dopamine receptor were determined in the caudate nuclei of these brains using tritiated spiperone as the ligand. The adjusted Kd was significantly lower in alcoholic than in nonalcoholic subjects. In subjects with the A1 allele, in whom a high association with alcoholism was found, the Bmax was significantly reduced compared with the Bmax of subjects with the A2 allele. Moreover, a progressively reduced Bmax was found in subjects with A2/A2, A1/A2, and A1/A1 alleles, with subjects with A2/A2 having the highest mean values, and subjects with A1/A1, the lowest. The polymorphic pattern of the D2 dopamine receptor gene and its differential expression of receptors suggests the involvement of the dopaminergic system in conferring susceptibility to at least one subtype of severe alcoholism

  20. Progress on the application of ligand receptor binding assays in radiopharmaceuticals

    International Nuclear Information System (INIS)

    Zhou Xue; Qian Jinping; Kong Aiying; Zhu Lin

    2010-01-01

    Receptor binding assay is an important drug screening method, which can quickly and inexpensively study the interactions between the targeted receptor and the potential ligands in vitro and provide the information of the relative binding affinity of ligand-receptor. The imaging of many radiopharmaceuticals is based on highly selective radioligand-receptor binding. The technique plays an important role in the design and screening of receptor-targeting radiopharmaceuticals. (authors)

  1. Dopamine and μ-opioid receptor dysregulation in the brains of binge-eating female rats - possible relevance in the psychopathology and treatment of binge-eating disorder.

    Science.gov (United States)

    Heal, David J; Hallam, Michelle; Prow, Michael; Gosden, Jane; Cheetham, Sharon; Choi, Yong K; Tarazi, Frank; Hutson, Peter

    2017-06-01

    Adult, female rats given irregular, limited access to chocolate develop binge-eating behaviour with normal bodyweight and compulsive/perseverative and impulsive behaviours similar to those in binge-eating disorder. We investigated whether (a) dysregulated central nervous system dopaminergic and opioidergic systems are part of the psychopathology of binge-eating and (b) these neurotransmitter systems may mediate the actions of drugs ameliorating binge-eating disorder psychopathology. Binge-eating produced a 39% reduction of striatal D 1 receptors with 22% and 23% reductions in medial and lateral caudate putamen and a 22% increase of striatal μ-opioid receptors. There was no change in D 1 receptor density in nucleus accumbens, medial prefrontal cortex or dorsolateral frontal cortex, striatal D 2 receptors and dopamine reuptake transporter sites, or μ-opioid receptors in frontal cortex. There were no changes in ligand affinities. The concentrations of monoamines, metabolites and estimates of dopamine (dopamine/dihydroxyphenylacetic acid ratio) and serotonin/5-hydroxyindolacetic acid ratio turnover rates were unchanged in striatum and frontal cortex. However, turnover of dopamine and serotonin in the hypothalamus was increased ~20% and ~15%, respectively. Striatal transmission via D 1 receptors is decreased in binge-eating rats while μ-opioid receptor signalling may be increased. These changes are consistent with the attenuation of binge-eating by lisdexamfetamine, which increases catecholaminergic neurotransmission, and nalmefene, a μ-opioid antagonist.

  2. DMPD: LPS-binding proteins and receptors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 9665271 LPS-binding proteins and receptors. Fenton MJ, Golenbock DT. J Leukoc Biol.... 1998 Jul;64(1):25-32. (.png) (.svg) (.html) (.csml) Show LPS-binding proteins and receptors. PubmedID 9665271 Title LPS-binding prot...eins and receptors. Authors Fenton MJ, Golenbock DT. Publication J Leukoc Biol. 199

  3. Beta 2-adrenergic receptors on eosinophils. Binding and functional studies

    International Nuclear Information System (INIS)

    Yukawa, T.; Ukena, D.; Kroegel, C.; Chanez, P.; Dent, G.; Chung, K.F.; Barnes, P.J.

    1990-01-01

    We have studied the binding characteristics and functional effects of beta-adrenoceptors on human and guinea pig eosinophils. We determined the binding of the beta-antagonist radioligand [125I]pindolol (IPIN) to intact eosinophils obtained from the peritoneal cavity of guinea pigs and from blood of patients with eosinophilia. Specific binding was saturable, and Scatchard analysis showed a single binding site with a dissociation constant (Kd) of 24.6 pM and maximal number of binding sites (Bmax) of 7,166 per cell. ICI 118,551, a beta 2-selective antagonist, inhibited IPIN binding with a Ki value of 0.28 nM and was approximately 5,000-fold more effective than the beta 1-selective antagonist, atenolol. Isoproterenol increased cAMP levels about 5.5-fold above basal levels (EC50 = 25 microM); albuterol, a beta 2-agonist, behaved as a partial agonist with a maximal stimulation of 80%. Binding to human eosinophils gave similar results with a Kd of 25.3 pM and a Bmax corresponding to 4,333 sites per cell. Incubation of both human and guinea pig eosinophils with opsonized zymosan (2 mg/ml) or with phorbol myristate acetate (PMA) (10(-8) and 10(-6) M) resulted in superoxide anion generation and the release of eosinophil peroxidase; albuterol (10(-7) to 10(-5) M) had no inhibitory effect on the release of these products. Thus, eosinophils from patients with eosinophilia and from the peritoneal cavity of guinea pigs possess beta-receptors of the beta 2-subtype that are coupled to adenylate cyclase; however, these receptors do not modulate oxidative metabolism or degranulation. The possible therapeutic consequences of these observations to asthma are discussed

  4. Five of Five VHHs Neutralizing Poliovirus Bind the Receptor-Binding Site.

    Science.gov (United States)

    Strauss, Mike; Schotte, Lise; Thys, Bert; Filman, David J; Hogle, James M

    2016-01-13

    Nanobodies, or VHHs, that recognize poliovirus type 1 have previously been selected and characterized as candidates for antiviral agents or reagents for standardization of vaccine quality control. In this study, we present high-resolution cryo-electron microscopy reconstructions of poliovirus with five neutralizing VHHs. All VHHs bind the capsid in the canyon at sites that extensively overlap the poliovirus receptor-binding site. In contrast, the interaction involves a unique (and surprisingly extensive) surface for each of the five VHHs. Five regions of the capsid were found to participate in binding with all five VHHs. Four of these five regions are known to alter during the expansion of the capsid associated with viral entry. Interestingly, binding of one of the VHHs, PVSS21E, resulted in significant changes of the capsid structure and thus seems to trap the virus in an early stage of expansion. We describe the cryo-electron microscopy structures of complexes of five neutralizing VHHs with the Mahoney strain of type 1 poliovirus at resolutions ranging from 3.8 to 6.3Å. All five VHHs bind deep in the virus canyon at similar sites that overlap extensively with the binding site for the receptor (CD155). The binding surfaces on the VHHs are surprisingly extensive, but despite the use of similar binding surfaces on the virus, the binding surface on the VHHs is unique for each VHH. In four of the five complexes, the virus remains essentially unchanged, but for the fifth there are significant changes reminiscent of but smaller in magnitude than the changes associated with cell entry, suggesting that this VHH traps the virus in a previously undescribed early intermediate state. The neutralizing mechanisms of the VHHs and their potential use as quality control agents for the end game of poliovirus eradication are discussed. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  5. Binding Mode of Insulin Receptor and Agonist Peptide

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Insulin is a protein hormone secreted by pancreatic β cells. One of its main functions is to keep the balance of glucose inside the body by regulating the absorption and metabolism of glucose in the periphery tissue, as well as the production and storage of hepatic glycogen. The insulin receptor is a transmembrane glycoprotein in which two α subunits with a molecular weight of 135 kD and twoβ subunits with a molecular weight of 95 kD are joined by a disulfide bond to form a β-α-α-β structure. The extracellular α subunit, especially, its three domains near the N-terminal are partially responsible for signal transduction or ligand-binding, as indicated by the experiments. The extracellular α subunits are involved in binding the ligands. The experimental results indicate that the three domains of the N-terminal of the α subunits are the main determinative parts of the insulin receptor to bind the insulin or mimetic peptide.We employed the extracellular domain (PDBID: 1IGR) of the insulin-like growth factor-1 receptor (IGF-1 R ) as the template to simulate and optimize the spatial structures of the three domains in the extracellular domain of the insulin receptor, which includes 468 residues. The work was accomplished by making use of the homology program in the Insight Ⅱ package on an Origin3800 server. The docking calculations of the insulin receptor obtained by homology with hexapeptides were carried out by means of the program Affinity. The analysis indicated that there were hydrogen bonding, and electrostatic and hydrophobic effects in the docking complex of the insulin receptor with hexapeptides.Moreover, we described the spatial orientation of a mimetic peptide with agonist activity in the docking complex. We obtained a rough model of binding of DLAPSQ or STIVYS with the insulin receptor, which provides the powerful theoretical support for designing the minimal insulin mimetic peptide with agonist activity, making it possible to develop oral small

  6. Changes in mu-opioid receptor expression and function in the mesolimbic system after long-term access to a palatable diet.

    Science.gov (United States)

    Pitman, Kimberley A; Borgland, Stephanie L

    2015-10-01

    The incidence of obesity in both adults and children is rising. In order to develop effective treatments for obesity, it is important to understand how diet can induce changes in the brain that could promote excessive intake of high-calorie foods and alter the efficacy of therapeutic targets. The mu-opioid receptor is involved in regulating the motivation for and hedonic reaction to food. Here, we review the literature examining changes in the expression and function of mu-opioid receptors in the mesolimbic system of rodents after extended access to a high-fat diet. We also review how maternal diet can induce long-term changes in the expression or function of mu-opioid receptors in the mesolimbic system of offspring. Understanding the behavioural and therapeutic implications of these changes requires further study. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Glycosylation of immunoglobulin A influences its receptor binding.

    Science.gov (United States)

    Basset, C; Devauchelle, V; Durand, V; Jamin, C; Pennec, Y L; Youinou, P; Dueymes, M

    1999-12-01

    Immunoglobulin A (IgA), which is heavily glycosylated, interacts with a variety of receptors, e.g. the asialoglycoprotein receptor (ASGP-R), which binds terminal galactose residues, and the Fcalpha receptor (FcalphaRI). It has thus been proposed that elevated serum levels of IgA in primary Sjögren's syndrome (pSS) are caused by its defective clearance. To test this hypothesis, we developed a method (based on sialyl transferases eluted from a hepatoma cell line) to increase the amount of sialic acid (SA) on IgA, and used a battery of IgA1- and IgA2-specific glycosidases to reduce this amount. Binding of IgA1 and IgA2 to ASGP-R and FcalphaRI was found to be sugar dependent because oversialylated IgA bound less than native or desialylated IgA. However, individual sugars did not play a direct role in this binding. Given that IgA are oversialylated in pSS, defective clearance of IgA may indeed be ascribed to an excess of SA in IgA1 and IgA2.

  8. Brain catalase activity inhibition as well as opioid receptor antagonism increases ethanol-induced HPA axis activation.

    Science.gov (United States)

    Pastor, Raúl; Sanchis-Segura, Carles; Aragon, Carlos M G

    2004-12-01

    Growing evidence indicates that brain catalase activity is involved in the psychopharmacological actions of ethanol. Recent data suggest that participation of this enzymatic system in some ethanol effects could be mediated by the endogenous opioid system. The present study assessed whether brain catalase has a role in ethanol-induced activation of the HPA axis, a neuroendocrine system modulated by the endogenous opioid neurotransmission. Swiss male mice received an intraperitoneal injection of the catalase inhibitor 3-amino-1,2,4-triazole (AT; 0-1 g/kg), and 0 to 20 hr after this administration, animals received an ethanol (0-4 g/kg; intraperitoneally) challenge. Thirty, 60, or 120 min after ethanol administration, plasma corticosterone levels were determined immunoenzymatically. In addition, we tested the effects of 45 mg/kg of cyanamide (another catalase inhibitor) and 0 to 2 mg/kg of naltrexone (nonselective opioid receptor antagonist) on ethanol-induced enhancement in plasma corticosterone values. The present study revealed that AT boosts ethanol-induced increase in plasma corticosterone levels in a dose- and time-dependent manner. However, it did not affect corticosterone values when measured after administration of saline, cocaine (4 mg/kg, intraperitoneally), or morphine (30 mg/kg, intraperitoneally). The catalase inhibitor cyanamide (45 mg/kg, intraperitoneally) also increased ethanol-related plasma corticosterone levels. These effects of AT and cyanamide on ethanol-induced corticosterone values were observed under treatment conditions that decreased significantly brain catalase activity. Indeed, a significant correlation between effects of catalase manipulations on both variables was found. Finally, we found that the administration of naltrexone enhanced the levels of plasma corticosterone after the administration of saline or ethanol. This study shows that the inhibition of brain catalase increases ethanol-induced plasma corticosterone levels. Results are

  9. Long-term antagonism of κ opioid receptors prevents escalation of and increased motivation for heroin intake.

    Science.gov (United States)

    Schlosburg, Joel E; Whitfield, Timothy W; Park, Paula E; Crawford, Elena F; George, Olivier; Vendruscolo, Leandro F; Koob, George F

    2013-12-04

    The abuse of opioid drugs, both illicit and prescription, is a persistent problem in the United States, accounting for >1.2 million users who require treatment each year. Current treatments rely on suppressing immediate withdrawal symptoms and replacing illicit drug use with long-acting opiate drugs. However, the mechanisms that lead to preventing opiate dependence are still poorly understood. We hypothesized that κ opioid receptor (KOR) activation during chronic opioid intake contributes to negative affective states associated with withdrawal and the motivation to take increasing amounts of heroin. Using a 12 h long-access model of heroin self-administration, rats showed escalation of heroin intake over several weeks. This was prevented by a single high dose (30 mg/kg) of the long-acting KOR antagonist norbinaltorphimine (nor-BNI), paralleled by reduced motivation to respond for heroin on a progressive-ratio schedule of reinforcement, a measure of compulsive-like responding. Systemic nor-BNI also significantly decreased heroin withdrawal-associated anxiety-like behavior. Immunohistochemical analysis showed prodynorphin content increased in the nucleus accumbens core in all heroin-exposed rats, but selectively increased in the nucleus accumbens shell in long-access rats. Local infusion of nor-BNI (4 μg/side) into accumbens core altered the initial intake of heroin but not the rate of escalation, while local injection into accumbens shell selectively suppressed increases in heroin intake over time without altering initial intake. These data suggest that dynorphin activity in the nucleus accumbens mediates the increasing motivation for heroin taking and compulsive-like responding for heroin, suggesting that KOR antagonists may be promising targets for the treatment of opioid addiction.

  10. Preparation and Evaluation at the Delta Opioid Receptor of a Series of Linear Leu-Enkephalin Analogues Obtained by Systematic Replacement of the Amides

    Science.gov (United States)

    2013-01-01

    Leu-enkephalin analogues, in which the amide bonds were sequentially and systematically replaced either by ester or N-methyl amide bonds, were prepared using classical organic chemistry as well as solid phase peptide synthesis (SPPS). The peptidomimetics were characterized using competition binding, ERK1/2 phosphorylation, receptor internalization, and contractility assays to evaluate their pharmacological profile over the delta opioid receptor (DOPr). The lipophilicity (LogD7.4) and plasma stability of the active analogues were also measured. Our results revealed that the last amide bond can be successfully replaced by either an ester or an N-methyl amide bond without significantly decreasing the biological activity of the corresponding analogues when compared to Leu-enkephalin. The peptidomimetics with an N-methyl amide function between residues Phe and Leu were found to be more lipophilic and more stable than Leu-enkephalin. Findings from the present study further revealed that the hydrogen-bond donor properties of the fourth amide of Leu-enkephalin are not important for its biological activity on DOPr. Our results show that the systematic replacement of amide bonds by isosteric functions represents an efficient way to design and synthesize novel peptide analogues with enhanced stability. Our findings further suggest that such a strategy can also be useful to study the biological roles of amide bonds. PMID:23650868

  11. Synthesis, modelling, and mu-opioid receptor affinity of N-3(9)-arylpropenyl-N-9(3)-propionyl-3,9-diazabicycl.

    Science.gov (United States)

    Pinna, G A; Murineddu, G; Curzu, M M; Villa, S; Vianello, P; Borea, P A; Gessi, S; Toma, L; Colombo, D; Cignarella, G

    2000-08-01

    A series of N-3-arylpropenyl-N-9-propionyl-3,9-diazabicyclo[3.3.1]nonanes (1a-g) and of reverted N-3-propionyl-N-9-arylpropenyl isomers (2a-g), as homologues of the previously reported analgesic 3,8-diazabicyclo[3.2.1]octanes (I-II), were synthesized and evaluated for the binding affinity towards opioid receptor subtypes mu, delta and kappa. Compounds 1a-g and 2a-g exhibited a strong selective mu-affinity with Ki values in the nanomolar range, which favourably compared with those of I and II. In addition, contrary to the trend observed for DBO-I, II, the mu-affinity of series 2 is markedly higher than that of the isomeric series 1. This aspect was discussed on the basis of the conformational studies performed on DBN which allowed hypotheses on the mode of interaction of these compounds with the mu receptor.

  12. (/sup 3/H)diprenorphine binding to kappa-sites in guinea-pig and rat brain: Evidence for apparent heterogeneity

    Energy Technology Data Exchange (ETDEWEB)

    Wood, M.S.; Traynor, J.R.

    1989-07-01

    The binding of the unselective opioid antagonist (/sup 3/H)diprenorphine to homogenates prepared from rat brain and from guinea-pig brain and cerebellum has been studied in HEPES buffer containing 10 mM Mg2+ ions. Sequential displacement of bound (/sup 3/H)diprenorphine by ligands with selectivity for mu-, delta-, and kappa-opioid receptors uncovers the multiple components of binding. In the presence of cold ligands that occupy all mu-, delta-, and kappa-sites, opioid binding still remains. This binding represents 20% of total specific sites and is displaced by naloxone. The nature of these undefined opioid binding sites is discussed.

  13. Cholinergic receptor binding in the frontal cortex of suicide victims

    International Nuclear Information System (INIS)

    Stanley, M.

    1986-01-01

    Because there is a high incidence of individuals diagnosed as having an affective disorder who subsequently commit suicide, the author thought it would be of interest to determine QNB binding in the brains of a large sample of suicide victims, and to compare the findings with a well-matched control group. Brain samples were obtained at autopsy from 22 suicide victims and 22 controls. Frontal cortex samples were diseected, frozen, and stored until assayed. Samples of tissue homogenate were incubated in duplicate with 10 concentrations of tritium-QNB. Specific binding was determined with and without atropine. The results confirmed previous studies in which no changes were noted in suicide versus control brains. While the findings neither disprove nor support the cholinergic hypothesis of depression, they do suggest that the neurochemical basis for the in vivo observations of increased responsivity of depressed individuals to muscarinic cholinergic agents might not involve changes in receptors estimated by QNB binding

  14. Intrauterine growth restriction modifies the hedonic response to sweet taste in newborn pups - Role of the accumbal μ-opioid receptors.

    Science.gov (United States)

    Laureano, D P; Dalle Molle, R; Alves, M B; Luft, C; Desai, M; Ross, M G; Silveira, P P

    2016-05-13

    Intrauterine growth restriction (IUGR) is associated with increased preference for palatable foods. The hedonic response to sweet taste, modulated by the nucleus accumbens μ-opioid-receptors, may be involved. We investigated hedonic responses and receptor levels in IUGR and Control animals. From pregnancy day 10, Sprague-Dawley dams received either an ad libitum (Control), or a 50% food restricted (FR) diet. At birth, pups were cross-fostered, and nursed by Adlib fed dams. The hedonic response was evaluated at 1 day after birth and at 90 days of life, by giving sucrose solution or water and analyzing the hedonic facial responses (within 60s). Control pups exposed either to water or sucrose resolved their hedonic responses after 16 and 18s, respectively, while FR hedonic responses to sucrose persisted over 20s. FR pups had deceased phospho-μ-opioid-receptor (p=0.009) and reduced phosphor:total mu opioid receptor ratio compared to controls pups (p=0.003). In adults, there was an interaction between group and solution at the end of the evaluation (p=0.044): Control decreased the response after sucrose solution, FR did not change over time. There were no differences in phosphorylation of μ-opioid-receptor in adults. These results demonstrate IUGR newborn rats exhibit alterations in hedonic response accompanied by a decrease in μ-opioid-receptor phosphorylation, though these alterations do not persist at 3 months of age. Opioid system alterations in early life may contribute to the development of preference for highly palatable foods and contribute to rapid weight gain and obesity in IUGR offspring. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  15. Delta opioid receptor on equine sperm cells: subcellular localization and involvement in sperm motility analyzed by computer assisted sperm analyzer (CASA

    Directory of Open Access Journals (Sweden)

    Lacalandra Giovanni M

    2010-06-01

    Full Text Available Abstract Background Opioid receptors and endogenous opioid peptides act not only in the control of nociceptive pathways, indeed several reports demonstrate the effects of opiates on sperm cell motility and morphology suggesting the importance of these receptors in the modulation of reproduction in mammals. In this study we investigated the expression of delta opioid receptors on equine spermatozoa by western blot/indirect immunofluorescence and its relationship with sperm cell physiology. Methods We analyzed viability, motility, capacitation, acrosome reaction and mitochondrial activity in the presence of naltrindole and DPDPE by means of a computer assisted sperm analyzer and a fluorescent confocal microscope. The evaluation of viability, capacitation and acrosome reaction was carried out by the double CTC/Hoechst staining, whereas mitochondrial activity was assessed by means of MitoTracker Orange dye. Results We showed that in equine sperm cells, delta opioid receptor is expressed as a doublet of 65 and 50 kDa molecular mass and is localized in the mid piece of tail; we also demonstrated that naltrindole, a delta opioid receptor antagonist, could be utilized in modulating several physiological parameters of the equine spermatozoon in a dose-dependent way. We also found that low concentrations of the antagonist increase sperm motility whereas high concentrations show the opposite effect. Moreover low concentrations hamper capacitation, acrosome reaction and viability even if the percentage of cells with active mitochondria seems to be increased; the opposite effect is exerted at high concentrations. We have also observed that the delta opioid receptor agonist DPDPE is scarcely involved in affecting the same parameters at the employed concentrations. Conclusions The results described in this paper add new important details in the comprehension of the mammalian sperm physiology and suggest new insights for improving reproduction and for

  16. Safety and efficacy of an oxycodone vaccine: Addressing some of the unique considerations posed by opioid abuse.

    Directory of Open Access Journals (Sweden)

    M D Raleigh

    Full Text Available Among vaccines aimed at treating substance use disorders, those targeting opioids present several unique medication development challenges. 1 Opioid overdose is a common complication of abuse, so it is desirable for an opioid vaccine to block the toxic as well as the addictive effects of opioids. 2 It is important that an opioid vaccine not interfere with the action of opioid antagonists used to reverse opioid overdose or treat addiction. 3 Some opioids are immunosuppressive and chronic ongoing opioid use could interfere with vaccine immunogenicity. 4 Although antibody-bound oxycodone is unable to enter the brain because of its size, it might still be able to activate peripheral opioid receptors. To assess vaccine impact on opioid toxicity, rats vaccinated with oxycodone conjugated to keyhole limpet hemocyanin subunit dimer (OXY-dKLH adsorbed to alum or controls vaccinated with dKLH were compared with regard to oxycodone-induced hotplate analgesia and oxycodone-induced respiratory depression and bradycardia. Vaccination shifted the dose-response curves to the right, representing protection, for each of these endpoints. Naloxone was equally effective in both OXY-dKLH and control groups, providing complete and rapid reversal of respiratory depression. The administration of a long-acting naltrexone formulation during vaccination did not impair vaccine immunogenicity in mice. Similarly, serum anti-oxycodone antibody titers were not altered by continuous morphine infusion during vaccination compared to opioid-naïve controls. Competitive ELISA assay showed negligible or low affinity of immune antiserum for endogenous opioids or opioid antagonists. In vitro receptor binding assays showed that antibody-bound oxycodone does not activate mu opioid receptors. These data support further study of OXY-dKLH as a potential treatment for oxycodone abuse and suggest that vaccination might also reduce the severity of oxycodone overdose.

  17. Anti-nociceptive effect of patchouli alcohol: Involving attenuation of cyclooxygenase 2 and modulation of mu-opioid receptor.

    Science.gov (United States)

    Yu, Xuan; Wang, Xin-Pei; Yan, Xiao-Jin; Jiang, Jing-Fei; Lei, Fan; Xing, Dong-Ming; Guo, Yue-Ying; Du, Li-Jun

    2017-08-09

    To explore the anti-nociceptive effect of patchouli alcohol (PA), the essential oil isolated from Pogostemon cablin (Blanco) Bent, and determine the mechanism in molecular levels. The acetic acid-induced writhing test and formalin-induced plantar injection test in mice were employed to confifirm the effect in vivo. Intracellular calcium ion was imaged to verify PA on mu-opioid receptor (MOR). Cyclooxygenase 2 (COX2) and MOR of mouse brain were expressed for determination of PA's target. Cellular experiments were carried out to find out COX2 and MOR expression induced by PA. PA significantly reduced latency period of visceral pain and writhing induced by acetic acid saline solution (Peffect of PA. A decrease in the intracellular calcium level (Peffect. PA showed the characters of enhancing the MOR expression and reducing the intracellular calcium ion similar to opioid effect. Both COX2 and MOR are involved in the mechanism of PA's anti-nociceptive effect, and the up-regulation of the receptor expression and the inhibition of intracellular calcium are a new perspective to PA's effect on MOR.

  18. Development of N-substituted quinolinimides, as potential PET tracers for the visualisation of δ-opioid receptors

    International Nuclear Information System (INIS)

    Bourdier, Th.

    2005-12-01

    In order to develop radiotracers for in vivo studies of δ-opioid receptors by Positron Emission Tomography (PET) or Single Photon Emission computed Tomography (SPECT), we undertook the synthesis of halogenated analogues (chlorinated and brominated) of compound 12. These analogues were prepared by a convergent synthesis and from these novel structures a halogen exchange reaction has been performed to complete this series. These molecules were tested to determine their in vitro affinity and selectivity toward δ opioid receptors. The compounds 12 and 15 were labelled with carbon-11. The radiosynthesis of compound 12, in weak radioactivity chemistry, was performed first by the Stille reaction and second by a new methodology based on the transfer reaction of [ 11 C]-methyl group. This new methodology used a mono-organotin compound prepared by addition of [ 11 C]-iodomethane onto Lappert's stannylene. The compound [ 11 C]-12 was obtained with 60 and 10% radiochemical yield respectively. In order to produce higher radioactivity quantities, the Stille reaction was automated. The compounds [ 11 C]-12 and [ 11 C]-15 were obtained in 40 minutes with a specific radioactivity ranging from 322 to 747 mCi/μmol. (author)

  19. Local analgesic effect of tramadol is not mediated by opioid receptors in early postoperative pain in rats

    Directory of Open Access Journals (Sweden)

    Angela Maria Sousa

    2015-05-01

    Full Text Available Background and objectives: Tramadol is known as a central acting analgesic drug, used for the treatment of moderate to severe pain. Local analgesic effect has been demonstrated, in part due to local anesthetic-like effect, but other mechanisms remain unclear. The role of peripheral opioid receptors in the local analgesic effect is not known. In this study, we examined role of peripheral opioid receptors in the local analgesic effect of tramadol in the plantar incision model. Methods: Young male Wistar rats were divided into seven groups: control, intraplantar tramadol, intravenous tramadol, intravenous naloxone-intraplantar tramadol, intraplantar naloxone-intraplantar tramadol, intravenous naloxone-intravenous tramadol, and intravenous naloxone. After receiving the assigned drugs (tramadol 5 mg, naloxone 200 μg or 0.9% NaCl, rats were submitted to plantar incision, and withdrawal thresholds after mechanical stimuli with von Frey filaments were assessed at baseline, 10, 15, 30, 45 and 60 min after incision. Results: Plantar incision led to marked mechanical hyperalgesia during the whole period of observation in the control group, no mechanical hyperalgesia were observed in intraplantar tramadol group, intraplantar naloxone-intraplantar tramadol group and intravenous naloxone-intraplantar tramadol. In the intravenous tramadol group a late increase in withdrawal thresholds (after 45 min was observed, the intravenous naloxone-intravenous tramadol group and intravenous naloxone remained hyperalgesic during the whole period. Conclusions: Tramadol presented an early local analgesic effect decreasing mechanical hyperalgesia induced by plantar incision. This analgesic effect was not mediated by peripheral opioid receptors. Resumo: Justificativa e objetivos: Tramadol é conhecido como um fármaco analgésico de ação central, usado para o tratamento de dor moderada a grave. O efeito analgésico local foi demonstrado, em parte devido ao efeito

  20. An autoradiographic map of (3H)diprenorphine binding in rat brain: effects of social interaction

    International Nuclear Information System (INIS)

    Panksepp, J.; Bishop, P.

    1981-01-01

    (3H)Diprenorphine binding was analyzed autoradiographically in the brains of 33 day old rat pups. A photographic atlas of diprenorphine binding in the coronal plane is provided to highlight the dispersion of opioid receptor systems through the brain. To determine whether brain opioid release may be induced by social interactions, half the animals were sacrificed following a 30 min period of social interaction while the other half were sacrificed following 30 min of social isolation. Opioid binding was higher in isolate-tested animals than socially-tested ones, suggesting that social interaction may promote endogenous brain opioid release

  1. Different amounts of ejaculatory activity, a natural rewarding behavior, induce differential mu and delta opioid receptor internalization in the rat's ventral tegmental area.

    Science.gov (United States)

    Garduño-Gutiérrez, René; León-Olea, Martha; Rodríguez-Manzo, Gabriela

    2013-12-06

    Opioid receptors internalize upon specific agonist stimulation. The in vivo significance of receptor internalization is not well established, partly due to the limited in vivo models used to study this phenomenon. Ejaculation promotes endogenous opioid release which activates opioid receptors at the brain, including the mesolimbic system and medial preoptic area. The objective of the present work was to analyze if there was a correlation between the degree of in vivo mu (MOR) and delta opioid receptor (DOR) internalization in the ventral tegmental area and the execution of different amounts of ejaculatory behavior of male rats. To this aim, we analyzed the brains of rats that ejaculated once or six successive times and of sexually exhausted rats with an established sexual inhibition, using immunofluorescence and confocal microscopy. Results showed that MOR and DOR internalization increased as a consequence of ejaculation. There was a relationship between the amount of sexual activity executed and the degree of internalization for MOR, but not for DOR. MOR internalization was larger in rats that ejaculated repeatedly than in animals ejaculating only once. Significant DOR internalization was found only in animals ejaculating once. Changes in MOR, DOR and beta arrestin2 detection, associated to sexual activity, were also found. It is suggested that copulation to satiety might be useful as a model system to study the biological significance of receptor internalization. © 2013 Published by Elsevier B.V.

  2. Blocking opioid receptors alters short-term feed intake and oro-sensorial preferences in weaned calves.

    Science.gov (United States)

    Montoro, C; Ipharraguerre, I R; Bach, A

    2012-05-01

    Opioid peptides may participate in the control of feed intake through mechanisms involving pleasure reward linked to consumption of palatable feed. The objective of this study was to determine whether blocking opioid receptors might void oro-sensorial preferences of calves, and affect circulating glucose, insulin, and anorexigenic hormones in fasted and fed calves. Two experiments involved 32 Holstein calves [body weight (BW)=86.5±1.73 kg, age=72±0.6 d]. In experiment 1, all calves received an ad libitum choice of the same feed either unflavored or flavored with a sweetener (Luctarom SFS-R, Lucta, Montornès del Vallès, Spain). Feed consumption was recorded every 2 h from 0800 to 1400 h for 3 consecutive days to verify the establishment of an oro-sensorial preference for sweet feed (SF). The SF was preferred over the control feed (CF) at all recorded times. In experiment 2, calves were subjected to a 2 × 2 factorial design to study the interaction between opioid activity and metabolic state. Half of the calves were fasted for 14 h (FAS), whereas the other half remained well fed (FED). Within each of these groups, at feeding time (0800 h), half of the calves received an i.v. injection of naloxone (NAL, an opioid receptor antagonist; 1 mg/kg of BW) and the other half was injected with saline solution (SAL; 0.9% NaCl). Therefore, treatments were FED-NAL, FED-SAL, FAS-NAL, and FAS-SAL. Blood samples were taken at -10, 20, 180, and 240 min relative to NAL or SAL injections. As expected, cumulative consumption of starter feed was greater in FAS than in FED calves. Total feed consumption 2 h after feeding was lower in NAL than in SAL calves. Calves in the FAS group did not discern between CF and SF during the first 4 h after feed offer. Preference for SF was greater in SAL than in NAL calves. Calves in the FED-SAL treatment preferred SF at 2 and 6 h after feed offer and tended to prefer SF at 4 h after feeding. However, FED-NAL calves did not discern between SF and CF

  3. Radiolabelled GLP-1 receptor antagonist binds to GLP-1 receptor-expressing human tissues

    Energy Technology Data Exchange (ETDEWEB)

    Waser, Beatrice; Reubi, Jean Claude [University of Berne, Division of Cell Biology and Experimental Cancer Research, Institute of Pathology, PO Box 62, Berne (Switzerland)

    2014-06-15

    Radiolabelled glucagon-like peptide 1 (GLP-1) receptor agonists have recently been shown to successfully image benign insulinomas in patients. For the somatostatin receptor targeting of tumours, however, it was recently reported that antagonist tracers were superior to agonist tracers. The present study therefore evaluated various forms of the {sup 125}iodinated-Bolton-Hunter (BH)-exendin(9-39) antagonist tracer for the in vitro visualization of GLP-1 receptor-expressing tissues in rats and humans and compared it with the agonist tracer {sup 125}I-GLP-1(7-36)amide. Receptor autoradiography studies with {sup 125}I-GLP-1(7-36)amide agonist or {sup 125}I-BH-exendin(9-39) antagonist radioligands were performed in human and rat tissues. The antagonist {sup 125}I-BH-exendin(9-39) labelled at lysine 19 identifies all human and rat GLP-1 target tissues and GLP-1 receptor-expressing tumours. Binding is of high affinity and is comparable in all tested tissues in its binding properties with the agonist tracer {sup 125}I-GLP-1(7-36)amide. For comparison, {sup 125}I-BH-exendin(9-39) with the BH labelled at lysine 4 did identify the GLP-1 receptor in rat tissues but not in human tissues. The GLP-1 receptor antagonist exendin(9-39) labelled with {sup 125}I-BH at lysine 19 is an excellent GLP-1 radioligand that identifies human and rat GLP-1 receptors in normal and tumoural tissues. It may therefore be the molecular basis to develop suitable GLP-1 receptor antagonist radioligands for in vivo imaging of GLP-1 receptor-expressing tissues in patients. (orig.)

  4. Radiolabelled GLP-1 receptor antagonist binds to GLP-1 receptor-expressing human tissues

    International Nuclear Information System (INIS)

    Waser, Beatrice; Reubi, Jean Claude

    2014-01-01

    Radiolabelled glucagon-like peptide 1 (GLP-1) receptor agonists have recently been shown to successfully image benign insulinomas in patients. For the somatostatin receptor targeting of tumours, however, it was recently reported that antagonist tracers were superior to agonist tracers. The present study therefore evaluated various forms of the 125 iodinated-Bolton-Hunter (BH)-exendin(9-39) antagonist tracer for the in vitro visualization of GLP-1 receptor-expressing tissues in rats and humans and compared it with the agonist tracer 125 I-GLP-1(7-36)amide. Receptor autoradiography studies with 125 I-GLP-1(7-36)amide agonist or 125 I-BH-exendin(9-39) antagonist radioligands were performed in human and rat tissues. The antagonist 125 I-BH-exendin(9-39) labelled at lysine 19 identifies all human and rat GLP-1 target tissues and GLP-1 receptor-expressing tumours. Binding is of high affinity and is comparable in all tested tissues in its binding properties with the agonist tracer 125 I-GLP-1(7-36)amide. For comparison, 125 I-BH-exendin(9-39) with the BH labelled at lysine 4 did identify the GLP-1 receptor in rat tissues but not in human tissues. The GLP-1 receptor antagonist exendin(9-39) labelled with 125 I-BH at lysine 19 is an excellent GLP-1 radioligand that identifies human and rat GLP-1 receptors in normal and tumoural tissues. It may therefore be the molecular basis to develop suitable GLP-1 receptor antagonist radioligands for in vivo imaging of GLP-1 receptor-expressing tissues in patients. (orig.)

  5. Plasticity of Signaling by Spinal Estrogen Receptor α, κ-Opioid Receptor, and Metabotropic Glutamate Receptors over the Rat Reproductive Cycle Regulates Spinal Endomorphin 2 Antinociception: Relevance of Endogenous-Biased Agonism.

    Science.gov (United States)

    Liu, Nai-Jiang; Murugaiyan, Vijaya; Storman, Emiliya M; Schnell, Stephen A; Kumar, Arjun; Wessendorf, Martin W; Gintzler, Alan R

    2017-11-15

    We previously showed that intrathecal application of endomorphin 2 [EM2; the highly specific endogenous μ-opioid receptor (MOR) ligand] induces antinociception that varies with stage of the rat estrous cycle: minimal during diestrus and prominent during proestrus. Earlier studies, however, did not identify proestrus-activated signaling strategies that enable spinal EM2 antinociception. We now report that in female rats, increased spinal dynorphin release and κ-opioid receptor (KOR) signaling, as well as the emergence of glutamate-activated metabotropic glutamate receptor 1 (mGluR 1 ) signaling, are critical to the transition from an EM2 nonresponsive state (during diestrus) to an analgesically responsive state (during proestrus). Differential signaling by mGluR 1 , depending on its activation by membrane estrogen receptor α (mERα; during diestrus) versus glutamate (during proestrus), concomitant with the ebb and flow of spinal dynorphin/KOR signaling, functions as a switch, preventing or promoting, respectively, spinal EM2 antinociception. Importantly, EM2 and glutamate-containing varicosities appose spinal neurons that express MOR along with mGluRs and mERα, suggesting that signaling mechanisms regulating analgesic effectiveness of intrathecally applied EM2 also pertain to endogenous EM2. Regulation of spinal EM2 antinociception by both the nature of the endogenous mGluR 1 activator (i.e., endogenous biased agonism at mGluR 1 ) and changes in spinal dynorphin/KOR signaling represent a novel mechanism for modulating analgesic responsiveness to endogenous EM2 (and perhaps other opioids). This points the way for developing noncanonical pharmacological approaches to pain management by harnessing endogenous opioids for pain relief. SIGNIFICANCE STATEMENT The current prescription opioid abuse epidemic underscores the urgency to develop alternative pharmacotherapies for managing pain. We find that the magnitude of spinal endomorphin 2 (EM2) antinociception not only

  6. The effects of opioid drugs on dopamine mediated locomotor activity in rats

    International Nuclear Information System (INIS)

    Leathern, L.L.

    1986-12-01

    Opioid drugs influence various behavioural parameters including locomotor activity in experimental animals. The interaction between the opioid and dopaminergic systems is one possible explanation for the effect of opioid drugs on locomotor activity. In this study behavioural and biochemical assays were done to investigate the interaction between the opioid and dopaminergic systems. Behavioural studies were done by measurement of locomotor activity (LA) of rats after acute or chronic pretreatment with opioid and/or dopaminergic drugs. Biochemical studies were in the form of radioligand binding assays, the effect on the number (Bmax) and affinity (K D ) of receptors was measured after chronic pretreatment with opioid and/or dopaminergic drugs. The opioid drugs used are morphine, nalbuphine and naloxone. Dopaminergic drugs used included: agonists-apomorphine and piribedil; antagonists-pimozide, haloperidol, chlorpromazine. In the acute situation increased LA was obtained with morphine and the DA agonists. A correlation between the behavioural and biochemical assays was found. Chronic pretreatment with morphine enhanced apomorphine induced LA, this supersensitivity was also measured as an increased receptor density (Bmax) of D2 receptors in the striatum. Chronic morphine pretreatment caused a decrease in morphine induced LA, while this subsensitivity was not apparent in the ligand binding assays - where no change in receptor number was observed. Chronic naloxone pretreatment enhanced morphine induced LA, as well as increased the Bmax of opioid receptors in the whole brain. It is concluded that an interaction between the opioid and dopaminergic systems does exist, and may account for the mechanism of action of the opioids

  7. The glycocalyx promotes cooperative binding and clustering of adhesion receptors.

    Science.gov (United States)

    Xu, Guang-Kui; Qian, Jin; Hu, Jinglei

    2016-05-18

    Cell adhesion plays a pivotal role in various biological processes, e.g., immune responses, cancer metastasis, and stem cell differentiation. The adhesion behaviors depend subtly on the binding kinetics of receptors and ligands restricted at the cell-substrate interfaces. Although much effort has been directed toward investigating the kinetics of adhesion molecules, the role of the glycocalyx, anchored on cell surfaces as an exterior layer, is still unclear. In this paper, we propose a theoretical approach to study the collective binding kinetics of a few and a large number of binders in the presence of the glycocalyx, representing the cases of initial and mature adhesions of cells, respectively. The analytical results are validated by finding good agreement with our Monte Carlo simulations. In the force loading case, the on-rate and affinity increase as more bonds form, whereas this cooperative effect is not observed in the displacement loading case. The increased thickness and stiffness of the glycocalyx tend to decrease the affinity for a few bonds, while they have less influence on the affinity for a large number of bonds. Moreover, for a flexible membrane with thermally-excited shape fluctuations, the glycocalyx is exhibited to promote the formation of bond clusters, mainly due to the cooperative binding of binders. This study helps to understand the cooperative kinetics of adhesion receptors under physiologically relevant loading conditions and sheds light on the novel role of the glycocalyx in cell adhesion.

  8. Receptor-ligand binding sites and virtual screening.

    Science.gov (United States)

    Hattotuwagama, Channa K; Davies, Matthew N; Flower, Darren R

    2006-01-01

    Within the pharmaceutical industry, the ultimate source of continuing profitability is the unremitting process of drug discovery. To be profitable, drugs must be marketable: legally novel, safe and relatively free of side effects, efficacious, and ideally inexpensive to produce. While drug discovery was once typified by a haphazard and empirical process, it is now increasingly driven by both knowledge of the receptor-mediated basis of disease and how drug molecules interact with receptors and the wider physiome. Medicinal chemistry postulates that to understand a congeneric ligand series, or set thereof, is to understand the nature and requirements of a ligand binding site. Likewise, structural molecular biology posits that to understand a binding site is to understand the nature of ligands bound therein. Reality sits somewhere between these extremes, yet subsumes them both. Complementary to rules of ligand design, arising through decades of medicinal chemistry, structural biology and computational chemistry are able to elucidate the nature of binding site-ligand interactions, facilitating, at both pragmatic and conceptual levels, the drug discovery process.

  9. Ablation of kappa-opioid receptors from brain dopamine neurons has anxiolytic-like effects and enhances cocaine-induced plasticity.

    Science.gov (United States)

    Van't Veer, Ashlee; Bechtholt, Anita J; Onvani, Sara; Potter, David; Wang, Yujun; Liu-Chen, Lee-Yuan; Schütz, Günther; Chartoff, Elena H; Rudolph, Uwe; Cohen, Bruce M; Carlezon, William A

    2013-07-01

    Brain kappa-opioid receptors (KORs) are implicated in states of motivation and emotion. Activation of KORs negatively regulates mesolimbic dopamine (DA) neurons, and KOR agonists produce depressive-like behavioral effects. To further evaluate how KOR function affects behavior, we developed mutant mice in which exon 3 of the KOR gene (Oprk1) was flanked with Cre-lox recombination (loxP) sites. By breeding these mice with lines that express Cre-recombinase (Cre) in early embryogenesis (EIIa-Cre) or only in DA neurons (dopamine transporter (DAT)-Cre), we developed constitutive KOR knockouts (KOR(-/-)) and conditional knockouts that lack KORs in DA-containing neurons (DAT-KOR(lox/lox)). Autoradiography demonstrated complete ablation of KOR binding in the KOR(-/-) mutants, and reduced binding in the DAT-KOR(lox/lox) mutants. Quantitative reverse transcription PCR (qPCR) studies confirmed that KOR mRNA is undetectable in the constitutive mutants and reduced in the midbrain DA systems of the conditional mutants. Behavioral characterization demonstrated that these mutant lines do not differ from controls in metrics, including hearing, vision, weight, and locomotor activity. Whereas KOR(-/-) mice appeared normal in the open field and light/dark box tests, DAT-KOR(lox/lox) mice showed reduced anxiety-like behavior, an effect that is broadly consistent with previously reported effects of KOR antagonists. Sensitization to the locomotor-stimulating effects of cocaine appeared normal in KOR(-/-) mutants, but was exaggerated in DAT-KOR(lox/lox) mutants. Increased sensitivity to cocaine in the DAT-KOR(lox/lox) mutants is consistent with a role for KORs in negative regulation of DA function, whereas the lack of differences in the KOR(-/-) mutants suggests compensatory adaptations after constitutive receptor ablation. These mouse lines may be useful in future studies of KOR function.

  10. Effect of Tramadol (μ-opioid receptor agonist on orthodontic tooth movements in a rat model

    Directory of Open Access Journals (Sweden)

    E. Javadi

    2012-01-01

    Full Text Available Objective: Tramadol is a synthetic analgesic of opioids which has more flexible mechanisms of action than typical opioids. Since it has been reported in previous study that typical opioids like morphine can affect the bone homeostasis, it is worthwhile to examine the effects of tramadol on tooth movement. In this study we investigated effects of tramadol on orthodontic tooth movement in rats.Materials and Methods: 30 male wistar rats were selected and received orthodontic appliance. 3 groups were designed based on the substance that they received daily injections of during a 2-week orthodontic treatment. 1. Control group with no injection.2.Control group with normal saline injection.3. the tramadol group. After the two-week treatment period the amount of tooth movement were measured in all the groups. Also the histological analysis was performed assessing the root resorption, osteoclasts numbers and bone resorption.Results: The amount of tooth movement was not significantl in the tramadol group comparing to the other groups (P>0.05.The results of 3 histological parameters (amount of root resorption, osteoclastic numbers and bone resorption were statistically insignificant (P>0.05.Conclusion: Tramadol as an atypical opioid does not interfere with the process of bone remodeling and tooth movement in rat. Tramadol does not affect osteoclastic activity and bone resorption and it does not cause to change the resulted root resorption either.

  11. Sex differences in subcellular distribution of delta opioid receptors in the rat hippocampus in response to acute and chronic stress

    Directory of Open Access Journals (Sweden)

    Sanoara Mazid

    2016-12-01

    Full Text Available Drug addiction requires associative learning processes that critically involve hippocampal circuits, including the opioid system. We recently found that acute and chronic stress, important regulators of addictive processes, affect hippocampal opioid levels and mu opioid receptor trafficking in a sexually dimorphic manner. Here, we examined whether acute and chronic stress similarly alters the levels and trafficking of hippocampal delta opioid receptors (DORs. Immediately after acute immobilization stress (AIS or one-day after chronic immobilization stress (CIS, the brains of adult female and male rats were perfusion-fixed with aldehydes. The CA3b region and the dentate hilus of the dorsal hippocampus were quantitatively analyzed by light microscopy using DOR immunoperoxidase or dual label electron microscopy for DOR using silver intensified immunogold particles (SIG and GABA using immunoperoxidase. At baseline, females compared to males had more DORs near the plasmalemma of pyramidal cell dendrites and about 3 times more DOR-labeled CA3 dendritic spines contacted by mossy fibers. In AIS females, near-plasmalemmal DOR-SIGs decreased in GABAergic hilar dendrites. However, in AIS males, near-plasmalemmal DOR-SIGs increased in CA3 pyramidal cell and hilar GABAergic dendrites and the percentage of CA3 dendritic spines contacted by mossy fibers increased to about half that seen in unstressed females. Conversely, after CIS, near-plasmalemmal DOR-SIGs increased in hilar GABA-labeled dendrites of females whereas in males plasmalemmal DOR-SIGs decreased in CA3 pyramidal cell dendrites and near-plasmalemmal DOR-SIGs decreased hilar GABA-labeled dendrites. As CIS in females, but not males, redistributed DOR-SIGs near the plasmalemmal of hilar GABAergic dendrites, a subsequent experiment examined the acute affect of oxycodone on the redistribution of DOR-SIGs in a separate cohort of CIS females. Plasmalemmal DOR-SIGs were significantly elevated on hilar

  12. Human myometrial adrenergic receptors during pregnancy: identification of the alpha-adrenergic receptor by [3H] dihydroergocryptine binding

    International Nuclear Information System (INIS)

    Jacobs, M.M.; Hayashida, D.; Roberts, J.M.

    1985-01-01

    The radioactive alpha-adrenergic antagonist [ 3 H] dihydroergocryptine binds to particulate preparations of term pregnant human myometrium in a manner compatible with binding to the alpha-adrenergic receptor (alpha-receptor). [ 3 H] Dihydroergocryptine binds with high affinity (KD = 2 nmol/L and low capacity (receptor concentration = 100 fmol/mg of protein). Adrenergic agonists compete for [ 3 H] dihydroergocryptine binding sites stereo-selectively ([-]-norepinephrine is 100 times as potent as [+]-norepinephrine) and in a manner compatible with alpha-adrenergic potencies (epinephrine approximately equal to norepinephrine much greater than isoproterenol). Studies in which prazosin, an alpha 1-antagonist, and yohimbine, and alpha 2-antagonist, competed for [ 3 H] dihydroergocryptine binding sites in human myometrium indicated that approximately 70% are alpha 2-receptors and that 30% are alpha 1-receptors. [ 3 H] dihydroergocryptine binding to human myometrial membrane particulate provides an important tool with which to study the molecular mechanisms of uterine alpha-adrenergic response

  13. Reduced striatal D2 receptor binding in myoclonus-dystonia

    International Nuclear Information System (INIS)

    Beukers, R.J.; Weisscher, N.; Tijssen, M.A.J.; Booij, J.; Zijlstra, F.; Amelsvoort, T.A.M.J. van

    2009-01-01

    To study striatal dopamine D 2 receptor availability in DYT11 mutation carriers of the autosomal dominantly inherited disorder myoclonus-dystonia (M-D). Fifteen DYT11 mutation carriers (11 clinically affected) and 15 age- and sex-matched controls were studied using 123 I-IBZM SPECT. Specific striatal binding ratios were calculated using standard templates for striatum and occipital areas. Multivariate analysis with corrections for ageing and smoking showed significantly lower specific striatal to occipital IBZM uptake ratios (SORs) both in the left and right striatum in clinically affected patients and also in all DYT11 mutation carriers compared to control subjects. Our findings are consistent with the theory of reduced dopamine D 2 receptor (D2R) availability in dystonia, although the possibility of increased endogenous dopamine, and consequently, competitive D2R occupancy cannot be ruled out. (orig.)

  14. Fluorescent Receptor Binding Assay for Detecting Ciguatoxins in Fish.

    Science.gov (United States)

    Hardison, D Ransom; Holland, William C; McCall, Jennifer R; Bourdelais, Andrea J; Baden, Daniel G; Darius, H Taiana; Chinain, Mireille; Tester, Patricia A; Shea, Damian; Quintana, Harold A Flores; Morris, James A; Litaker, R Wayne

    2016-01-01

    Ciguatera fish poisoning is an illness suffered by > 50,000 people yearly after consumption of fish containing ciguatoxins (CTXs). One of the current methodologies to detect ciguatoxins in fish is a radiolabeled receptor binding assay (RBA(R)). However, the license requirements and regulations pertaining to radioisotope utilization can limit the applicability of the RBA(R) in certain labs. A fluorescence based receptor binding assay (RBA(F)) was developed to provide an alternative method of screening fish samples for CTXs in facilities not certified to use radioisotopes. The new assay is based on competition binding between CTXs and fluorescently labeled brevetoxin-2 (BODIPY®-PbTx-2) for voltage-gated sodium channel receptors at site 5 instead of a radiolabeled brevetoxin. Responses were linear in fish tissues spiked from 0.1 to 1.0 ppb with Pacific ciguatoxin-3C (P-CTX-3C) with a detection limit of 0.075 ppb. Carribean ciguatoxins were confirmed in Caribbean fish by LC-MS/MS analysis of the regional biomarker (C-CTX-1). Fish (N = 61) of six different species were screened using the RBA(F). Results for corresponding samples analyzed using the neuroblastoma cell-based assay (CBA-N2a) correlated well (R2 = 0.71) with those of the RBA(F), given the low levels of CTX present in positive fish. Data analyses also showed the resulting toxicity levels of P-CTX-3C equivalents determined by CBA-N2a were consistently lower than the RBA(F) affinities expressed as % binding equivalents, indicating that a given amount of toxin bound to the site 5 receptors translates into corresponding lower cytotoxicity. Consequently, the RBA(F), which takes approximately two hours to perform, provides a generous estimate relative to the widely used CBA-N2a which requires 2.5 days to complete. Other RBA(F) advantages include the long-term (> 5 years) stability of the BODIPY®-PbTx-2 and having similar results as the commonly used RBA(R). The RBA(F) is cost-effective, allows high sample

  15. Fluorescent Receptor Binding Assay for Detecting Ciguatoxins in Fish.

    Directory of Open Access Journals (Sweden)

    D Ransom Hardison

    Full Text Available Ciguatera fish poisoning is an illness suffered by > 50,000 people yearly after consumption of fish containing ciguatoxins (CTXs. One of the current methodologies to detect ciguatoxins in fish is a radiolabeled receptor binding assay (RBA(R. However, the license requirements and regulations pertaining to radioisotope utilization can limit the applicability of the RBA(R in certain labs. A fluorescence based receptor binding assay (RBA(F was developed to provide an alternative method of screening fish samples for CTXs in facilities not certified to use radioisotopes. The new assay is based on competition binding between CTXs and fluorescently labeled brevetoxin-2 (BODIPY®-PbTx-2 for voltage-gated sodium channel receptors at site 5 instead of a radiolabeled brevetoxin. Responses were linear in fish tissues spiked from 0.1 to 1.0 ppb with Pacific ciguatoxin-3C (P-CTX-3C with a detection limit of 0.075 ppb. Carribean ciguatoxins were confirmed in Caribbean fish by LC-MS/MS analysis of the regional biomarker (C-CTX-1. Fish (N = 61 of six different species were screened using the RBA(F. Results for corresponding samples analyzed using the neuroblastoma cell-based assay (CBA-N2a correlated well (R2 = 0.71 with those of the RBA(F, given the low levels of CTX present in positive fish. Data analyses also showed the resulting toxicity levels of P-CTX-3C equivalents determined by CBA-N2a were consistently lower than the RBA(F affinities expressed as % binding equivalents, indicating that a given amount of toxin bound to the site 5 receptors translates into corresponding lower cytotoxicity. Consequently, the RBA(F, which takes approximately two hours to perform, provides a generous estimate relative to the widely used CBA-N2a which requires 2.5 days to complete. Other RBA(F advantages include the long-term (> 5 years stability of the BODIPY®-PbTx-2 and having similar results as the commonly used RBA(R. The RBA(F is cost-effective, allows high sample

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

    Directory of Open Access Journals (Sweden)

    Abrar Ashoor

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

  17. Plasma membrane cholesterol level and agonist-induced internalization of delta-opioid receptors; colocalization study with intracellular membrane markers of Rab family\

    Czech Academy of Sciences Publication Activity Database

    Brejchová, Jana; Vošahlíková, Miroslava; Roubalová, Lenka; Parenti, M.; Mauri, M.; Chernyavskiy, Oleksandr; Svoboda, Petr

    2016-01-01

    Roč. 48, č. 4 (2016), s. 375-396 ISSN 0145-479X R&D Projects: GA ČR(CZ) GAP207/12/0919 Institutional support: RVO:67985823 Keywords : cholesterol * plasma membrane * delta-opioid receptor * internalization * Rab proteins Subject RIV: CE - Biochemistry Impact factor: 2.576, year: 2016

  18. High Efficacy but Low Potency of delta-Opioid Receptor-G Protein Coupling in Brij-58-Treated, Low-Density Plasma Membrane Fragments

    Czech Academy of Sciences Publication Activity Database

    Roubalová, Lenka; Vošahlíková, Miroslava; Brejchová, Jana; Sýkora, Jan; Rudajev, Vladimír; Svoboda, Petr

    2015-01-01

    Roč. 10, č. 8 (2015), e0135664 E-ISSN 1932-6203 R&D Projects: GA ČR(CZ) GAP207/12/0919 Institutional support: RVO:67985823 ; RVO:61388955 Keywords : delta - opioid receptor * G protein coupling * detergent * efficacy * potency Subject RIV: CE - Biochemistry; CF - Physical ; Theoretical Chemistry (UFCH-W) Impact factor: 3.057, year: 2015

  19. The influences of reproductive status and acute stress on the levels of phosphorylated mu opioid receptor immunoreactivity in rat hippocampus

    Directory of Open Access Journals (Sweden)

    Keith L. Gonzales

    2011-08-01

    Full Text Available Opioids play a critical role in hippocampally dependent behavior and plasticity. In the hippocampal formation, mu opioid receptors (MOR are prominent in parvalbumin (PARV containing interneurons. Previously we found that gonadal hormones modulate the trafficking of MORs in PARV interneurons. Although sex differences in response to stress are well documented, the point at which opioids, sex and stress interact to influence hippocampal function remains elusive. Thus, we used quantitative immunocytochemistry in combination with light and electron microscopy for the phosphorylated MOR at the SER375 carboxy-terminal residue (pMOR in male and female rats to assess these interactions. In both sexes, pMOR-immunoreactivity (ir was prominent in axons and terminals and in a few neuronal somata and dendrites, some of which contained PARV in the mossy fiber pathway region of the dentate gyrus (DG hilus and CA3 stratum lucidum. In unstressed rats, the levels of pMOR-ir in the DG or CA3 were not affected by sex or estrous cycle stage. However, immediately following 30 minutes of acute immobilization stress (AIS, males had higher levels of pMOR-ir whereas females at proestrus and estrus (high estrogen stages had lower levels of pMOR-ir within the DG. In contrast, the number and types of neuronal profiles with pMOR-ir were not altered by AIS in either males or proestrus females. These data demonstrate that although gonadal steroids do not affect pMOR levels at resting conditions, they are differentially activated both pre- and post-synaptic MORs following stress. These interactions may contribute to the reported sex differences in hippocampally dependent behaviors in stressed animals.

  20. Opioid Antagonists and the A118G Polymorphism in the μ-Opioid Receptor Gene: Effects of GSK1521498 and Naltrexone in Healthy Drinkers Stratified by OPRM1 Genotype

    Science.gov (United States)

    Ziauddeen, Hisham; Nestor, Liam J; Subramaniam, Naresh; Dodds, Chris; Nathan, Pradeep J; Miller, Sam R; Sarai, Bhopinder K; Maltby, Kay; Fernando, Disala; Warren, Liling; Hosking, Louise K; Waterworth, Dawn; Korzeniowska, Anna; Win, Beta; Richards, Duncan B; Vasist Johnson, Lakshmi; Fletcher, Paul C; Bullmore, Edward T

    2016-01-01

    The A118G single-nucleotide polymorphism (SNP rs1799971) in the μ-opioid receptor gene, OPRM1, has been much studied in relation to alcohol use disorders. The reported effects of allelic variation at this SNP on alcohol-related behaviors, and on opioid receptor antagonist treatments, have been inconsistent. We investigated the pharmacogenetic interaction between A118G variation and the effects of two μ-opioid receptor antagonists in a clinical lab setting. Fifty-six overweight and moderate–heavy drinkers were prospectively stratified by genotype (29 AA homozygotes, 27 carriers of at least 1 G allele) in a double-blind placebo-controlled, three-period crossover design with naltrexone (NTX; 25 mg OD for 2 days, then 50 mg OD for 3 days) and GSK1521498 (10 mg OD for 5 days). The primary end point was regional brain activation by the contrast between alcohol and neutral tastes measured using functional magnetic resonance imaging (fMRI). Secondary end points included other fMRI contrasts, subjective responses to intravenous alcohol challenge, and food intake. GSK1521498 (but not NTX) significantly attenuated fMRI activation by appetitive tastes in the midbrain and amygdala. GSK1521498 (and NTX to a lesser extent) significantly affected self-reported responses to alcohol infusion. Both drugs reduced food intake. Across all end points, there was less robust evidence for significant effects of OPRM1 allelic variation, or for pharmacogenetic interactions between genotype and drug treatment. These results do not support strong modulatory effects of OPRM1 genetic variation on opioid receptor antagonist attenuation of alcohol- and food-related behaviors. However, they do support further investigation of GSK1521498 as a potential therapeutic for alcohol use and eating disorders. PMID:27109624

  1. Alterations in opioid parameters in the hypothalamus of rats with estradiol-induced polycystic ovarian disease

    International Nuclear Information System (INIS)

    Desjardins, G.C.; Beaudet, A.; Brawer, J.R.

    1990-01-01

    The distribution and density of selectively labeled mu-, delta-, and kappa-opioid binding sites were examined by in vitro radioautography in the hypothalamus of normal, estradiol valerate (EV)-injected, and estradiol (E2)-implanted female rats. Hypothalamic beta-endorphin concentration was also examined by RIA in these three groups of animals. Quantitative analysis of film radioautographs demonstrated a selective increase in mu-opioid binding in the medial preoptic area of EV-treated, but not of E2-implanted rats. However, both these estrogenized groups exhibited a reduction in the density of delta-opioid binding in the suprachiasmatic nucleus. Statistically significant changes between either estrogenized groups were not observed for kappa-opioid binding. Results on the hypothalamic concentration of beta-endorphin indicated a marked reduction in EV-injected animals with respect to controls. In contrast, the E2-implanted animals exhibited beta-endorphin concentrations similar to controls. The present results confirm the increase in opioid receptor binding previously reported in the hypothalamus of EV-treated rats and further demonstrate that this increase is confined to the medial preoptic area and exclusively concerns mu-opioid receptors. The concomitant reduction in beta-endorphin levels observed in the same group of animals suggests that the observed increase in mu-opioid binding could reflect a chronic up-regulation of the receptor in response to compromised beta-endorphin input. Given the restriction of this effect to the site of origin of LHRH neurons and the demonstrated inhibitory role of opioids on LHRH release, it is tempting to postulate that such up-regulation could lead to the suppression of the plasma LH pattern that characterizes polycystic ovarian disease in the EV-treated rat

  2. Alterations in opioid parameters in the hypothalamus of rats with estradiol-induced polycystic ovarian disease

    Energy Technology Data Exchange (ETDEWEB)

    Desjardins, G.C.; Beaudet, A.; Brawer, J.R. (McGill Univ., Quebec (Canada))

    1990-12-01

    The distribution and density of selectively labeled mu-, delta-, and kappa-opioid binding sites were examined by in vitro radioautography in the hypothalamus of normal, estradiol valerate (EV)-injected, and estradiol (E2)-implanted female rats. Hypothalamic beta-endorphin concentration was also examined by RIA in these three groups of animals. Quantitative analysis of film radioautographs demonstrated a selective increase in mu-opioid binding in the medial preoptic area of EV-treated, but not of E2-implanted rats. However, both these estrogenized groups exhibited a reduction in the density of delta-opioid binding in the suprachiasmatic nucleus. Statistically significant changes between either estrogenized groups were not observed for kappa-opioid binding. Results on the hypothalamic concentration of beta-endorphin indicated a marked reduction in EV-injected animals with respect to controls. In contrast, the E2-implanted animals exhibited beta-endorphin concentrations similar to controls. The present results confirm the increase in opioid receptor binding previously reported in the hypothalamus of EV-treated rats and further demonstrate that this increase is confined to the medial preoptic area and exclusively concerns mu-opioid receptors. The concomitant reduction in beta-endorphin levels observed in the same group of animals suggests that the observed increase in mu-opioid binding could reflect a chronic up-regulation of the receptor in response to compromised beta-endorphin input. Given the restriction of this effect to the site of origin of LHRH neurons and the demonstrated inhibitory role of opioids on LHRH release, it is tempting to postulate that such up-regulation could lead to the suppression of the plasma LH pattern that characterizes polycystic ovarian disease in the EV-treated rat.

  3. Dendroaspis natriuretic peptide binds to the natriuretic peptide clearance receptor

    International Nuclear Information System (INIS)

    Johns, Douglas G.; Ao, Zhaohui; Heidrich, Bradley J.; Hunsberger, Gerald E.; Graham, Taylor; Payne, Lisa; Elshourbagy, Nabil; Lu, Quinn; Aiyar, Nambi; Douglas, Stephen A.

    2007-01-01

    Dendroaspis natriuretic peptide (DNP) is a newly-described natriuretic peptide which lowers blood pressure via vasodilation. The natriuretic peptide clearance receptor (NPR-C) removes natriuretic peptides from the circulation, but whether DNP interacts with human NPR-C directly is unknown. The purpose of this study was to test the hypothesis that DNP binds to NPR-C. ANP, BNP, CNP, and the NPR-C ligands AP-811 and cANP(4-23) displaced [ 125 I]-ANP from NPR-C with pM-to-nM K i values. DNP displaced [ 125 I]-ANP from NPR-C with nM potency, which represents the first direct demonstration of binding of DNP to human NPR-C. DNP showed high pM affinity for the GC-A receptor and no affinity for GC-B (K i > 1000 nM). DNP was nearly 10-fold more potent than ANP at stimulating cGMP production in GC-A expressing cells. Blockade of NPR-C might represent a novel therapeutic approach in augmenting the known beneficial actions of DNP in cardiovascular diseases such as hypertension and heart failure

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

    International Nuclear Information System (INIS)

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

    1987-01-01

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

  5. Distinct phosphotyrosines on a growth factor receptor bind to specific molecules that mediate different signaling pathways.

    Science.gov (United States)

    Fantl, W J; Escobedo, J A; Martin, G A; Turck, C W; del Rosario, M; McCormick, F; Williams, L T

    1992-05-01

    The receptor for platelet-derived growth factor (PDGF) binds two proteins containing SH2 domains, GTPase activating protein (GAP) and phosphatidylinositol 3-kinase (PI3-kinase). The sites on the receptor that mediate this interaction were identified by using phosphotyrosine-containing peptides representing receptor sequences to block specifically binding of either PI3-kinase or GAP. These results suggested that PI3-kinase binds two phosphotyrosine residues, each located in a 5 aa motif with an essential methionine at the fourth position C-terminal to the tyrosine. Point mutations at these sites caused a selective elimination of PI3-kinase binding and loss of PDGF-stimulated DNA synthesis. Mutation of the binding site for GAP prevented the receptor from associating with or phosphorylating GAP, but had no effect on PI3-kinase binding and little effect on DNA synthesis. Therefore, GAP and PI3-kinase interact with the receptor by binding to different phosphotyrosine-containing sequence motifs.

  6. Genetic, functional and molecular features of glucocorticoid receptor binding.

    Directory of Open Access Journals (Sweden)

    Francesca Luca

    Full Text Available Glucocorticoids (GCs are key mediators of stress response and are widely used as pharmacological agents to treat immune diseases, such as asthma and inflammatory bowel disease, and certain types of cancer. GCs act mainly by activating the GC receptor (GR, which interacts with other transcription factors to regulate gene expression. Here, we combined different functional genomics approaches to gain molecular insights into the mechanisms of action of GC. By profiling the transcriptional response to GC over time in 4 Yoruba (YRI and 4 Tuscans (TSI lymphoblastoid cell lines (LCLs, we suggest that the transcriptional response to GC is variable not only in time, but also in direction (positive or negative depending on the presence of specific interacting transcription factors. Accordingly, when we performed ChIP-seq for GR and NF-κB in two YRI LCLs treated with GC or with vehicle control, we observed that features of GR binding sites differ for up- and down-regulated genes. Finally, we show that eQTLs that affect expression patterns only in the presence of GC are 1.9-fold more likely to occur in GR binding sites, compared to eQTLs that affect expression only in its absence. Our results indicate that genetic variation at GR and interacting transcription factors binding sites influences variability in gene expression, and attest to the power of combining different functional genomic approaches.

  7. Benzodiazepine receptor binding in vivo with (/sup 3/)-Ro 15-1788

    Energy Technology Data Exchange (ETDEWEB)

    Goeders, N.E.; Kuhar, M.J.

    1985-07-29

    In vivo benzodiazepine receptor binding has generally been studied by ex vivo techniques. In this investigation, the authors identify the conditions where (/sup 3/H)-Ro 15-1788 labels benzodiazepine receptors by true in vivo binding, i.e. where workable specific to nonspecific ratios are obtained in intact tissues without homogenization or washing. (/sup 3/H)-Flunitrazepam and (/sup 3/H)-clonazepam did not exhibit useful in vivo receptor binding. 39 references, 5 figures, 1 table.

  8. Ligand- and cell-dependent determinants of internalization and cAMP modulation by delta opioid receptor (DOR) agonists

    Science.gov (United States)

    Charfi, Iness; Nagi, Karim; Mnie-Filali, Ouissame; Thibault, Dominic; Balboni, Gianfranco; Schiller, Peter W.; Trudeau, Louis-Eric

    2014-01-01

    Signaling bias refers to G protein-coupled receptor ligand ability to preferentially activate one type of signal over another. Bias to evoke signaling as opposed to sequestration has been proposed as a predictor of opioid ligand potential for generating tolerance. Here we measured whether delta opioid receptor agonists preferentially inhibited cyclase activity over internalization in HEK cells. Efficacy (τ) and affinity (KA) values were estimated from functional data and bias was calculated from efficiency coefficients (log τ/KA). This approach better represented the data as compared to alternative methods that estimate bias exclusively from τ values. Log (τ/KA) coefficients indicated that SNC-80 and UFP-512 promoted cyclase inhibition more efficiently than DOR internalization as compared to DPDPE (bias factor for SNC-80: 50 and for UFP-512: 132). Molecular determinants of internalization were different in HEK293 cells and neurons with βarrs contributing to internalization in both cell types, while PKC and GRK2 activities were only involved in neurons. Rank orders of ligand ability to engage different internalization mechanisms in neurons were compared to rank order of Emax values for cyclase assays in HEK cells. Comparison revealed a significant reversal in rank order for cyclase Emax values and βarr-dependent internalization in neurons, indicating that these responses were ligand-specific. Despite this evidence, and because kinases involved in internalization were not the same across cellular backgrounds, it is not possible to assert if the magnitude and nature of bias revealed by rank orders of maximal responses is the same as the one measured in HEK cells. PMID:24022593

  9. Characterizing low affinity epibatidine binding to α4β2 nicotinic acetylcholine receptors with ligand depletion and nonspecific binding

    Science.gov (United States)

    2011-01-01

    Background Along with high affinity binding of epibatidine (Kd1≈10 pM) to α4β2 nicotinic acetylcholine receptor (nAChR), low affinity binding of epibatidine (Kd2≈1-10 nM) to an independent binding site has been reported. Studying this low affinity binding is important because it might contribute understanding about the structure and synthesis of α4β2 nAChR. The binding behavior of epibatidine and α4β2 AChR raises a question about interpreting binding data from two independent sites with ligand depletion and nonspecific binding, both of which can affect equilibrium binding of [3H]epibatidine and α4β2 nAChR. If modeled incorrectly, ligand depletion and nonspecific binding lead to inaccurate estimates of binding constants. Fitting total equilibrium binding as a function of total ligand accurately characterizes a single site with ligand depletion and nonspecific binding. The goal of this study was to determine whether this approach is sufficient with two independent high and low affinity sites. Results Computer simulations of binding revealed complexities beyond fitting total binding for characterizing the second, low affinity site of α4β2 nAChR. First, distinguishing low-affinity specific binding from nonspecific binding was a potential problem with saturation data. Varying the maximum concentration of [3H]epibatidine, simultaneously fitting independently measured nonspecific binding, and varying α4β2 nAChR concentration were effective remedies. Second, ligand depletion helped identify the low affinity site when nonspecific binding was significant in saturation or competition data, contrary to a common belief that ligand depletion always is detrimental. Third, measuring nonspecific binding without α4β2 nAChR distinguished better between nonspecific binding and low-affinity specific binding under some circumstances of competitive binding than did presuming nonspecific binding to be residual [3H]epibatidine binding after adding a large concentration of

  10. Characterizing low affinity epibatidine binding to α4β2 nicotinic acetylcholine receptors with ligand depletion and nonspecific binding

    Directory of Open Access Journals (Sweden)

    Person Alexandra M

    2011-11-01

    Full Text Available Abstract Background Along with high affinity binding of epibatidine (Kd1≈10 pM to α4β2 nicotinic acetylcholine receptor (nAChR, low affinity binding of epibatidine (Kd2≈1-10 nM to an independent binding site has been reported. Studying this low affinity binding is important because it might contribute understanding about the structure and synthesis of α4β2 nAChR. The binding behavior of epibatidine and α4β2 AChR raises a question about interpreting binding data from two independent sites with ligand depletion and nonspecific binding, both of which can affect equilibrium binding of [3H]epibatidine and α4β2 nAChR. If modeled incorrectly, ligand depletion and nonspecific binding lead to inaccurate estimates of binding constants. Fitting total equilibrium binding as a function of total ligand accurately characterizes a single site with ligand depletion and nonspecific binding. The goal of this study was to determine whether this approach is sufficient with two independent high and low affinity sites. Results Computer simulations of binding revealed complexities beyond fitting total binding for characterizing the second, low affinity site of α4β2 nAChR. First, distinguishing low-affinity specific binding from nonspecific binding was a potential problem with saturation data. Varying the maximum concentration of [3H]epibatidine, simultaneously fitting independently measured nonspecific binding, and varying α4β2 nAChR concentration were effective remedies. Second, ligand depletion helped identify the low affinity site when nonspecific binding was significant in saturation or competition data, contrary to a common belief that ligand depletion always is detrimental. Third, measuring nonspecific binding without α4β2 nAChR distinguished better between nonspecific binding and low-affinity specific binding under some circumstances of competitive binding than did presuming nonspecific binding to be residual [3H]epibatidine binding after

  11. Evidence for endogenous opioid release in the amygdala during positive emotion.

    Science.gov (United States)

    Koepp, M J; Hammers, A; Lawrence, A D; Asselin, M C; Grasby, P M; Bench, C J

    2009-01-01

    Endogenous opioid release has been linked to relief from aversive emotional memories, thereby promoting a euphoric state and subsequent interactions towards social stimuli resulting in the formation of social preferences. However, this theory remains controversial. Using positron emission tomography and [(11)C]diprenorphine (DPN) in healthy volunteers, we found significantly reduced DPN binding to opioid receptor in the hippocampus during positive mood induction compared to neutral mood. Furthermore, the magnitude of positive mood change correlated negatively with DPN binding in the amygdala bilaterally. Our finding of reduced DPN binding is consistent with increased release of endogenous opioids, providing direct evidence that localised release of endogenous opioids is involved in the regulation of positive emotion in humans.

  12. Kappa-opioid receptor signaling in the striatum as a potential modulator of dopamine transmission in cocaine dependence

    Directory of Open Access Journals (Sweden)

    Pierre eTrifilieff

    2013-06-01

    Full Text Available Cocaine addiction is accompanied by a decrease in striatal dopamine signaling, measured as a decrease in dopamine D2 receptor binding as well as blunted dopamine release in the striatum. These alterations in dopamine transmission have clinical relevance, and have been shown to correlate with cocaine-seeking behavior and response to treatment for cocaine dependence. However, the mechanisms contributing to the hypodopaminergic state in cocaine addiction remain unknown. Here we review the Positron Emission Tomography (PET imaging studies showing alterations in D2 receptor binding potential and dopamine transmission in cocaine abusers and their significance in cocaine-seeking behavior. Based on animal and human studies, we propose that the kappa receptor/dynorphin system, because of its impact on dopamine transmission and upregulation following cocaine exposure, could contribute to the hypodopaminergic state reported in cocaine addiction, and could thus be a relevant target for treatment development.

  13. Tritium labelling of highly selective probes for. delta. -opioid receptors: ( sup 3 H)Tyr-D-Ser(O-t-Bu)-Gly-Phe-Leu-Thr(DSTBULET) and ( sup 3 H)Tyr-D-Ser(O-t-Bu)-Gly-Phe-Leu-Thr(O-t-Bu)(BUBU)

    Energy Technology Data Exchange (ETDEWEB)

    Fellion, E.; Gacel, G.; Roques, B.P. (Institut National de la Sante et de la Recherche Medicale (INSERM), 75 - Paris (France)); Roy, J.; Morgat, J.L. (CEA Centre d' Etudes Nucleaires de Saclay, 91 - Gif-sur-Yvette (France). Service de Biochimie)

    1990-08-01

    The introduction of bulky residue(s) in linear enkephalin-related hexapeptides represents a new approach in the design of selective probes for {delta}-opioid receptors, displaying the appropriate criteria to investigate biological and pharmacological properties of the assumed binding site ({delta}) of endogenous enkephalins. The selectivities and high affinities of Tyr-D-Ser(O-t-Bu)-Gly-Phe-Leu-Thr(DSTBULET) and especially Tyr-D-Ser(O-t-Bu)Gly-Phe-Leu-Thr(O-t-Bu) (BUBU) associated with a satisfactory resistance to peptidases, make them the most suitable {delta}-probes reported to date. In the present paper, we report the synthesis of DSTBULET and BUBU under tritiated forms with high specific radioactivities. These radio-labelled probes will enable extensive in vitro and in vivo investigations of {delta}-opioid receptors properties to be carried out. (author).

  14. Opioid intoxication

    Science.gov (United States)

    ... easily result in intoxication. The provider prescribes a sleep medicine (sedative) in addition to the opioid. The provider ... an opioid with certain other drugs, such as sleep medicines or alcohol Taking the opioid in ways not ...

  15. Methods for quantifying T cell receptor binding affinities and thermodynamics

    Science.gov (United States)

    Piepenbrink, Kurt H.; Gloor, Brian E.; Armstrong, Kathryn M.; Baker, Brian M.

    2013-01-01

    αβ T cell receptors (TCRs) recognize peptide antigens bound and presented by class I or class II major histocompatibility complex (MHC) proteins. Recognition of a peptide/MHC complex is required for initiation and propagation of a cellular immune response, as well as the development and maintenance of the T cell repertoire. Here we discuss methods to quantify the affinities and thermodynamics of interactions between soluble ectodomains of TCRs and their peptide/MHC ligands, focusing on titration calorimetry, surface plasmon resonance, and fluorescence anisotropy. As TCRs typically bind ligand with weak-to-moderate affinities, we focus the discussion on means to enhance the accuracy and precision of low affinity measurements. In addition to further elucidating the biology of the T cell mediated immune response, more reliable low affinity measurements will aid with more probing studies with mutants or altered peptides that can help illuminate the physical underpinnings of how TCRs achieve their remarkable recognition properties. PMID:21609868

  16. Analysis of the hormone-binding domain of steroid receptors using chimeras generated by homologous recombination

    International Nuclear Information System (INIS)

    Martinez, Elisabeth D.; Pattabiraman, Nagarajan; Danielsen, Mark

    2005-01-01

    The glucocorticoid receptor and the mineralocorticoid receptor are members of the steroid receptor family that exhibit ligand cross-reactivity. Specificity of steroid receptor action is investigated in the present work by the construction and characterization of chimeras between the glucocorticoid receptor and the mineralocorticoid receptor. We used an innovative approach to make novel steroid receptor proteins in vivo that in general, contrary to our expectations, show increased ligand specificity compared to the parental receptors. We describe a receptor that is specific for the potent synthetic glucocorticoid triamcinolone acetonide and does not bind aldosterone. A further set of chimeras has an increased ability to discriminate between ligands, responding potently to mineralocorticoids and only very weakly to synthetic glucocorticoids. A chimera with the fusion site in the hinge highlights the importance of the region between the DNA-binding and the hormone-binding domains since, unlike both the glucocorticoid and mineralocorticoid receptors, it only responds to mineralocorticoids. One chimera has reduced specificity in that it acts as a general corticoid receptor, responding to glucocorticoids and mineralocorticoids with similar potency and efficacy. Our data suggest that regions of the glucocorticoid and mineralocorticoid receptor hormone-binding domains are functionally non-reciprocal. We present transcriptional, hormone-binding, and structure-modeling evidence that suggests that receptor-specific interactions within and across domains mediate aspects of specificity in transcriptional responses to steroids

  17. Nuclear thyroid hormone receptors in rabbit heart: reduced triiodothyronine binding in atrium compared with ventricle

    International Nuclear Information System (INIS)

    Banerjee, S.K.; Ulrich, J.M.; Kaldor, G.J.

    1988-01-01

    Radiolabeled triiodothyronine (T3) binding to isolated nuclei was measured to compare the binding characteristics of the nuclear receptors in rabbit ventricular and atrial muscle cells. Scatchard analysis of the binding data yielded a maximum binding capacity of 170 +/- 20 fmol per mg DNA and apparent dissociation constant of 525 +/- 100 pM for ventricular nuclei. The binding capacity and the dissociation constant for the atrial muscle cell nuclei were 55 +/- 10 fmol per mg DNA and 500 +/- 75 pM, respectively. The results suggest that the binding capacity for T3 receptor in the atrium is considerably lower than that found in the ventricle. The reduced binding capacity of the T3 receptor in the atrium might reflect differences in the nuclear T3 receptors between ventricle and atrium

  18. Synthesis and Pharmacology of Halogenated δ-Opioid-Selective [D-Ala2]Deltorphin II Peptide Analogues

    Science.gov (United States)

    Pescatore, Robyn; Marrone, Gina F.; Sedberry, Seth; Vinton, Daniel; Finkelstein, Netanel; Katlowitz, Yitzchak E.; Pasternak, Gavril W.; Wilson, Krista R.; Majumdar, Susruta

    2015-01-01

    Deltorphins are naturally occurring peptides produced by the skin of the giant monkey frog (Phyllomedusa bicolor). They are δ-opioid receptor-selective agonists. Herein, we report the design and synthesis of a peptide, Tyr-D-Ala-(pI)Phe-Glu-Ile-Ile-Gly-NH2 3 (GATE3-8), based on the [D-Ala2]deltorphin II template, which is δ-selective in in vitro radioligand binding assays over the μ- and κ-opioid receptors. It is a full agonist in [35S]GTPγS functional assays and analgesic when administered supraspinally to mice. Analgesia of 3 (GATE3-8) is blocked by the selective δ receptor antagonist naltrindole, indicating that the analgesic action of 3 is mediated by the δ-opioid receptor. We have established a radioligand in which 125I isincorporated into 3 (GATE3-8). The radioligand has a KD of 0.1 nM in Chinese hamster ovary (CHO) cells expressing the δ receptor. Additionally, a series of peptides based on 3 (GATE3-8) was synthesized by incorporating various halogens in the para position on the aromatic ring of Phe3. The peptides were characterized for binding affinity at the μ-, δ-, and κ-opioid receptors, which showed a linear correlation between binding affinity and the size of the halogen substituent. These peptides may be interesting tools for probing δ-opioid receptor pharmacology. PMID:25844930

  19. Synthesis and pharmacology of halogenated δ-opioid-selective [d-Ala(2)]deltorphin II peptide analogues.

    Science.gov (United States)

    Pescatore, Robyn; Marrone, Gina F; Sedberry, Seth; Vinton, Daniel; Finkelstein, Netanel; Katlowitz, Yitzchak E; Pasternak, Gavril W; Wilson, Krista R; Majumdar, Susruta

    2015-06-17

    Deltorphins are naturally occurring peptides produced by the skin of the giant monkey frog (Phyllomedusa bicolor). They are δ-opioid receptor-selective agonists. Herein, we report the design and synthesis of a peptide, Tyr-d-Ala-(pI)Phe-Glu-Ile-Ile-Gly-NH2 3 (GATE3-8), based on the [d-Ala(2)]deltorphin II template, which is δ-selective in in vitro radioligand binding assays over the μ- and κ-opioid receptors. It is a full agonist in [(35)S]GTPγS functional assays and analgesic when administered supraspinally to mice. Analgesia of 3 (GATE3-8) is blocked by the selective δ receptor antagonist naltrindole, indicating that the analgesic action of 3 is mediated by the δ-opioid receptor. We have established a radioligand in which (125)I is incorporated into 3 (GATE3-8). The radioligand has a KD of 0.1 nM in Chinese hamster ovary (CHO) cells expressing the δ receptor. Additionally, a series of peptides based on 3 (GATE3-8) was synthesized by incorporating various halogens in the para position on the aromatic ring of Phe(3). The peptides were characterized for binding affinity at the μ-, δ-, and κ-opioid receptors, which showed a linear correlation between binding affinity and the size of the halogen substituent. These peptides may be interesting tools for probing δ-opioid receptor pharmacology.

  20. Detection and Quantization of the Expression of Two mu-Opioid Receptor Splice Variants mRNA (hMOR-1A and hMOR-1O in Peripheral Blood Lymphocytes of Long-Term Abstinent Former Opioid Addicts

    Directory of Open Access Journals (Sweden)

    N Vousooghi, Pharm

    2012-05-01

    Full Text Available

    Background and Objectives

    The mu-Opioid receptor (MOR exerts a critical role on effects of opiodis. The objective of this study is to find a peripheral bio-marker in addiction studies through quantization of the expression of two MOR splice variants mRNA (hMOR-1A and hMOR-1O in peripheral blood lymphocytes (PBLs of long-term abstinent former opioids addicts.

    Methods

    In this case-control study, case and control people were male and divided in two groups: people who gave up addiction to opioids (case and healthy individuals without history of addiction (control. The mRNA expression in PBLs of participants was detected and measured by real-time Polymerase Chain Reaction (PCR using SYBR Green Dye.

    Results

    The hMOR-1A mRNA expression in PBLs of abstinent group was significantly reduced and reached to 0.33 of the control group (p<0.001. Similar results were obtained for the other splice variant with the mRNA expression of hMOR-1O in PBLs of abstinent group reaching to 0.38 of that of the control group (p < 0.001.

    Conclusion

    mRNA expression deficiency of two mu-opioid receptor splice variants, hMOR-1A and nMOR-1O, seams to be a risk factor making individuals vulnerable to drug addiction. Based on this analysis measuring the amount of mRNA expression of these two splice variants in PBLs can serve as a peripheral bio-marker for detecting people at risk.

  1. Dopamine D2/3- and μ-opioid receptor antagonists reduce cue-induced responding and reward impulsivity in humans.

    Science.gov (United States)

    Weber, S C; Beck-Schimmer, B; Kajdi, M-E; Müller, D; Tobler, P N; Quednow, B B

    2016-07-05

    Increased responding to drug-associated stimuli (cue reactivity) and an inability to tolerate delayed gratification (reward impulsivity) have been implicated in the development and maintenance of drug addiction. Whereas data from animal studies suggest that both the dopamine and opioid system are involved in these two reward-related processes, their role in humans is less clear. Moreover, dopaminergic and opioidergic drugs have not been directly compared with regard to these functions, even though a deeper understanding of the underlying mechanisms might inform the development of specific treatments for elevated cue reactivity and reward impulsivity. In a randomized, double-blind, between-subject design we administered the selective dopamine D2/D3 receptor antagonist amisulpride (400 mg, n=41), the unspecific opioid receptor antagonist naltrexone (50 mg, n=40) or placebo (n=40) to healthy humans and measured cue-induced responding with a Pavlovian-instrumental transfer task and reward impulsivity with a delay discounting task. Mood was assessed using a visual analogue scale. Compared with placebo, amisulpride significantly suppressed cue-induced responding and reward impulsivity. The effects of naltrexone were similar, although less pronounced. Both amisulpride and naltrexone decreased average mood ratings compared with placebo. Our results demonstrate that a selective blockade of dopamine D2/D3 receptors reduces cue-induced responding and reward impulsivity in healthy humans. Antagonizing μ-opioid receptors has similar effects for cue-induced responding and to a lesser extent for reward impulsivity.

  2. An examination of the effects of subthalamic nucleus inhibition or μ-opioid receptor stimulation on food-directed motivation in the non-deprived rat

    Science.gov (United States)

    Pratt, Wayne E.; Choi, Eugene; Guy, Elizabeth G.

    2012-01-01

    The subthalamic nucleus (STN) serves important functions in regulating movement, cognition, and motivation and is connected with cortical and basal ganglia circuits that process reward and reinforcement. In order to further examine the role of the STN on motivation toward food in non-deprived rats, these experiments studied the effects of pharmacological inhibition or μ-opioid receptor stimulation of the STN on the 2-hr intake of a sweetened fat diet, the amount of work exerted to earn sucrose on a progressive ratio 2 (PR-2) schedule of reinforcement, and performance on a differential reinforcement of low-rate responding (DRL) schedule for sucrose reward. Separate behavioral groups (N = 6–9) were tested following bilateral inhibition of the STN with the GABAA receptor agonist muscimol (at 0–5 ng/0.5 μl/side) or following μ-opioid receptor stimulation with the agonist D-Ala2, N-MePhe4, Gly-ol-enkephalin (DAMGO; at 0, 0.025 or 0.25 μg/0.5 μl/side). Although STN inhibition increased ambulatory behavior during 2-hr feeding sessions, it did not significantly alter intake of the sweetened fat diet. STN inhibition also did not affect the breakpoint for sucrose pellets during a 1-hr PR-2 reinforcement schedule or impact the number of reinforcers earned on a 1-hr DRL-20 sec reinforcement schedule in non-deprived rats. In contrast, STN μ-opioid receptor stimulation significantly increased feeding on the palatable diet and reduced the reinforcers earned on a DRL-20 schedule, although DAMGO microinfusions had no effect on PR-2 performance. These data suggest that STN inhibition does not enhance incentive motivation for food in the absence of food restriction and that STN μ-opioid receptors play an important and unique role in motivational processes. PMID:22391117

  3. Opioid/NMDA receptors blockade reverses the depressant-like behavior of foot shock stress in the mouse forced swimming test.

    Science.gov (United States)

    Haj-Mirzaian, Arya; Ostadhadi, Sattar; Kordjazy, Nastaran; Dehpour, Ahmad Reza; Ejtemaei Mehr, Shahram

    2014-07-15

    Opioid and glutamatergic receptors have a key role in depression following stress. In this study, we assessed opioid and glutamatergic receptors interaction with the depressant-like behavior of acute foot-shock stress in the mouse forced swimming test. Stress was induced by intermittent foot shock stimulation during 30min and swim periods were afterwards conducted by placing mice in separated glass cylinders filled with water for 6min. The immobility time during the last 4min of the test was considered. Acute foot-shock stress significantly increased the immobility time of mice compared to non-stressed control group (P≤0.01). Administration of non-selective opioid receptors antagonist, naltrexone (1 and 2mg/kg, i.p.), and the selective non-competitive NMDA receptor antagonist, MK-801 (0.05mg/kg, i.p.), and the selective serotonin reuptake inhibitor, fluoxetine (5mg/kg), significantly reduced the immobility time in stressed animals (P≤0.01). Lower doses of MK-801 (0.01mg/kg), naltrexone (0.3mg/kg), NMDA (75mg/kg) and morphine(5mg/kg) had no effect on foot-shock stressed mice. Combined treatment of sub-effective doses of naltrexone and MK-801 significantly showed an antidepressant-like effect (P≤0.001). On the other hand, co-administration of non-effective doses of NMDA and morphine with effective doses of naltrexone and MK-801 reversed the anti-immobility effect of these drugs. Taken together, we have for the first time demonstrated the possible role of opioid/NMDA receptors signaling in the depressant-like effect of foot-shock stress, and proposed the use of drugs that act like standard anti-depressants in stress-induced depression. Copyright © 2014. Published by Elsevier B.V.

  4. Examining the role of mu opioid receptor endocytosis in the beneficial and side-effects of prolonged opioid use: From a symposium on new concepts in mu-opioid pharmacology

    OpenAIRE

    Whistler, Jennifer L.

    2012-01-01

    Opioid drugs remain the gold standard for the treatment of severe pain, both acute/post-surgical and chronic. However, the utility of opioid drugs for the treatment of chronic pain is compromised by the development of analgesic tolerance which, in turn, leads to dose-escalation and increased likelihood of dangerous side effects, including dependence. Consequently, there remains resistance among clinicians and the general population to using opiates for pain management because of risk of “addi...

  5. Familial defective apolipoprotein B-100: low density lipoproteins with abnormal receptor binding

    International Nuclear Information System (INIS)

    Innerarity, T.L.; Weisgraber, K.H.; Arnold, K.S.; Mahley, R.W.; Krauss, R.M.; Vega, G.L.; Grundy, S.M.

    1987-01-01

    Previous in vivo turnover studies suggested that retarded clearance of low density lipoproteins (LDL) from the plasma of some hypercholesterolemic patients is due to LDL with defective receptor binding. The present study examined this postulate directly by receptor binding experiments. The LDL from a hypercholesterolemic patient (G.R.) displayed a reduced ability to bind to the LDL receptors on normal human fibroblasts. The G.R. LDL possessed 32% of normal receptor binding activity. Likewise, the G.R. LDL were much less effective than normal LDL in competing with 125 I-labeled normal LDL for cellular uptake and degradation and in stimulating intracellular cholesteryl ester synthesis. The defect in LDL binding appears to be due to a genetic abnormality of apolipoprotein B-100: two brothers of the proband possess LDL defective in receptor binding, whereas a third brother and the proband's son have normally binding LDL. Further, the defect in receptor binding does not appear to be associated wit an abnormal lipid composition or structure of the LDL. Normal and abnormal LDL subpopulations were partially separated from plasma of two subjects by density-gradient ultracentrifugation, a finding consistent with the presence of a normal and a mutant allele. The affected family members appear to be heterozygous for this disorder, which has been designated familial defective apolipoprotein B-100. These studies indicate that the defective receptor binding results in inefficient clearance of LDL and the hypercholesterolemia observed in these patients

  6. Neuronal low-density lipoprotein receptor-related protein 1 binds and endocytoses prion fibrils via receptor cluster 4

    DEFF Research Database (Denmark)

    Jen, Angela; Parkyn, Celia J; Mootoosamy, Roy C

    2010-01-01

    For infectious prion protein (designated PrP(Sc)) to act as a template to convert normal cellular protein (PrP(C)) to its distinctive pathogenic conformation, the two forms of prion protein (PrP) must interact closely. The neuronal receptor that rapidly endocytoses PrP(C) is the low......-density lipoprotein receptor-related protein 1 (LRP1). We show here that on sensory neurons LRP1 is also the receptor that binds and rapidly endocytoses smaller oligomeric forms of infectious prion fibrils, and recombinant PrP fibrils. Although LRP1 binds two molecules of most ligands independently to its receptor...... both prion and LRP1 biology....

  7. m-Trifluoromethyl-diphenyl diselenide promotes resilience to social avoidance induced by social defeat stress in mice: Contribution of opioid receptors and MAPKs.

    Science.gov (United States)

    Rosa, Suzan Gonçalves; Pesarico, Ana Paula; Nogueira, Cristina Wayne

    2018-03-02

    Depressive symptoms precipitated by stress are prevalent in population. In experimental models of social stress, endogenous opioids mediate different aspects of defensive and submissive behaviors. The present study investigated the opioid receptors, mitogen-activated protein kinase (MAPKs) and protein kinase B (Akt) contribution to m-trifluoromethyl-diphenyl diselenide [(m-CF 3 -PhSe) 2 ] effects on social avoidance induced by social defeat stress (SDS). Adult Swiss mice were subjected to SDS and treated with (m-CF 3 -PhSe) 2 (5 to 25mg/kg) for 7days. After that, the mice performed locomotor and social avoidance tests. The opioid receptors, MAPKs and Akt protein contents were determined in the prefrontal cortical samples of mice. Firstly, the mice were segregated in susceptible or resilient subpopulation based on their social avoidance induced by stress. (m-CF 3 -PhSe) 2 (25mg/kg) was effective against the stress-induced social avoidance and improved social interaction behavior in mice. SDS increased the μ and κ protein contents but reduced those of δ opioid receptors in susceptible mice. Resilient and (m-CF 3 -PhSe) 2 -treated mice had no alteration in the levels of opioid receptors. Moreover, (m-CF 3 -PhSe) 2 was effective against the increase of c-Jun N-terminal kinase (JNK) and the decrease of Akt phosphorylation protein contents induced by SDS in susceptible mice. The protein content of extracellular signal-regulated kinase (ERK) phosphorylation was reduced in both susceptible and resilient mice, whereas p38 mitogen-activated protein kinase (p38 MAPK) phosphorylation was increased only in resilient mice. (m-CF 3 -PhSe) 2 was partially effective against the pERK decrease and ineffective against the increase in p38 MAPK phosphorylation in mice subjected to SDS. These results suggest that the modulation of protein contents of opioid receptors, JNK and Akt phosphorylation is associated with resilience to SDS promoted by (m-CF 3 -PhSe) 2 in mice. Copyright

  8. Evidence for the involvement of MC4 receptors in the central mechanisms of opioid antinociception

    NARCIS (Netherlands)

    Starowicz, Katarzyna

    2005-01-01

    The data described in this thesis extend general knowledge of the involvement of the MC4 receptor in mechanisms of analgesia. The following aspects outlined below constitute novel information. Firstly, the MC4R localization in the DRG is demonstrated. The MC4 receptor was assumed to exist

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

    International Nuclear Information System (INIS)

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

    1987-01-01

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

  10. General, kappa, delta and mu opioid receptor antagonists mediate feeding elicited by the GABA-B agonist baclofen in the ventral tegmental area and nucleus accumbens shell in rats: reciprocal and regional interactions.

    Science.gov (United States)

    Miner, Patricia; Shimonova, Lyudmila; Khaimov, Arthur; Borukhova, Yaffa; Ilyayeva, Ester; Ranaldi, Robert; Bodnar, Richard J

    2012-03-14

    Food intake is significantly increased following administration of agonists of GABA and opioid receptors into the nucleus accumbens shell (NACs) and ventral tegmental area (VTA). GABA-A or GABA-B receptor antagonist pretreatment within the VTA or NACs differentially affects mu-opioid agonist-induced feeding elicited from the same site. Correspondingly, general or selective opioid receptor antagonist pretreatment within the VTA or NACs differentially affects GABA agonist-induced feeding elicited from the same site. Regional interactions have been evaluated in feeding studies by administering antagonists in one site prior to agonist administration in a second site. Thus, opioid antagonist-opioid agonist and GABA antagonist-GABA agonist feeding interactions have been identified between the VTA and NACs. However, pretreatment with GABA-A or GABA-B receptor antagonists in the VTA failed to affect mu opioid agonist-induced feeding elicited from the NACs, and correspondingly, these antagonists administered in the NACs failed to affect mu opioid-induced feeding elicited from the VTA. To evaluate whether regional and reciprocal VTA and NACs feeding interactions occur for opioid receptor modulation of GABA agonist-mediated feeding, the present study examined whether feeding elicited by the GABA-B agonist, baclofen microinjected into the NACs was dose-dependently blocked by pretreatment with general (naltrexone: NTX), mu (beta-funaltrexamine: BFNA), kappa (nor-binaltorphamine: NBNI) or delta (naltrindole: NTI) opioid antagonists in the VTA, and correspondingly, whether VTA baclofen-induced feeding was dose-dependently blocked by NACs pretreatment with NTX, BFNA, NBNI or NTI in rats. Bilateral pairs of cannulae aimed at the VTA and NACs were stereotaxically implanted in rats, and their food intakes were assessed following vehicle and baclofen (200 ng) in each site. Baclofen produced similar magnitudes of increased food intake following VTA and NACs treatment. Baclofen

  11. Opioid system and human emotions.

    Science.gov (United States)

    Nummenmaa, Lauri; Tuominen, Lauri

    2017-04-10

    Emotions are states of vigilant readiness that guide human and animal behaviour during survival-salient situations. Categorical models of emotions posit neurally and physiologically distinct basic human emotions (anger, fear, disgust, happiness, sadness and surprise) that govern different survival functions. Opioid receptors are expressed abundantly in the mammalian emotion circuit, and the opioid system modulates a variety of functions related to arousal and motivation. Yet, its specific contribution to different basic emotions has remained poorly understood. Here, we review how the endogenous opioid system and particularly the μ receptor contribute to emotional processing in humans. Activation of the endogenous opioid system is consistently associated with both pleasant and unpleasant emotions. In general, exogenous opioid agonists facilitate approach-oriented emotions (anger, pleasure) and inhibit avoidance-oriented emotions (fear, sadness). Opioids also modulate social bonding and affiliative behaviour, and prolonged opioid abuse may render both social bonding and emotion recognition circuits dysfunctional. However, there is no clear evidence that the opioid system is able to affect the emotions associated with surprise and disgust. Taken together, the opioid systems contribute to a wide array of positive and negative emotions through their general ability to modulate the approach versus avoidance motivation associated with specific emotions. Because of the protective effects of opioid system-mediated prosociality and positive mood, the opioid system may constitute an important factor contributing to psychological and psychosomatic resilience. © 2017 The British Pharmacological Society.

  12. Nicotine and endogenous opioids: neurochemical and pharmacological evidence.

    Science.gov (United States)

    Hadjiconstantinou, Maria; Neff, Norton H

    2011-06-01

    Although the mesolimbic dopamine hypothesis is the most influential theory of nicotine reward and reinforcement, there has been a consensus that other neurotransmitter systems contribute to the addictive properties of nicotine as well. In this regard, the brain opioidergic system is of interest. Striatum is rich in opioid peptides and opioid receptors, and striatal opioidergic neurons are engaged in a bidirectional communication with midbrain dopaminergic neurons, closely regulating each other's activity. Enkephalins and dynorphins exert opposing actions on dopaminergic neurons, increasing and decreasing dopamine release respectively, and are components of circuits promoting positive or negative motivational and affective states. Moreover, dopamine controls the synthesis of striatal enkephalins and dynorphins. Evidence suggests that opioidergic function is altered after nicotine and endogenous opioids are involved in nicotine's behavioral effects. 1) The synthesis and release of β-endorphin, met-enkephalin and dynorphin in brain, especially nucleus accumbens (NAc), are altered after acute or chronic nicotine treatment and during nicotine withdrawal. 2) Although opioid receptor binding and mRNA do not appear to change in the striatum during nicotine withdrawal, the activity of κ-opioid (KOPr) and δ-opioid (DOPr) receptors is attenuated in NAc. 3) The nicotine withdrawal syndrome reminisces that of opiates, and naloxone precipitates some of its somatic, motivational, and affective signs. 4) Genetic and pharmacological studies indicate that μ-opioid (MOPr) receptors are mainly involved in nicotine reward, while DOPrs contribute to the emotional and KOPrs to the aversive responses of nicotine. 5) Finally, MOPrs and enkephalin, but not β-endorphin or dynorphin, are necessary for the physical manifestations of nicotine withdrawal. This article is part of a Special Issue entitled 'Trends in neuropharmacology: in memory of Erminio Costa'. Copyright © 2010 Elsevier

  13. Characterisation of the human NMDA receptor subunit NR3A glycine binding site

    DEFF Research Database (Denmark)

    Nilsson, A; Duan, J; Mo-Boquist, L-L

    2007-01-01

    In this study, we characterise the binding site of the human N-methyl-d-aspartate (NMDA) receptor subunit NR3A. Saturation radioligand binding of the NMDA receptor agonists [(3)H]-glycine and [(3)H]-glutamate showed that only glycine binds to human NR3A (hNR3A) with high affinity (K(d)=535nM (277...

  14. Chronic suppression of μ-opioid receptor signaling in the nucleus accumbens attenuates development of diet-induced obesity in rats.

    Science.gov (United States)

    Lenard, N R; Zheng, H; Berthoud, H-R

    2010-06-01

    To test the hypothesis that micro-opioid receptor signaling in the nucleus accumbens contributes to hedonic (over)eating and obesity. To investigate the effects of chronic micro-opioid antagonism in the nucleus accumbens core or shell on intake of a palatable diet, and the development of diet-induced obesity in rats. Chronic blockade of micro-opioid receptor signaling in the nucleus accumbens core or shell was achieved by means of repeated injections (every 4-5 days) of the irreversible receptor antagonist beta-funaltrexamine (BFNA) over 3-5 weeks. The diet consisted of either a choice of high-fat chow, chocolate-flavored Ensure and regular chow (each nutritionally complete) or regular chow only. Intake of each food item, body weight and body fat mass were monitored throughout the study. The BFNA injections aimed at either the core or shell of the nucleus accumbens resulted in significantly attenuated intake of palatable diet, body weight gain and fat accretion, compared with vehicle control injections. The injection of BFNA in the core did not significantly change these parameters in chow-fed control rats. The injection of BFNA in the core and shell differentially affected intake of the two palatable food items: in the core, BFNA significantly reduced the intake of high-fat, but not of Ensure, whereas in the shell, it significantly reduced the intake of Ensure, but not of high-fat, compared with vehicle treatment. Endogenous micro-opioid receptor signaling in the nucleus accumbens core and shell is necessary for palatable diet-induced hyperphagia and obesity to fully develop in rats. Sweet and non-sweet fatty foods may be differentially processed in subcomponents of the ventral striatum.

  15. Binding modes of dihydroquinoxalinones in a homology model of bradykinin receptor 1.

    Science.gov (United States)

    Ha, Sookhee N; Hey, Pat J; Ransom, Rick W; Harrell, C Meacham; Murphy, Kathryn L; Chang, Ray; Chen, Tsing-Bau; Su, Dai-Shi; Markowitz, M Kristine; Bock, Mark G; Freidinger, Roger M; Hess, Fred J

    2005-05-27

    We report the first homology model of human bradykinin receptor B1 generated from the crystal structure of bovine rhodopsin as a template. Using an automated docking procedure, two B1 receptor antagonists of the dihydroquinoxalinone structural class were docked into the receptor model. Site-directed mutagenesis data of the amino acid residues in TM1, TM3, TM6, and TM7 were incorporated to place the compounds in the binding site of the homology model of the human B1 bradykinin receptor. The best pose in agreement with the mutation data was selected for detailed study of the receptor-antagonist interaction. To test the model, the calculated antagonist-receptor binding energy was correlated with the experimentally measured binding affinity (K(i)) for nine dihydroquinoxalinone analogs. The model was used to gain insight into the molecular mechanism for receptor function and to optimize the dihydroquinoxalinone analogs.

  16. Structure-based Understanding of Binding Affinity and Mode of Estrogen Receptor α Agonists and Antagonists.

    Science.gov (United States)

    The flexible hydrophobic ligand binding pocket (LBP) of estrogen receptor α (ERα) allows the binding of a wide variety of endocrine disruptors. Upon ligand binding, the LBP reshapes around the contours of the ligand and stabilizes the complex by complementary hydrophobic interact...

  17. Combined sodium ion sensitivity in agonist binding and internalization of vasopressin V1b receptors.

    Science.gov (United States)

    Koshimizu, Taka-Aki; Kashiwazaki, Aki; Taniguchi, Junichi

    2016-05-03

    Reducing Na(+) in the extracellular environment may lead to two beneficial effects for increasing agonist binding to cell surface G-protein coupled receptors (GPCRs): reduction of Na(+)-mediated binding block and reduce of receptor internalization. However, such combined effects have not been explored. We used Chinese Hamster Ovary cells expressing vasopressin V1b receptors as a model to explore Na(+) sensitivity in agonist binding and receptor internalization. Under basal conditions, a large fraction of V1b receptors is located intracellularly, and a small fraction is in the plasma membrane. Decreases in external Na(+) increased cell surface [(3)H]AVP binding and decreased receptor internalization. Substitution of Na(+) by Cs(+) or NH4(+) inhibited agonist binding. To suppress receptor internalization, the concentration of NaCl, but not of CsCl, had to be less than 50 mM, due to the high sensitivity of the internalization machinery to Na(+) over Cs(+). Iso-osmotic supplementation of glucose or NH4Cl maintained internalization of the V1b receptor, even in a low-NaCl environment. Moreover, iodide ions, which acted as a counter anion, inhibited V1b agonist binding. In summary, we found external ionic conditions that could increase the presence of high-affinity state receptors at the cell surface with minimum internalization during agonist stimulations.

  18. Effects of Chronic Social Defeat Stress on Sleep and Circadian Rhythms Are Mitigated by Kappa-Opioid Receptor Antagonism.

    Science.gov (United States)

    Wells, Audrey M; Ridener, Elysia; Bourbonais, Clinton A; Kim, Woori; Pantazopoulos, Harry; Carroll, F Ivy; Kim, Kwang-Soo; Cohen, Bruce M; Carlezon, William A

    2017-08-09

    Stress plays a critical role in the neurobiology of mood and anxiety disorders. Sleep and circadian rhythms are affected in many of these conditions. Here we examined the effects of chronic social defeat stress (CSDS), an ethological form of stress, on sleep and circadian rhythms. We exposed male mice implanted with wireless telemetry transmitters to a 10 day CSDS regimen known to produce anhedonia (a depressive-like effect) and social avoidance (an anxiety-like effect). EEG, EMG, body temperature, and locomotor activity data were collected continuously during the CSDS regimen and a 5 day recovery period. CSDS affected numerous endpoints, including paradoxical sleep (PS) and slow-wave sleep (SWS), as well as the circadian rhythmicity of body temperature and locomotor activity. The magnitude of the effects increased with repeated stress, and some changes (PS bouts, SWS time, body temperature, locomotor activity) persisted after the CSDS regimen had ended. CSDS also altered mRNA levels of the circadian rhythm-related gene mPer2 within brain areas that regulate motivation and emotion. Administration of the κ-opioid receptor (KOR) antagonist JDTic (30 mg/kg, i.p.) before CSDS reduced stress effects on both sleep and circadian rhythms, or hastened their recovery, and attenuated changes in mPer2 Our findings show that CSDS produces persistent disruptions in sleep and circadian rhythmicity, mimicking attributes of stress-related conditions as they appear in humans. The ability of KOR antagonists to mitigate these disruptions is consistent with previously reported antistress effects. Studying homologous endpoints across species may facilitate the development of improved treatments for psychiatric illness. SIGNIFICANCE STATEMENT Stress plays a critical role in the neurobiology of mood and anxiety disorders. We show that chronic social defeat stress in mice produces progressive alterations in sleep and circadian rhythms that resemble features of depression as it appears in

  19. In vitro binding and receptor-mediated activity of terlipressin at vasopressin receptors V1 and V2.

    Science.gov (United States)

    Jamil, Khurram; Pappas, Stephen Chris; Devarakonda, Krishna R

    2018-01-01

    Terlipressin, a synthetic, systemic vasoconstrictor with selective activity at vasopressin-1 (V 1 ) receptors, is a pro-drug for the endogenous/natural porcine hormone [Lys 8 ]-vasopressin (LVP). We investigated binding and receptor-mediated cellular activities of terlipressin, LVP, and endogenous human hormone [Arg 8 ]-vasopressin (AVP) at V 1 and vasopressin-2 (V 2 ) receptors. Cell membrane homogenates of Chinese hamster ovary cells expressing human V 1 and V 2 receptors were used in competitive binding assays to measure receptor-binding activity. These cells were used in functional assays to measure receptor-mediated cellular activity of terlipressin, LVP, and AVP. Binding was measured by [ 3 H]AVP counts, and the activity was measured by fluorometric detection of intracellular calcium mobilization (V 1 ) and cyclic adenosine monophosphate (V 2 ). Binding potency at V 1 and V 2 was AVP>LVP>terlipressin. LVP and terlipressin had approximately sixfold higher affinity for V 1 than for V 2 . Cellular activity potency was also AVP>LVP>terlipressin. Terlipressin was a partial agonist at V 1 and a full agonist at V 2 ; LVP was a full agonist at both V 1 and V 2 . The in vivo response to terlipressin is likely due to the partial V 1 agonist activity of terlipressin and full V 1 agonist activity of its metabolite, LVP. These results provide supportive evidence for previous findings and further establish terlipressin pharmacology for vasopressin receptors.

  20. μ-opioid modulation of HIV-1 coreceptor expressionand HIV-1 replication

    International Nuclear Information System (INIS)

    Steele, Amber D.; Henderson, Earl E.; Rogers, Thomas J.

    2003-01-01

    A substantial proportion of HIV-1-infected individuals are intravenous drug users (IVDUs) who abuse opiates. Opioids induce a number of immunomodulatory effects that may directly influence HIV-1 disease progression. In the present report, we have investigated the effect of opioids on the expression of the major HIV-1 coreceptors CXCR4 and CCR5. For these studies we have focused on opiates which are ligands for the μ-opioid receptor. Our results show that DAMGO, a selective μ-opioid agonist, increases CXCR4 and CCR5 expression in both CD3 + lymphoblasts and CD14 + monocytes three- to fivefold. Furthermore, DAMGO-induced elevation of HIV-1 coreceptor expression translates into enhanced replication of both X4 and R5 viral strains of HIV-1. We have confirmed the role of the μ-opioid receptor based on the ability of a μ-opioid receptor-selective antagonist to block the effects of DAMGO. We have also found that morphine enhances CXCR4 and CCR5 expression and subsequently increases both X4 and R5 HIV-1 infection. We suggest that the capacity of μ-opioids to increase HIV-1 coreceptor expression and replication may promote viral binding, trafficking of HIV-1-infected cells, and enhanced disease progression

  1. Nanodiscs for immobilization of lipid bilayers and membrane receptors: kinetic analysis of cholera toxin binding to a glycolipid receptor

    DEFF Research Database (Denmark)

    Borch, Jonas; Torta, Federico; Sligar, Stephen G

    2008-01-01

    nanodiscs and their incorporated membrane receptors can be attached to surface plasmon resonance sensorchips and used to measure the kinetics of the interaction between soluble molecules and membrane receptors inserted in the bilayer of nanodiscs. Cholera toxin and its glycolipid receptor G(M1) constitute...... a system that can be considered a paradigm for interactions of soluble proteins with membrane receptors. In this work, we have investigated different technologies for capturing nanodiscs containing the glycolipid receptor G(M1) in lipid bilayers, enabling measurements of binding of its soluble interaction...

  2. Connections between EM2-containing terminals and GABA/μ-opioid receptor co-expressing neurons in the rat spinal trigeminal caudal nucleus

    Science.gov (United States)

    Li, Meng-Ying; Wu, Zhen-Yu; Lu, Ya-Cheng; Yin, Jun-Bin; Wang, Jian; Zhang, Ting; Dong, Yu-Lin; Wang, Feng

    2014-01-01

    Endomorphin-2 (EM2) demonstrates a potent antinociceptive effect via the μ-opioid receptor (MOR). To provide morphological evidence for the pain control effect of EM2, the synaptic connections between EM2-immunoreactive (IR) axonal terminals and γ-amino butyric acid (GABA)/MOR co-expressing neurons in lamina II of the spinal trigeminal caudal nucleus (Vc) were investigated in the rat. Dense EM2-, MOR- and GABA-IR fibers and terminals were mainly observed in lamina II of the Vc. Within lamina II, GABA- and MOR-neuronal cell bodies were also encountered. The results of immunofluorescent histochemical triple-staining showed that approximately 14.2 or 18.9% of GABA-IR or MOR-IR neurons also showed MOR- or GABA-immunopositive staining in lamina II; approximately 45.2 and 36.1% of the GABA-IR and MOR-IR neurons, respectively, expressed FOS protein in their nuclei induced by injecting formalin into the left lower lip of the mouth. Most of the GABA/MOR, GABA/FOS, and MOR/FOS double-labeled neurons made close contacts with EM2-IR fibers and terminals. Immuno-electron microscopy confirmed that the EM2-IR terminals formed synapses with GABA-IR or MOR-IR dendritic processes and neuronal cell bodies in lamina II of the Vc. These results suggest that EM2 might participate in pain transmission and modulation by binding to MOR-IR and GABAergic inhibitory interneuron in lamina II of the Vc to exert inhibitory effect on the excitatory interneuron in lamina II and projection neurons in laminae I and III. PMID:25386121

  3. Suppressed Fat Appetite after Roux-en-Y Gastric Bypass Surgery Associates with Reduced Brain μ-opioid Receptor Availability in Diet-Induced Obese Male Rats.

    Science.gov (United States)

    Hankir, Mohammed K; Patt, Marianne; Patt, Jörg T W; Becker, Georg A; Rullmann, Michael; Kranz, Mathias; Deuther-Conrad, Winnie; Schischke, Kristin; Seyfried, Florian; Brust, Peter; Hesse, Swen; Sabri, Osama; Krügel, Ute; Fenske, Wiebke K

    2016-01-01

    Brain μ-opioid receptors (MORs) stimulate high-fat (HF) feeding and have been implicated in the distinct long term outcomes on body weight of bariatric surgery and dieting. Whether alterations in fat appetite specifically following these disparate weight loss interventions relate to changes in brain MOR signaling is unknown. To address this issue, diet-induced obese male rats underwent either Roux-en-Y gastric bypass (RYGB) or sham surgeries. Postoperatively, animals were placed on a two-choice diet consisting of low-fat (LF) and HF food and sham-operated rats were further split into ad libitum fed (Sham-LF/HF) and body weight-matched (Sham-BWM) to RYGB groups. An additional set of sham-operated rats always only on a LF diet (Sham-LF) served as lean controls, making four experimental groups in total. Corresponding to a stage of weight loss maintenance for RYGB rats, two-bottle fat preference tests in conjunction with small-animal positron emission tomography (PET) imaging studies with the selective MOR radioligand [ 11 C]carfentanil were performed. Brains were subsequently collected and MOR protein levels in the hypothalamus, striatum, prefrontal cortex and orbitofrontal cortex were analyzed by Western Blot. We found that only the RYGB group presented with intervention-specific changes: having markedly suppressed intake and preference for high concentration fat emulsions, a widespread reduction in [ 11 C]carfentanil binding potential (reflecting MOR availability) in various brain regions, and a downregulation of striatal and prefrontal MOR protein levels compared to the remaining groups. These findings suggest that the suppressed fat appetite caused by RYGB surgery is due to reduced brain MOR signaling, which may contribute to sustained weight loss unlike the case for dieting.

  4. Definition of the G protein-coupled receptor transmembrane bundle binding pocket and calculation of receptor similarities for drug design

    DEFF Research Database (Denmark)

    Gloriam, David Erik Immanuel; Foord, Steven M; Blaney, Frank E

    2009-01-01

    currently available crystal structures. This was used to characterize pharmacological relationships of Family A/Rhodopsin family GPCRs, minimizing evolutionary influence from parts of the receptor that do not generally affect ligand binding. The resultant dendogram tended to group receptors according...

  5. Differences in both glycosylation and binding properties between rat and mouse liver prolactin receptors.

    Science.gov (United States)

    Lascols, O; Cherqui, G; Munier, A; Picard, J; Capeau, J

    1994-05-01

    To investigate whether glycanic chains of prolactin receptors (PRL-R) play a role in hormone binding activity, comparison was made of rat and mouse liver solubilized receptors with respect to both their affinity for the hormone and their glycosylation properties. As compared with rat receptors, mouse receptors exhibited a 2-fold higher affinity for human growth hormone (hGH), the hormone being bound by both tissues with a lactogenic specificity. Along with this increased affinity, mouse receptors had a 2 lower M(r) relative to rat receptors (62 kDa versus 64 kDa as measured on hGH cross-linked receptors). These differences could be ascribed to different glycosylation properties of the receptors from the two species, as supported by the followings. 1) After treatment with endoglycosidase F (endo F), rat and mouse PRL-R no longer exhibited any difference in their M(r) (54 kDa for both cross-linked receptors). 2) Neuraminidase treatment increased by 37% the binding of hGH to mouse receptors, but was ineffective on the hormone-binding to rat receptors. Conversely, wheat germ agglutinin (WGA), another sialic acid specific probe, decreased hGH binding to rat receptors by 25%, but had no effect on this process for mouse ones. 3) Marked differences were observed in the recoveries of rat and mouse hormone-receptor (HR) complexes from ricin-1- (RCA1-), concanavalin A- (ConA-) and WGA-immobilized lectins. These differences were reduced (RCA1 and ConA) or abolished (WGA) after rat and mouse receptor desialylation by neuraminidase, a treatment which decreased the M(r) of both receptors by 2 kDa. Taken together, these results strongly suggest that the PRL-R from rat and mouse liver contain biantennary N-linked oligosaccharidic chains with distinct type of sialylation, which may account for their differential hormone-binding affinities.

  6. A Novel Approach for Effectively Treating SCI Pain, Improving Opioid Efficacy, and Preventing Opioid-Induced Constipation: Key Role of Toll-Like Receptor 4 (TLR4)

    Science.gov (United States)

    2016-12-01

    tested whether performing midline 150-kDyn contusion SCI at T9 with 1 s dwell elicited more robust pain symptoms. For locomotor recovery (BBB scale...addition, we tested the hypotheses that acute morphine would worsen locomotor recovery and chronic neuropathic pain, and that acute (+)-naltrexone could...This proposal will test a clinically relevant therapeutic, (+)-naltrexone, that we predict will improve the efficacy of opioids for controlling SCI

  7. A Novel Approach for Effectively Treating SCI Pain, Improving Opioid Efficacy, and Preventing Opioid Induced Constipation: Key Role of Toll-Like Receptor 4 (TLR4)

    Science.gov (United States)

    2016-10-01

    tested whether performing midline 150-kDyn contusion SCI at T9 with 1 s dwell elicited more robust pain symptoms. For locomotor recovery (BBB scale...addition, we tested the hypotheses that acute morphine would worsen locomotor recovery and chronic neuropathic pain, and that acute (+)-naltrexone could...This proposal will test a clinically relevant therapeutic, (+)-naltrexone, that we predict will improve the efficacy of opioids for controlling SCI

  8. NOpiates: Novel Dual Action Neuronal Nitric Oxide Synthase Inhibitors with μ-Opioid Agonist Activity.

    Science.gov (United States)

    Renton, Paul; Green, Brenda; Maddaford, Shawn; Rakhit, Suman; Andrews, John S

    2012-03-08

    A novel series of benzimidazole designed multiple ligands (DMLs) with activity at the neuronal nitric oxide synthase (nNOS) enzyme and the μ-opioid receptor was developed. Targeting of the structurally dissimilar heme-containing enzyme and the μ-opioid GPCR was predicated on the modulatory role of nitric oxide on μ-opioid receptor function. Structure-activity relationship studies yielded lead compound 24 with excellent nNOS inhibitory activity (IC50 = 0.44 μM), selectivity over both endothelial nitric oxide synthase (10-fold) and inducible nitric oxide synthase (125-fold), and potent μ-opioid binding affinity, K i = 5.4 nM. The functional activity as measured in the cyclic adenosine monosphospate secondary messenger assay resulted in full agonist activity (EC50 = 0.34 μM). This work represents a novel approach in the development of new analgesics for the treatment of pain.

  9. Neurotensin receptor binding levels in basal ganglia are not altered in Huntington's chorea or schizophrenia

    International Nuclear Information System (INIS)

    Palacios, J.M.; Chinaglia, G.; Rigo, M.; Ulrich, J.; Probst, A.

    1991-01-01

    Autoradiographic techniques were used to examine the distribution and levels of neurotensin receptor binding sites in the basal ganglia and related regions of the human brain. Monoiodo ( 125 I-Tyr3)neurotensin was used as a ligand. High amounts of neurotensin receptor binding sites were found in the substantia nigra pars compacta. Lower but significant quantities of neurotensin receptor binding sites characterized the caudate, putamen, and nucleus accumbens, while very low quantities were seen in both medial and lateral segments of the globus pallidus. In Huntington's chorea, the levels of neurotensin receptor binding sites were found to be comparable to those of control cases. Only slight but not statistically significant decreases in amounts of receptor binding sites were detected in the dorsal part of the head and in the body of caudate nucleus. No alterations in the levels of neurotensin receptor binding sites were observed in the substantia nigra pars compacta and reticulata. These results suggest that a large proportion of neurotensin receptor binding sites in the basal ganglia are located on intrinsic neurons and on extrinsic afferent fibers that do not degenerate in Huntington's disease

  10. Unsaturated free fatty acids increase benzodiazepine receptor agonist binding depending on the subunit composition of the GABAA receptor complex.

    Science.gov (United States)

    Witt, M R; Westh-Hansen, S E; Rasmussen, P B; Hastrup, S; Nielsen, M

    1996-11-01

    It has been shown previously that unsaturated free fatty acids (FFAs) strongly enhance the binding of agonist benzodiazepine receptor ligands and GABAA receptor ligands in the CNS in vitro. To investigate the selectivity of this effect, recombinant human GABAA/benzodiazepine receptor complexes formed by different subunit compositions (alpha x beta y gamma 2, x = 1, 2, 3, and 5; y = 1, 2, and 3) were expressed using the baculovirus-transfected Sf9 insect cell system. At 10(-4) M, unsaturated FFAs, particularly arachidonic (20:4) and docosahexaenoic (22:6) acids, strongly stimulated (> 200% of control values) the binding of [3H]flunitrazepam ([3H]FNM) to the alpha 3 beta 2 gamma 2 receptor combination in whole cell preparations. No effect or small increases in levels of unsaturated FFAs on [3H]FNM binding to alpha 1 beta x gamma 2 and alpha 2 beta x gamma 2 receptor combinations were observed, and weak effects (130% of control values) were detected using the alpha 5 beta 2 gamma 2 receptor combination. The saturated FFAs, stearic and palmitic acids, were without effect on [3H]FNM binding to any combination of receptor complexes. The hydroxylated unsaturated FFAs, ricinoleic and ricinelaidic acids, were shown to decrease the binding of [3H]FNM only if an alpha 1 beta 2 gamma 2 receptor combination was used. Given the heterogeneity of the GABAA/ benzodiazepine receptor subunit distribution in the CNS, the effects of FFAs on the benzodiazepine receptor can be assumed to vary at both cellular and regional levels.

  11. Hypothalamic kappa opioid receptor mediates both diet-induced and melanin concentrating hormone-induced liver damage through inflammation and endoplasmic reticulum stress.

    Science.gov (United States)

    Imbernon, Monica; Sanchez-Rebordelo, Estrella; Romero-Picó, Amparo; Kalló, Imre; Chee, Melissa J; Porteiro, Begoña; Al-Massadi, Omar; Contreras, Cristina; Fernø, Johan; Senra, Ana; Gallego, Rosalia; Folgueira, Cintia; Seoane, Luisa M; van Gestel, Margriet; Adan, Roger A; Liposits, Zsolt; Dieguez, Carlos; López, Miguel; Nogueiras, Ruben

    2016-10-01

    The opioid system is widely known to modulate the brain reward system and thus affect the behavior of humans and other animals, including feeding. We hypothesized that the hypothalamic opioid system might also control energy metabolism in peripheral tissues. Mice lacking the kappa opioid receptor (κOR) and adenoviral vectors overexpressing or silencing κOR were stereotaxically delivered in the lateral hypothalamic area (LHA) of rats. Vagal denervation was performed to assess its effect on liver metabolism. Endoplasmic reticulum (ER) stress was inhibited by pharmacological (tauroursodeoxycholic acid) and genetic (overexpression of the chaperone glucose-regulated protein 78 kDa) approaches. The peripheral effects on lipid metabolism were assessed by histological techniques and western blot. We show that in the LHA κOR directly controls hepatic lipid metabolism through the parasympathetic nervous system, independent of changes in food intake and body weight. κOR colocalizes with melanin concentrating hormone receptor 1 (MCH-R1) in the LHA, and genetic disruption of κOR reduced melanin concentrating hormone-induced liver steatosis. The functional relevance of these findings was given by the fact that silencing of κOR in the LHA attenuated both methionine choline-deficient, diet-induced and choline-deficient, high-fat diet-induced ER stress, inflammation, steatohepatitis, and fibrosis, whereas overexpression of κOR in this area promoted liver steatosis. Overexpression of glucose-regulated protein 78 kDa in the liver abolished hypothalamic κOR-induced steatosis by reducing hepatic ER stress. This study reveals a novel hypothalamic-parasympathetic circuit modulating hepatic function through inflammation and ER stress independent of changes in food intake or body weight; these findings might have implications for the clinical use of opioid receptor antagonists. (Hepatology 2016;64:1086-1104). © 2016 The Authors. (Hepatology published by Wiley Periodicals, Inc., on

  12. BOLD Imaging in Awake Wild-Type and Mu-Opioid Receptor Knock-Out Mice Reveals On-Target Activation Maps in Response to Oxycodone

    Directory of Open Access Journals (Sweden)

    Kelsey Moore

    2016-11-01

    Full Text Available Blood oxygen level dependent (BOLD imaging in awake mice was used to identify differences in brain activity between wild-type, and Mu (µ opioid receptor knock-outs (MuKO in response to oxycodone (OXY. Using a segmented, annotated MRI mouse atlas and computational analysis, patterns of integrated positive and negative BOLD activity were identified across 122 brain areas. The pattern of positive BOLD showed enhanced activation across the brain in WT mice within 15 min of intraperitoneal administration of 2.5 mg of OXY. BOLD activation was detected in 72 regions out of 122, and was most prominent in areas of high µ opioid receptor density (thalamus, ventral tegmental area, substantia nigra, caudate putamen, basal amygdala and hypothalamus, and focus on pain circuits indicated strong activation in major pain processing centers (central amygdala, solitary tract, parabrachial area, insular cortex, gigantocellularis area, ventral thalamus primary sensory cortex and prelimbic cortex. Importantly, the OXY-induced positive BOLD was eliminated in MuKO mice in most regions, with few exceptions (some cerebellar nuclei, CA3 of the hippocampus, medial amygdala and preoptic areas. This result indicates that most effects of OXY on positive BOLD are mediated by the µ opioid receptor (on-target effects. OXY also caused an increase in negative BOLD in WT mice in few regions (16 out of 122 and, unlike the positive BOLD response the negative BOLD was only partially eliminated in the MuKO mice (cerebellum, and in some case intensified (hippocampus. Negative BOLD analysis therefore shows activation and deactivation events in the absence of the µ receptor for some areas where receptor expression is normally extremely low or absent (off-target effects. Together, our approach permits establishing opioid-induced BOLD activation maps in awake mice. In addition, comparison of WT and MuKO mutant mice reveals both on-target and off-target activation events, and set an OXY

  13. Molecular characterization of the receptor binding structure-activity relationships of influenza B virus hemagglutinin.

    Science.gov (United States)

    Carbone, V; Kim, H; Huang, J X; Baker, M A; Ong, C; Cooper, M A; Li, J; Rockman, S; Velkov, T

    2013-01-01

    Selectivity of α2,6-linked human-like receptors by B hemagglutinin (HA) is yet to be fully understood. This study integrates binding data with structure-recognition models to examine the impact of regional-specific sequence variations within the receptor-binding pocket on selectivity and structure activity relationships (SAR). The receptor-binding selectivity of influenza B HAs corresponding to either B/Victoria/2/1987 or the B/Yamagata/16/88 lineages was examined using surface plasmon resonance, solid-phase ELISA and gel-capture assays. Our SAR data showed that the presence of asialyl sugar units is the main determinant of receptor preference of α2,6 versus α2,3 receptor binding. Changes to the type of sialyl-glycan linkage present on receptors exhibit only a minor effect upon binding affinity. Homology-based structural models revealed that structural properties within the HA pocket, such as a glyco-conjugate at Asn194 on the 190-helix, sterically interfere with binding to avian receptor analogs by blocking the exit path of the asialyl sugars. Similarly, naturally occurring substitutions in the C-terminal region of the 190-helix and near the N-terminal end of the 140-loop narrows the horizontal borders of the binding pocket, which restricts access of the avian receptor analog LSTa. This study helps bridge the gap between ligand structure and receptor recognition for influenza B HA; and provides a consensus SAR model for the binding of human and avian receptor analogs to influenza B HA.

  14. Libidibia ferrea mature seeds promote antinociceptive effect by peripheral and central pathway: possible involvement of opioid and cholinergic receptors.

    Science.gov (United States)

    Sawada, Luis Armando; Monteiro, Vanessa Sâmia da Conçeição; Rabelo, Guilherme Rodrigues; Dias, Germana Bueno; Da Cunha, Maura; do Nascimento, José Luiz Martins; Bastos, Gilmara de Nazareth Tavares

    2014-01-01

    Libidibia ferrea (LF) is a medicinal plant that holds many pharmacological properties. We evaluated the antinociceptive effect in the LF aqueous seed extract and Lipidic Portion of Libidibia ferrea (LPLF), partially elucidating their mechanisms. Histochemical tests and Gas chromatography of the LPLF were performed to characterize its fatty acids. Acetic acid-induced abdominal constriction, formalin-induced pain, and hot-plate test in mice were employed in the study. In all experiments, aqueous extract or LPLF was administered systemically at the doses of 1, 5, and 10 mg/kg. LF aqueous seed extract and LPLF demonstrated a dose-dependent antinociceptive effect in all tests indicating both peripheral anti-inflammatory and central analgesia properties. Also, the use of atropine (5 mg/kg), naloxone (5 mg/kg) in the abdominal writhing test was able to reverse the antinociceptive effect of the LPLF, indicating that at least one of LF lipids components is responsible for the dose related antinociceptive action in chemical and thermal models of nociception in mice. Together, the present results suggested that Libidibia ferrea induced antinociceptive activity is possibly related to its ability to inhibit opioid, cholinergic receptors, and cyclooxygenase-2 pathway, since its main component, linoleic acid, has been demonstrated to produce such effect in previous studies.

  15. Libidibia ferrea Mature Seeds Promote Antinociceptive Effect by Peripheral and Central Pathway: Possible Involvement of Opioid and Cholinergic Receptors

    Directory of Open Access Journals (Sweden)

    Luis Armando Sawada

    2014-01-01

    Full Text Available Libidibia ferrea (LF is a medicinal plant that holds many pharmacological properties. We evaluated the antinociceptive effect in the LF aqueous seed extract and Lipidic Portion of Libidibia ferrea (LPLF, partially elucidating their mechanisms. Histochemical tests and Gas chromatography of the LPLF were performed to characterize its fatty acids. Acetic acid-induced abdominal constriction, formalin-induced pain, and hot-plate test in mice were employed in the study. In all experiments, aqueous extract or LPLF was administered systemically at the doses of 1, 5, and 10 mg/kg. LF aqueous seed extract and LPLF demonstrated a dose-dependent antinociceptive effect in all tests indicating both peripheral anti-inflammatory and central analgesia properties. Also, the use of atropine (5 mg/kg, naloxone (5 mg/kg in the abdominal writhing test was able to reverse the antinociceptive effect of the LPLF, indicating that at least one of LF lipids components is responsible for the dose related antinociceptive action in chemical and thermal models of nociception in mice. Together, the present results suggested that Libidibia ferrea induced antinociceptive activity is possibly related to its ability to inhibit opioid, cholinergic receptors, and cyclooxygenase-2 pathway, since its main component, linoleic acid, has been demonstrated to produce such effect in previous studies.

  16. Effect of N-Methyl-D-Aspartate Receptor Antagonist Dextromethorphan on Opioid Analgesia in Pediatric Intensive Care Unit.

    Science.gov (United States)

    Naeem, Mohammed; Al Alem, Hala; Al Shehri, Ali; Al-Jeraisy, Majed

    2016-01-01

    Objective . Pain control is an essential goal in the management of critical children. Narcotics are the mainstay for pain control. Patients frequently need escalating doses of narcotics. In such cases an adjunctive therapy may be beneficial. Dextromethorphan (DM) is NMDA receptor antagonist and may prevent tolerance to narcotics; however, its definitive role is still unclear. We sought whether dextromethorphan addition could decrease the requirements of fentanyl to control pain in critical children. Design . Double-blind, randomized control trial (RCT). Setting . Pediatric multidisciplinary ICU in tertiary care center. Patients . Thirty-six pediatric patients 2-14 years of age in a multidisciplinary PICU requiring analgesia were randomized into dextromethorphan and placebo. The subjects in both groups showed similarity in most of the characteristics. Interventions . Subjects while receiving fentanyl for pain control received dextromethorphan or placebo through nasogastric/orogastric tubes for 96 hours. Pain was assessed using FLACC and faces scales. Measurements and Main Results . This study found no statistical significant difference in fentanyl requirements between subjects receiving dextromethorphan and those receiving placebo ( p = 0.127). Conclusions . Dextromethorphan has no effect on opioid requirement for control of acute pain in children admitted with acute critical care illness in PICU. The registration number for this trial is NCT01553435.

  17. Effect of N-Methyl-D-Aspartate Receptor Antagonist Dextromethorphan on Opioid Analgesia in Pediatric Intensive Care Unit

    Directory of Open Access Journals (Sweden)

    Mohammed Naeem

    2016-01-01

    Full Text Available Objective. Pain control is an essential goal in the management of critical children. Narcotics are the mainstay for pain control. Patients frequently need escalating doses of narcotics. In such cases an adjunctive therapy may be beneficial. Dextromethorphan (DM is NMDA receptor antagonist and may prevent tolerance to narcotics; however, its definitive role is still unclear. We sought whether dextromethorphan addition could decrease the requirements of fentanyl to control pain in critical children. Design. Double-blind, randomized control trial (RCT. Setting. Pediatric multidisciplinary ICU in tertiary care center. Patients. Thirty-six pediatric patients 2–14 years of age in a multidisciplinary PICU requiring analgesia were randomized into dextromethorphan and placebo. The subjects in both groups showed similarity in most of the characteristics. Interventions. Subjects while receiving fentanyl for pain control received dextromethorphan or placebo through nasogastric/orogastric tubes for 96 hours. Pain was assessed using FLACC and faces scales. Measurements and Main Results. This study found no statistical significant difference in fentanyl requirements between subjects receiving dextromethorphan and those receiving placebo (p=0.127. Conclusions. Dextromethorphan has no effect on opioid requirement for control of acute pain in children admitted with acute critical care illness in PICU. The registration number for this trial is NCT01553435.

  18. Partial separation of platelet and placental adenosine receptors from adenosine A2-like binding protein

    International Nuclear Information System (INIS)

    Zolnierowicz, S.; Work, C.; Hutchison, K.; Fox, I.H.

    1990-01-01

    The ubiquitous adenosine A2-like binding protein obscures the binding properties of adenosine receptors assayed with 5'-N-[ 3 H]ethylcarboxamidoadenosine [( 3 H]NECA). To solve this problem, we developed a rapid and simple method to separate adenosine receptors from the adenosine A2-like binding protein. Human platelet and placental membranes were solubilized with 1% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. The soluble platelet extract was precipitated with polyethylene glycol and the fraction enriched in adenosine receptors was isolated from the precipitate by differential centrifugation. The adenosine A2-like binding protein was removed from the soluble placental extract with hydroxylapatite and adenosine receptors were precipitated with polyethylene glycol. The specificity of the [ 3 H]NECA binding is typical of an adenosine A2 receptor for platelets and an adenosine A1 receptor for placenta. This method leads to enrichment of adenosine A2 receptors for platelets and adenosine A1 receptors for placenta. This provides a useful preparation technique for pharmacologic studies of adenosine receptors

  19. Free energy calculations offer insights into the influence of receptor flexibility on ligand-receptor binding affinities.

    Science.gov (United States)

    Dolenc, Jožica; Riniker, Sereina; Gaspari, Roberto; Daura, Xavier; van Gunsteren, Wilfred F

    2011-08-01

    Docking algorithms for computer-aided drug discovery and design often ignore or restrain the flexibility of the receptor, which may lead to a loss of accuracy of the relative free enthalpies of binding. In order to evaluate the contribution of receptor flexibility to relative binding free enthalpies, two host-guest systems have been examined: inclusion complexes of α-cyclodextrin (αCD) with 1-chlorobenzene (ClBn), 1-bromobenzene (BrBn) and toluene (MeBn), and complexes of DNA with the minor-groove binding ligands netropsin (Net) and distamycin (Dist). Molecular dynamics simulations and free energy calculations reveal that restraining of the flexibility of the receptor can have a significant influence on the estimated relative ligand-receptor binding affinities as well as on the predicted structures of the biomolecular complexes. The influence is particularly pronounced in the case of flexible receptors such as DNA, where a 50% contribution of DNA flexibility towards the relative ligand-DNA binding affinities is observed. The differences in the free enthalpy of binding do not arise only from the changes in ligand-DNA interactions but also from changes in ligand-solvent interactions as well as from the loss of DNA configurational entropy upon restraining.

  20. PI3K class II α regulates δ-opioid receptor export from the trans-Golgi network.

    Science.gov (United States)

    Shiwarski, Daniel J; Darr, Marlena; Telmer, Cheryl A; Bruchez, Marcel P; Puthenveedu, Manojkumar A

    2017-08-01

    The interplay between signaling and trafficking by G protein-coupled receptors (GPCRs) has focused mainly on endocytic trafficking. Whether and how surface delivery of newly synthesized GPCRs is regulated by extracellular signals is less understood. Here we define a signaling-regulated checkpoint at the trans -Golgi network (TGN) that controls the surface delivery of the delta opioid receptor (δR). In PC12 cells, inhibition of phosphoinositide-3 kinase (PI3K) activity blocked export of newly synthesized δR from the Golgi and delivery to the cell surface, similar to treatment with nerve growth factor (NGF). Depletion of class II phosphoinositide-3 kinase α (PI3K C2A), but not inhibition of class I PI3K, blocked δR export to comparable levels and attenuated δR-mediated cAMP inhibition. NGF treatment displaced PI3K C2A from the Golgi and optogenetic recruitment of the PI3K C2A kinase domain to the TGN-induced δR export downstream of NGF. Of importance, PI3K C2A expression promotes export of endogenous δR in primary trigeminal ganglion neurons. Taken together, our results identify PI3K C2A as being required and sufficient for δR export and surface delivery in neuronal cells and suggest that it could be a key modulator of a novel Golgi export checkpoint that coordinates GPCR delivery to the surface. © 2017 Shiwarski et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  1. Human myometrial adrenergic receptors: identification of the beta-adrenergic receptor by [3H]dihydroalprenolol binding

    International Nuclear Information System (INIS)

    Hayashida, D.N.; Leung, R.; Goldfien, A.; Roberts, J.M.

    1982-01-01

    The radioactive beta-adrenergic antagonist [ 3 H] dihydroalprenolol (DHA) binds to particulate preparations of human myometrium in a manner compatible with binding to the beta-adrenergic receptor. The binding of DHA is rapid (attaining equilibrium in 12 minutes), readily reversible (half time = 16 minutes), high affinity (K/sub D/ = 0.50 nM), low capacity (Bmax = 70 fmoles/mg of protein), and stereoselective ([-]-propranolol is 100 times as potent as [+] -propranolol in inhibiting DHA binding). Adrenergic agonists competed for DHA binding sites in a manner compatible with beta-adrenergic interactions and mirrored β 2 pharmacologic potencies: isoproterenol > epinephrine >> norepinephrine. Studies in which zinterol, a β 2 -adrenergic agonist, competed for DHA binding sites in human myometrial particulate indicated that at least 87% of the beta-adrenergic receptors present are β 2 -adrenergic receptors. Binding of DHA to human myometrial beta-adrenergic receptors provides a tool which may be used in the examination of gonadal hormonal modification of adrenergic response in human uterus as well as in the analysis of beta-adrenergic agents as potentially useful tocolytic agents

  2. DNA binding properties of dioxin receptors in wild-type and mutant mouse hepatoma cells

    International Nuclear Information System (INIS)

    Cuthill, S.; Poellinger, L.

    1988-01-01

    The current model of action of 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin) entails stimulation of target gene transcription via the formation of dioxin-receptor complexes and subsequent accumulation of the complexes within the cell nucleus. Here, the authors have analyzed the DNA binding properties of the dioxin receptor in wild-type mouse hepatoma (Hepa 1c1c7) cells and a class of nonresponsive mutant cells which fail to accumulate dioxin-receptor complexes within the nucleus in vivo. In vitro, both the wild-type and mutant [ 3 H]dioxin-receptor complexes exhibited low affinity for DNA-cellulose (5-8% and around 4% retention, respectively) in the absence of prior biochemical manipulations. However, following chromatography on heparin-Sepharose, the wild-type but not the mutant dioxin receptor was transformed to a species with an increased affinity for DNA (40-50% retention on DNA-cellulose). The gross molecular structure of the mutant, non DNA binding dioxin receptor did not appear to be altered as compared to that of the wild-type receptor. These results imply that the primary deficiency in the mutant dioxin receptor form may reside at the DNA binding level and that, in analogy to steroid hormone receptors, DNA binding of the receptor may be an essential step in the regulation of target gene transcription by dioxin

  3. Molecular mechanism of action of opioids in human neuroblastoma cells

    International Nuclear Information System (INIS)

    Yu, V.C.K.

    1987-01-01

    A series of human neuroblastoma cell lines was screened for the presence of opioid receptor sites. Of these cell lines, SK-N-SH was found to express approximately 50,000 μ and 10,000 δ opioid receptor sites/cell. In vitro characterization revealed that the binding properties of these receptor sites closely resembled those of human and rodent brain. Phosphatidylinositol turnover as a potential second messenger system for the μ receptor was examined in SK-N-SH cells. Neurotransmitter receptor systems were determined in the three sub-clones of SK-N-SH cells. Cells of the SH-SY5Y line, a phenotypically stable subclone of SK-N-SH cells, were induced to differentiate by treatment with various inducing agents, and changes of several neurotransmitter receptor systems were determined. Nerve growth factor (NGF) and retinoic acid (RA) up-regulated, while dBcAMP down-regulated opioid receptor sites. [ 3 H]Dopamine uptake was slightly enhanced only in RA-treated cells. Strikingly, the efficacy of PGE 1 -stimulated accumulation of cAMP was enhanced by 15- to 30-fold upon RA treatment

  4. Reduced post-synaptic serotonin type 1A receptor binding in bipolar depression

    Science.gov (United States)

    Nugent, Allison C.; Bain, Earle E.; Carlson, Paul J.; Neumeister, Alexander; Bonne, Omer; Carson, Richard E.; Eckelman, William; Herscovitch, Peter; Zarate, Carlos A.; Charney, Dennis S.; Drevets, Wayne C.

    2013-01-01

    Multiple lines of evidence suggest that serotonin type 1A (5-HT1A) receptor dysfunction is involved in the pathophysiology of mood disorders, and that alterations in 5-HT1A receptor function play a role in the mechanisms of antidepressant and mood stabilizer treatment. The literature is in disagreement, however, as to whether 5-HT1A receptor binding abnormalities exist in bipolar disorder (BD). We acquired PET images of 5-HT1A receptor binding in 26 unmedicated BD subjects and 37 healthy controls using [18F]FCWAY, a highly selective 5-HT1A receptor radio-ligand. The mean 5-HT1A receptor binding potential (BPP) was significantly lower in BD subjects compared to controls in cortical regions where 5-HT1A receptors are expressed post-synaptically, most prominently in the mesiotemporal cortex. Post-hoc assessments involving other receptor specific binding parameters suggested that this difference particularly affected the females with BD. The mean BPP did not differ between groups in the raphe nucleus, however, where 5-HT1A receptors are predominantly expressed pre-synaptically. Across subjects the BPP in the mesiotemporal cortex was inversely correlated with trough plasma cortisol levels, consistent with preclinical literature indicating that hippocampal 5-HT1A receptor expression is inhibited by glucocorticoid receptor stimulation. These findings suggest that 5-HT1A receptor binding is abnormally reduced in BD, and this abnormality may particularly involve the postsynaptic 5-HT1A receptor system of individuals with a tendency toward cortisol hypersecretion. PMID:23434290

  5. Does protein binding modulate the effect of angiotensin II receptor antagonists?

    Directory of Open Access Journals (Sweden)

    Marc P Maillard

    2001-03-01

    Full Text Available IntroductionAngiotensin II AT 1-receptor antagonists are highly bound to plasma proteins (≥ 99%. With some antagonists, such as DuP-532, the protein binding was such that no efficacy of the drug could be demonstrated clinically. Whether protein binding interferes with the efficacy of other antagonists is not known. We have therefore investigated in vitro how plasma proteins may affect the antagonistic effect of different AT1-receptor antagonists.MethodsA radio-receptor binding assay was used to analyse the interaction between proteins and the ability of various angiotensin II (Ang II antagonists to block AT1-receptors. In addition, the Biacore technology, a new technique which enables the real-time monitoring of binding events between two molecules, was used to evaluate the dissociation rate constants of five AT1-receptor antagonists from human serum albumin.ResultsThe in vitro AT 1-antagonistic effects of different Ang II receptor antagonists were differentially affected by the presence of human plasma, with rightward shifts of the IC50 ranging from one to several orders of magnitude. The importance of the shift correlates with the dissociation rate constants of these drugs from albumin. Our experiments also show that the way that AT1-receptor antagonists bind to proteins differs from one compound to another. These results suggest that the interaction with plasma proteins appears to modulate the efficacy of some Ang II antagonists.ConclusionAlthough the high binding level of Ang II receptor antagonist to plasma proteins appears to be a feature common to this class of compounds, the kinetics and characteristics of this binding is of great importance. With some antagonists, protein binding interferes markedly with their efficacy to block AT1-receptors.

  6. Allosteric regulation by oleamide of the binding properties of 5-hydroxytryptamine7 receptors.

    Science.gov (United States)

    Hedlund, P B; Carson, M J; Sutcliffe, J G; Thomas, E A

    1999-12-01

    Oleamide belongs to a family of amidated lipids with diverse biological activities, including sleep induction and signaling modulation of several 5-hydroxytryptamine (5-HT) receptor subtypes, including 5-HT1A, 5-HT2A/2C, and 5-HT7. The 5-HT7 receptor, predominantly localized in the hypothalamus, hippocampus, and frontal cortex, stimulates cyclic AMP formation and is thought to be involved in the regulation of sleep-wake cycles. Recently, it was proposed that oleamide acts at an allosteric site on the 5-HT7 receptor to regulate cyclic AMP formation. We have further investigated the interaction between oleamide and 5-HT7 receptors by performing radioligand binding assays with HeLa cells transfected with the 5-HT7 receptor. Methiothepin, clozapine, and 5-HT all displaced specific [3H]5-HT (100 nM) binding, with pK(D) values of 7.55, 7.85, and 8.39, respectively. Oleamide also displaced [3H]5-HT binding, but the maximum inhibition was only 40% of the binding. Taking allosteric (see below) cooperativity into account, a K(D) of 2.69 nM was calculated for oleamide. In saturation binding experiments, oleamide caused a 3-fold decrease in the affinity of [3H]5-HT for the 5-HT7 receptor, without affecting the number of binding sites. A Schild analysis showed that the induced shift in affinity of [3H]5-HT reached a plateau, unlike that of a competitive inhibitor, illustrating the allosteric nature of the interaction between oleamide and the 5-HT7 receptor. Oleic acid, the product of oleamide hydrolysis, had a similar effect on [3H]5-HT binding, whereas structural analogs of oleamide, trans-9,10-octadecenamide, cis-8,9-octadecenamide, and erucamide, did not alter [3H]5-HT binding significantly. The findings support the hypothesis that oleamide acts via an allosteric site on the 5-HT7 receptor regulating receptor affinity.

  7. The Roles of Hemagglutinin Phe-95 in Receptor Binding and Pathogenicity of Influenza B Virus

    Science.gov (United States)

    Ni, Fengyun; Mbawuike, Innocent Nnadi; Kondrashkina, Elena; Wang, Qinghua

    2014-01-01

    Diverged ~4,000 years ago, influenza B virus has several important differences from influenza A virus, including lower receptor-binding affinity and highly restricted host range. Based on our prior structural studies, we hypothesized that a single-residue difference in the receptor-binding site of hemagglutinin (HA), Phe-95 in influenza B virus versus Tyr-98 in influenza A/H1~H15, is possibly a key determinant for the low receptor-binding affinity. Here we demonstrate that the mutation Phe95→Tyr in influenza B virus HA restores all three hydrogen bonds made by Tyr-98 in influenza A/H3 HA and has the potential to enhance receptor binding. However, the full realization of this potential is influenced by the local environment into which the mutation is introduced. The binding and replication of the recombinant viruses correlate well with the receptor-binding capabilities of HA. These results are discussed in relation to the roles of Phe-95 in receptor binding and pathogenicity of influenza B virus. PMID:24503069

  8. Analyzing machupo virus-receptor binding by molecular dynamics simulations

    Directory of Open Access Journals (Sweden)

    Austin G. Meyer

    2014-02-01

    Full Text Available In many biological applications, we would like to be able to computationally predict mutational effects on affinity in protein–protein interactions. However, many commonly used methods to predict these effects perform poorly in important test cases. In particular, the effects of multiple mutations, non alanine substitutions, and flexible loops are difficult to predict with available tools and protocols. We present here an existing method applied in a novel way to a new test case; we interrogate affinity differences resulting from mutations in a host–virus protein–protein interface. We use steered molecular dynamics (SMD to computationally pull the machupo virus (MACV spike glycoprotein (GP1 away from the human transferrin receptor (hTfR1. We then approximate affinity using the maximum applied force of separation and the area under the force-versus-distance curve. We find, even without the rigor and planning required for free energy calculations, that these quantities can provide novel biophysical insight into the GP1/hTfR1 interaction. First, with no prior knowledge of the system we can differentiate among wild type and mutant complexes. Moreover, we show that this simple SMD scheme correlates well with relative free energy differences computed via free energy perturbation. Second, although the static co-crystal structure shows two large hydrogen-bonding networks in the GP1/hTfR1 interface, our simulations indicate that one of them may not be important for tight binding. Third, one viral site known to be critical for infection may mark an important evolutionary suppressor site for infection-resistant hTfR1 mutants. Finally, our approach provides a framework to compare the effects of multiple mutations, individually and jointly, on protein–protein interactions.

  9. Analyzing machupo virus-receptor binding by molecular dynamics simulations

    Science.gov (United States)

    Sawyer, Sara L.; Ellington, Andrew D.; Wilke, Claus O.

    2014-01-01

    In many biological applications, we would like to be able to computationally predict mutational effects on affinity in protein–protein interactions. However, many commonly used methods to predict these effects perform poorly in important test cases. In particular, the effects of multiple mutations, non alanine substitutions, and flexible loops are difficult to predict with available tools and protocols. We present here an existing method applied in a novel way to a new test case; we interrogate affinity differences resulting from mutations in a host–virus protein–protein interface. We use steered molecular dynamics (SMD) to computationally pull the machupo virus (MACV) spike glycoprotein (GP1) away from the human transferrin receptor (hTfR1). We then approximate affinity using the maximum applied force of separation and the area under the force-versus-distance curve. We find, even without the rigor and planning required for free energy calculations, that these quantities can provide novel biophysical insight into the GP1/hTfR1 interaction. First, with no prior knowledge of the system we can differentiate among wild type and mutant complexes. Moreover, we show that this simple SMD scheme correlates well with relative free energy differences computed via free energy perturbation. Second, although the static co-crystal structure shows two large hydrogen-bonding networks in the GP1/hTfR1 interface, our simulations indicate that one of them may not be important for tight binding. Third, one viral site known to be critical for infection may mark an important evolutionary suppressor site for infection-resistant hTfR1 mutants. Finally, our approach provides a framework to compare the effects of multiple mutations, individually and jointly, on protein–protein interactions. PMID:24624315

  10. Modelling the interdependence between the stoichiometry of receptor oligomerization and ligand binding for a coexisting dimer/tetramer receptor system.

    Science.gov (United States)

    Rovira, X; Vivó, M; Serra, J; Roche, D; Strange, P G; Giraldo, J

    2009-01-01

    Many G protein-coupled receptors have been shown to exist as oligomers, but the oligomerization state and the effects of this on receptor function are unclear. For some G protein-coupled receptors, in ligand binding assays, different radioligands provide different maximal binding capacities. Here we have developed mathematical models for co-expressed dimeric and tetrameric species of receptors. We have considered models where the dimers and tetramers are in equilibrium and where they do not interconvert and we have also considered the potential influence of the ligands on the degree of oligomerization. By analogy with agonist efficacy, we have considered ligands that promote, inhibit or have no effect on oligomerization. Cell surface receptor expression and the intrinsic capacity of receptors to oligomerize are quantitative parameters of the equations. The models can account for differences in the maximal binding capacities of radioligands in different preparations of receptors and provide a conceptual framework for simulation and data fitting in complex oligomeric receptor situations.

  11. Molecular characterization of a novel human hybrid-type receptor that binds the alpha2-macroglobulin receptor-associated protein

    DEFF Research Database (Denmark)

    Jacobsen, Linda; Madsen, P; Moestrup, S K

    1996-01-01

    the corresponding cDNA. The gene, designated SORL1, maps to chromosome 11q 23/24 and encodes a 2214-residue type 1 receptor containing a furin cleavage site immediately preceding the N terminus determined in the purified protein. The receptor, designated sorLA-1, has a short cytoplasmic tail containing a tyrosine...... density lipoprotein receptor gene family receptors, and 3) six tandemly arranged fibronectin type III repeats also found in certain neural adhesion proteins. sorLA-1 may therefore be classified as a hybrid receptor. Northern blotting revealed specific mRNA transcripts in brain, spinal cord, and testis......The 39-40-kDa receptor-associated protein (RAP) binds to the members of the low density lipoprotein receptor gene family and functions as a specialized endoplasmic reticulum/Golgi chaperone. Using RAP affinity chromatography, we have purified a novel approximately 250-kDa brain protein and isolated...

  12. Monoclonal antibody to the rat glucocorticoid receptor. Relationship between the immunoreactive and DNA-binding domain

    International Nuclear Information System (INIS)

    Eisen, L.P.; Reichman, M.E.; Thompson, E.B.; Gametchu, B.; Harrison, R.W.; Eisen, H.J.

    1985-01-01

    The region of the glucocorticoid receptor that reacted with a monoclonal antibody (BUGR-1) was identified. In order to identify the immunoreactive region, the rat liver glucocorticoid receptor was subjected to limited proteolysis; immunoreactive fragments were identified by Western blotting. The monoclonal antibody reacted with both the undigested Mr approximately 97,000 receptor subunit and a Mr approximately 45,000 fragment containing the steroid-binding and DNA-binding domains. Digestion by trypsin also produced two steroid-binding fragments of Mr approximately 27,000 and 31,000 which did not react with the antibody and an immunoreactive Mr approximately 16,000 fragment. This Mr approximately 16,000 fragment was shown to bind to DNA-cellulose, indicating that it contained a DNA-binding domain of the receptor. The undigested receptor must have steroid associated with it to undergo activation to a DNA-binding form. However, the Mr approximately 16,000 immunoreactive fragment binds to DNA-cellulose even if it is obtained by digestion of the steroid-free holoreceptor which does not itself bind to DNA

  13. Diminished hepatic growth hormone receptor binding in sex-linked dwarf broiler and leghorn chickens.

    Science.gov (United States)

    Leung, F C; Styles, W J; Rosenblum, C I; Lilburn, M S; Marsh, J A

    1987-02-01

    Hepatic growth hormone (GH) receptor binding was compared in normal and sex-linked dwarfs (SLD) from both Hubbard and Cornell strain chickens. At 6, 8, and 20 weeks of age, hepatic GH receptor binding in the Hubbard SLD chickens was significantly lower than that of normal fast-growing birds. At 20 weeks of age, only 2 of 22 SLD chickens in the Hubbard broiler strain showed positive binding at a high enough level to allow for Scatchard analysis. The affinity constants and binding capacities of these two SLD chickens were numerically (but not significantly) lower than those of the normal fast-growing birds. We further examined hepatic GH receptor binding in two closely related White Leghorn strains of chickens that have been maintained as closed breeding populations for many years. We observed no detectable hepatic GH binding in the Cornell SLD chickens (N = 20), as compared to the normal-growing control strain (K strain). In both SLD strains, pretreatment with 4 M MgCl2 did not enhance GH binding, suggesting that there was no endogenous GH binding to the receptor. Based on these data, we suggest that the lack, or greatly reduced number, of GH receptors may be a major contributing factor to the dwarfism observed in these strains.

  14. Substance P and substance K receptor binding sites in the human gastrointestinal tract: localization by autoradiography

    International Nuclear Information System (INIS)

    Gates, T.S.; Zimmerman, R.P.; Mantyh, C.R.; Vigna, S.R.; Maggio, J.E.; Welton, M.L.; Passaro, E.P. Jr.; Mantyh, P.W.

    1988-01-01

    Quantitative receptor autoradiography was used to localize and quantify the distribution of binding sites for 125 I-radiolabeled substance P (SP), substance K (SK) and neuromedin K (NK) in the human GI tract using histologically normal tissue obtained from uninvolved margins of resections for carcinoma. The distribution of SP and SK binding sites is different for each gastrointestinal (GI) segment examined. Specific SP binding sites are expressed by arterioles and venules, myenteric plexus, external circular muscle, external longitudinal muscle, muscularis mucosa, epithelial cells of the mucosa, and the germinal centers of lymph nodules. SK binding sites are distributed in a pattern distinct from SP binding sites and are localized to the external circular muscle, external longitudinal muscle, and the muscularis mucosa. Binding sites for NK were not detected in any part of the human GI tract. These results demonstrate that: (1) surgical specimens from the human GI tract can be effectively processed for quantitative receptor autoradiography; (2) of the three mammalian tachykinins tested, SP and SK, but not NK binding sites are expressed in detectable levels in the human GI tract; (3) whereas SK receptor binding sites are expressed almost exclusively by smooth muscle, SP binding sites are expressed by smooth muscle cells, arterioles, venules, epithelial cells of the mucosa and cells associated with lymph nodules; and (4) both SP and SK binding sites expressed by smooth muscle are more stable than SP binding sites expressed by blood vessels, lymph nodules, and mucosal cells

  15. Ascorbic acid enables reversible dopamine receptor 3H-agonist binding

    International Nuclear Information System (INIS)

    Leff, S.; Sibley, D.R.; Hamblin, M.; Creese, I.

    1981-01-01

    The effects of ascorbic acid on dopaminergic 3 H-agonist receptor binding were studied in membrane homogenates of bovine anterior pituitary and caudate, and rat striatum. In all tissues virtually no stereospecific binding (defined using 1uM (+)butaclamol) of the 3 H-agonists N-propylnorapomorphine (NPA), apomorphine, or dopamine could be demonstrated in the absence of ascorbic acid. Although levels of total 3 H-agonist binding were three to five times greater in the absence than in the presence of 0.1% ascorbic acid, the increased binding was entirely non-stereospecific. Greater amounts of dopamine-inhibitable 3 H-NPA binding could be demonstrated in the absence of 0.1% ascorbic acid, but this measure of ''specific binding'' was demonstrated not to represent dopamine receptor binding since several other catecholamines and catechol were equipotent with dopamine and more potent than the dopamine agonist (+/-)amino-6,7-dihydroxy-1,2,3,4-tetrahydronapthalene (ADTN) in inhibiting this binding. High levels of dopamine-displaceable 3 H-agonist binding were detected in fresh and boiled homogenates of cerebellum, an area of brain which receives no dopaminergic innervation, further demonstrating the non-specific nature of 3 H-agonist binding in the absence of ascorbic acid. These studies emphasize that under typical assay conditions ascorbic acid is required in order to demonstrate reversible and specific 3 H-agonist binding to dopamine receptors

  16. Targinact--opioid pain relief without constipation?

    Science.gov (United States)

    2010-12-01

    Targinact (Napp Pharmaceuticals Ltd) is a modified-release combination product containing the strong opioid oxycodone plus the opioid antagonist naloxone. It is licensed for "severe pain, which can be adequately managed only with opioid analgesics".1 The summary of product characteristics (SPC) states that "naloxone is added to counteract opioid-induced constipation by blocking the action of oxycodone at opioid receptors locally in the gut". Advertising for the product claims "better pain relief", "superior GI [gastrointestinal] tolerability" and "improved quality of life" "compared to previous treatment in a clinical practice study (n=7836)". Here we consider whether Targinact offers advantages over using strong opioids plus laxatives where required.

  17. [3H]cytisine binding to nicotinic cholinergic receptors in brain

    International Nuclear Information System (INIS)

    Pabreza, L.A.; Dhawan, S.; Kellar, K.J.

    1991-01-01

    Cytisine, a ganglionic agonist, competes with high affinity for brain nicotinic cholinergic receptors labeled by any of several nicotinic 3 H-agonist ligands. Here we have examined the binding of [ 3 H]cytisine in rat brain homogenates. [ 3 H]Cytisine binds with high affinity (Kd less than 1 nM), and specific binding represented 60-90% of total binding at all concentrations examined up to 15 nM. The nicotinic cholinergic agonists nicotine, acetylcholine, and carbachol compete with high affinity for [ 3 H]cytisine binding sites, whereas among nicotinic receptor antagonists only dihydro-beta-erythroidine competes with high affinity (in the nanomolar range). Comparison of binding in several brain regions showed that [ 3 H]cytisine binding is higher in the thalamus, striatum, and cortex than in the hippocampus, cerebellum, or hypothalamus. The pharmacology and brain regional distribution of [ 3 H]cytisine binding sites are those predicted for neuronal nicotinic receptor agonist recognition sites. The high affinity and low nonspecific binding of [ 3 H]cytisine should make it a very useful ligand for studying neuronal nicotinic receptors

  18. Is the isolated ligand binding domain a good model of the domain in the native receptor?

    Science.gov (United States)

    Deming, Dustin; Cheng, Qing; Jayaraman, Vasanthi

    2003-05-16

    Numerous studies have used the atomic level structure of the isolated ligand binding domain of the glutamate receptor to elucidate the agonist-induced activation and desensitization processes in this group of proteins. However, no study has demonstrated the structural equivalence of the isolated ligand binding fragments and the protein in the native receptor. In this report, using visible absorption spectroscopy we show that the electronic environment of the antagonist 6-cyano-7-nitro-2,3-dihydroxyquinoxaline is identical for the isolated protein and the native glutamate receptors expressed in cells. Our results hence establish that the local structure of the ligand binding site is the same in the two proteins and validate the detailed structure-function relationships that have been developed based on a comparison of the structure of the isolated ligand binding domain and electrophysiological consequences in the native receptor.

  19. Potential ligand-binding residues in rat olfactory receptors identified by correlated mutation analysis

    Science.gov (United States)

    Singer, M. S.; Oliveira, L.; Vriend, G.; Shepherd, G. M.

    1995-01-01

    A family of G-protein-coupled receptors is believed to mediate the recognition of odor molecules. In order to identify potential ligand-binding residues, we have applied correlated mutation analysis to receptor sequences from the rat. This method identifies pairs of sequence positions where residues remain conserved or mutate in tandem, thereby suggesting structural or functional importance. The analysis supported molecular modeling studies in suggesting several residues in positions that were consistent with ligand-binding function. Two of these positions, dominated by histidine residues, may play important roles in ligand binding and could confer broad specificity to mammalian odor receptors. The presence of positive (overdominant) selection at some of the identified positions provides additional evidence for roles in ligand binding. Higher-order groups of correlated residues were also observed. Each group may interact with an individual ligand determinant, and combinations of these groups may provide a multi-dimensional mechanism for receptor diversity.

  20. Absence of serum growth hormone binding protein in patients with growth hormone receptor deficiency (Laron dwarfism)

    Energy Technology Data Exchange (ETDEWEB)

    Daughaday, W.H.; Trivedi, B.

    1987-07-01

    It has recently been recognized that human serum contains a protein that specifically binds human growth hormone (hGH). This protein has the same restricted specificity for hGH as the membrane-bound GH receptor. To determine whether the GH-binding protein is a derivative of, or otherwise related to, the GH receptor, the authors have examined the serum of three patients with Laron-type dwarfism, a condition in which GH refractoriness has been attributed to a defect in the GH receptor. The binding of /sup 125/I-labeled hGH incubated with serum has been measured after gel filtration of the serum through an Ultrogel AcA 44 minicolumn. Results are expressed as percent of specifically bound /sup 125/I-hGH and as specific binding relative to that of a reference serum after correction is made for endogenous GH. The mean +/- SEM of specific binding of sera from eight normal adults (26-46 years of age) was 21.6 +/- 0.45%, and the relative specific binding was 101.1 +/- 8.6%. Sera from 11 normal children had lower specific binding of 12.5 +/- 1.95% and relative specific binding of 56.6 +/- 9.1%. Sera from three children with Laron-type dwarfism lacked any demonstrable GH binding, whereas sera from 10 other children with other types of nonpituitary short stature had normal relative specific binding. They suggest that the serum GH-binding protein is a soluble derivative of the GH receptor. Measurement of the serum GH-binding protein may permit recognition of other abnormalities of the GH receptor.

  1. Absence of serum growth hormone binding protein in patients with growth hormone receptor deficiency (Laron dwarfism)

    International Nuclear Information System (INIS)

    Daughaday, W.H.; Trivedi, B.

    1987-01-01

    It has recently been recognized that human serum contains a protein that specifically binds human growth hormone (hGH). This protein has the same restricted specificity for hGH as the membrane-bound GH receptor. To determine whether the GH-binding protein is a derivative of, or otherwise related to, the GH receptor, the authors have examined the serum of three patients with Laron-type dwarfism, a condition in which GH refractoriness has been attributed to a defect in the GH receptor. The binding of 125 I-labeled hGH incubated with serum has been measured after gel filtration of the serum through an Ultrogel AcA 44 minicolumn. Results are expressed as percent of specifically bound 125 I-hGH and as specific binding relative to that of a reference serum after correction is made for endogenous GH. The mean +/- SEM of specific binding of sera from eight normal adults (26-46 years of age) was 21.6 +/- 0.45%, and the relative specific binding was 101.1 +/- 8.6%. Sera from 11 normal children had lower specific binding of 12.5 +/- 1.95% and relative specific binding of 56.6 +/- 9.1%. Sera from three children with Laron-type dwarfism lacked any demonstrable GH binding, whereas sera from 10 other children with other types of nonpituitary short stature had normal relative specific binding. They suggest that the serum GH-binding protein is a soluble derivative of the GH receptor. Measurement of the serum GH-binding protein may permit recognition of other abnormalities of the GH receptor

  2. Low 5-HT1B receptor binding in the migraine brain

    DEFF Research Database (Denmark)

    Deen, Marie; Hansen, Hanne D; Hougaard, Anders

    2018-01-01

    Background The pathophysiology of migraine may involve dysfunction of serotonergic signaling. In particular, the 5-HT1B receptor is considered a key player due to the efficacy of 5-HT1B receptor agonists for treatment of migraine attacks. Aim To examine the cerebral 5-HT1B receptor binding....... Patients who reported migraine brain regions involved in pain modulation as regions of interest and applied a latent variable model (LVM) to assess the group effect on binding across these regions. Results Our data...... support a model wherein group status predicts the latent variable ( p = 0.038), with migraine patients having lower 5-HT1B receptor binding across regions compared to controls. Further, in a whole-brain voxel-based analysis, time since last migraine attack correlated positively with 5-HT1B receptor...

  3. Estrogen receptor-independent catechol estrogen binding activity: protein binding studies in wild-type, Estrogen receptor-alpha KO, and aromatase KO mice tissues.

    Science.gov (United States)

    Philips, Brian J; Ansell, Pete J; Newton, Leslie G; Harada, Nobuhiro; Honda, Shin-Ichiro; Ganjam, Venkataseshu K; Rottinghaus, George E; Welshons, Wade V; Lubahn, Dennis B

    2004-06-01

    Primary evidence for novel estrogen signaling pathways is based upon well-documented estrogenic responses not inhibited by estrogen receptor antagonists. In addition to 17beta-E2, the catechol estrogen 4-hydroxyestradiol (4OHE2) has been shown to elicit biological responses independent of classical estrogen receptors in estrogen receptor-alpha knockout (ERalphaKO) mice. Consequently, our research was designed to biochemically characterize the protein(s) that could be mediating the biological effects of catechol estrogens using enzymatically synthesized, radiolabeled 4-hydroxyestrone (4OHE1) and 4OHE2. Scatchard analyses identified a single class of high-affinity (K(d) approximately 1.6 nM), saturable cytosolic binding sites in several ERalphaKO estrogen-responsive tissues. Specific catechol estrogen binding was competitively inhibited by unlabeled catechol estrogens, but not by 17beta-E2 or the estrogen receptor antagonist ICI 182,780. Tissue distribution studies indicated significant binding differences both within and among various tissues in wild-type, ERalphaKO, and aromatase knockout female mice. Ligand metabolism experiments revealed extensive metabolism of labeled catechol estrogen, suggesting that catechol estrogen metabolites were responsible for the specific binding. Collectively, our data provide compelling evidence for the interaction of catechol estrogen metabolites with a novel binding protein that exhibits high affinity, specificity, and selective tissue distribution. The extensive biochemical characterization of this binding protein indicates that this protein may be a receptor, and thus may mediate ERalpha/beta-independent effects of catechol estrogens and their metabolites.

  4. Evaluation of the In Vivo and Ex Vivo Binding of Novel BC1 Cannabinoid Receptor Radiotracers

    Energy Technology Data Exchange (ETDEWEB)

    Miller, A.; Gatley, J.; Gifford, A.

    2002-01-01

    The primary active ingredient of marijuana, 9-tetrahydrocannabinol, exerts its psychoactive effects by binding to cannabinoid CB1 receptors. These receptors are found throughout the brain with high concentrations in the hippocampus and cerebellum. The current study was conducted to evaluate the binding of a newly developed putative cannabinoid antagonist, AM630, and a classical cannabinoid 8-tetrahydrocannabinol as potential PET and/or SPECT imaging agents for brain CB1 receptors. For both of these ligands in vivo and ex vivo studies in mice were conducted. AM630 showed good overall brain uptake (as measure by %IA/g) and a moderately rapid clearance from the brain with a half-clearance time of approximately 30 minutes. However, AM630 did not show selective binding to CB1 cannabinoid receptors. Ex vivo autoradiography supported the lack of selective binding seen in the in vivo study. Similar to AM630, 8-tetrahydrocanibol also failed to show selective binding to CB1 receptor rich brain areas. The 8-tetrahydrocanibol showed moderate overall brain uptake and relatively slow brain clearance as compared to AM630. Further studies were done with AM2233, a cannabinoid ligand with a similar structure as AM630. These studies were done to develop an ex vivo binding assay to quantify the displacement of [131I]AM2233 binding by other ligands in Swiss-Webster and CB1 receptor knockout mice. By developing this assay we hoped to determine the identity of an unknown binding site for AM2233 present in the hippocampus of CB1 knockout mice. Using an approach based on incubation of brain slices prepared from mice given intravenous [131I]AM2233 in either the presence or absence of AM2233 (unlabelled) it was possible to demonstrate a significant AM2233-displacable binding in the Swiss-Webster mice. Future studies will determine if this assay is appropriate for identifying the unknown binding site for AM2233 in the CB1 knockout mice.

  5. Association of dopamine D(3) receptors with actin-binding protein 280 (ABP-280).

    Science.gov (United States)

    Li, Ming; Li, Chuanyu; Weingarten, Paul; Bunzow, James R; Grandy, David K; Zhou, Qun Yong

    2002-03-01

    Proteins that bind to G protein-coupled receptors have been identified as regulators of receptor localization and signaling. In our previous studies, a cytoskeletal protein, actin-binding protein 280 (ABP-280), was found to associate with the third cytoplasmic loop of dopamine D(2) receptors. In this study, we demonstrate that ABP-280 also interacts with dopamine D(3) receptors, but not with D(4) receptors. Similar to the dopamine D(2) receptor, the D(3)/ABP-280 association is of signaling importance. In human melanoma M2 cells lacking ABP-280, D(3) receptors were unable to inhibit forskolin-stimulated cyclic AMP (cAMP) production significantly. D(4) receptors, however, exhibited a similar degree of inhibition of forskolin-stimulated cAMP production in ABP-280-deficient M2 cells and ABP-280-replent M2 subclones (A7 cells). Further experiments revealed that the D(3)/ABP-280 interaction was critically dependent upon a 36 amino acid carboxyl domain of the D(3) receptor third loop, which is conserved in the D(2) receptor but not in the D(4) receptor. Our results demonstrate a subtype-specific regulation of dopamine D(2)-family receptor signaling by the cytoskeletal protein ABP-280.

  6. Genome-wide identification of estrogen receptor alpha-binding sites in mouse liver

    DEFF Research Database (Denmark)

    Gao, Hui; Fält, Susann; Sandelin, Albin

    2007-01-01

    We report the genome-wide identification of estrogen receptor alpha (ERalpha)-binding regions in mouse liver using a combination of chromatin immunoprecipitation and tiled microarrays that cover all nonrepetitive sequences in the mouse genome. This analysis identified 5568 ERalpha-binding regions...... genes. The majority of ERalpha-binding regions lie in regions that are evolutionarily conserved between human and mouse. Motif-finding algorithms identified the estrogen response element, and variants thereof, together with binding sites for activator protein 1, basic-helix-loop-helix proteins, ETS...... signaling in mouse liver, by characterizing the first step in this signaling cascade, the binding of ERalpha to DNA in intact chromatin....

  7. Changes in 5-HT4 receptor and 5-HT transporter binding in olfactory bulbectomized and glucocorticoid receptor heterozygous mice

    DEFF Research Database (Denmark)

    Licht, Cecilie L; Kirkegaard, Lisbeth; Zueger, Maha

    2010-01-01

    . The olfactory bulbectomized mice displayed increased activity in the open field test, a characteristic depression-like feature of this model. After bulbectomy, 5-HT(4) receptor binding was increased in the ventral hippocampus (12%) but unchanged in the dorsal hippocampus, frontal and caudal caudate putamen......]citalopram in two murine models of depression-related states, olfactory bulbectomy and glucocorticoid receptor heterozygous (GR(+/-)) mice. The olfactory bulbectomy model is characterized by 5-HT system changes, while the GR(+/-) mice have a deficit in hypothalamic-pituitary-adrenal (HPA) system control....... Among post hoc analyzed regions, there was a 14% decrease in 5-HT(4) receptor binding in the olfactory tubercles. The 5-HTT binding was unchanged in the hippocampus and caudate putamen of bulbectomized mice but post hoc analysis showed small decreases in lateral septum and lateral globus pallidus...

  8. In Silico Investigation of the Neurotensin Receptor 1 Binding Site

    DEFF Research Database (Denmark)

    Lückmann, Michael; Holst, Birgitte; Schwartz, Thue W.

    2016-01-01

    structure of NTSR1 in complex with NTS8-13 has been detd., providing novel insights into peptide ligand recognition by 7TM receptors. SR48692, a potent and selective small mol. antagonist has previously been used extensively as a tool compd. to study NTSR1 receptor signaling properties. To investigate......The neurotensin receptor 1 (NTSR1) belongs to the family of 7TM, G protein-coupled receptors, and is activated by the 13-amino-acid peptide neurotensin (NTS) that has been shown to play important roles in neurol. disorders and the promotion of cancer cells. Recently, a high-resoln. x-ray crystal...

  9. The novel δ opioid receptor agonist KNT-127 produces antidepressant-like and antinociceptive effects in mice without producing convulsions.

    Science.gov (United States)

    Saitoh, Akiyoshi; Sugiyama, Azusa; Nemoto, Toru; Fujii, Hideaki; Wada, Keiji; Oka, Jun-Ichiro; Nagase, Hiroshi; Yamada, Mitsuhiko

    2011-10-01

    We previously reported that the δ opioid receptor (DOP) agonists SNC80 and TAN-67 produce potent antidepressant-like and antinociceptive effects in rodents. However, SNC80 produced convulsive effects. Recently, we succeeded in synthesizing a novel DOP agonist called KNT-127. The present study examined the convulsive, antidepressant-like, and antinociceptive effects of KNT-127 in mice. In contrast to SNC80, KNT-127 produced no convulsions at doses of up to 100mg/kg. In mice subjected to the forced swim test, a screening model for antidepressants, KNT-127 (1mg/kg, s.c.) significantly decreased the duration of immobility and increased the duration of swimming without influencing spontaneous locomotor activity. These behavioral changes were similar to that observed for the tricyclic antidepressant imipramine (6mg/kg). The antidepressant-like effect of KNT-127 in mice was antagonized by pretreatment with naltrindole (NTI), a selective DOP antagonist, or naltriben, a putative DOP(2) subtype antagonist. In addition, KNT-127 (3mg/kg, s.c.) significantly reduced the number of acetic acid-induced abdominal constrictions and the duration of licking time, respectively, in mice subjected to a writhing test and a formalin test. These antinociceptive effects were antagonized by pretreatment with either NTI or 7-benzylidenenaltrexone, a putative DOP(1) subtype antagonist. We propose that KNT-127 should be considered as a candidate compound for the development of DOP-based antidepressants and/or analgesics that lack convulsive effects. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

  10. Dynamic Regulation of Delta-Opioid Receptor in Rat Trigeminal Ganglion Neurons by Lipopolysaccharide-induced Acute Pulpitis.

    Science.gov (United States)

    Huang, Jin; Lv, Yiheng; Fu, Yunjie; Ren, Lili; Wang, Pan; Liu, Baozhu; Huang, Keqiang; Bi, Jing

    2015-12-01

    Delta-opioid receptor (DOR) and its endogenous ligands distribute in trigeminal system and play a very important role in modulating peripheral inflammatory pain. DOR activation can trigger p44/42 mitogen-activated protein kinase (ERK1/2) and Akt signaling pathways, which participate in anti-inflammatory and neuroprotective effects. In this study, our purpose was to determine the dynamic changes of DOR in trigeminal ganglion (TG) neurons during the process of acute dental pulp inflammation and elucidate its possible mechanism. Forty rats were used to generate lipopolysaccharide-induced acute pulpitis animal models at 6, 12, and 24 hours and sham-operated groups. Acute pulpitis was confirmed by hematoxylin-eosin staining, and TG neuron activation was determined by anti-c-Fos immunohistochemistry. DOR protein and gene expression in TG was investigated by immunohistochemistry, Western blotting, and real-time polymerase chain reaction, and DOR expression in trigeminal nerves and dental pulp was also determined by immunohistochemistry. To further investigate the mechanism of DOR modulating acute inflammation, the change of pErk1/2 and pAkt in TG was examined by immunohistochemistry. Lipopolysaccharide could successfully induce acute pulpitis and activated TG neurons. Acute pulpitis could dynamically increase DOR protein and gene expression at 6, 12, and 24 hours in TG, and DOR dimerization was significantly increased at 12 and 24 hours. Acute pulpitis also induced the dynamic change of DOR protein in trigeminal nerve and dental pulp. Furthermore, ERK1/2 and Akt signaling pathways were inhibited in TG after acute pulpitis. Increased DOR expression and dimerization may play important roles in peripheral acute inflammatory pain. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  11. Involvement of delta opioid receptors in alcohol withdrawal-induced mechanical allodynia in male C57BL/6 mice.

    Science.gov (United States)

    Alongkronrusmee, Doungkamol; Chiang, Terrance; van Rijn, Richard M

    2016-10-01

    As a legal drug, alcohol is commonly abused and it is estimated that 17 million adults in the United States suffer from alcohol use disorder. Heavy alcoholics can experience withdrawal symptoms including anxiety and mechanical allodynia that can facilitate relapse. The molecular mechanisms underlying this phenomenon are not well understood, which stifles development of new therapeutics. Here we investigate whether delta opioid receptors (DORs) play an active role in alcohol withdrawal-induced mechanical allodynia (AWiMA) and if DOR agonists may provide analgesic relief from AWiMA. To study AWiMA, adult male wild-type and DOR knockout C57BL/6 mice were exposed to alcohol by a voluntary drinking model or oral gavage exposure model, which we developed and validated here. We also used the DOR-selective agonist TAN-67 and antagonist naltrindole to examine the involvement of DORs in AWiMA, which was measured using a von Frey model of mechanical allodynia. We created a robust model of alcohol withdrawal-induced anxiety and mechanical allodynia by orally gavaging mice with 3g/kg alcohol for three weeks. AWiMA was exacerbated and prolonged in DOR knockout mice as well as by pharmacological blockade of DORs compared to control mice. However, analgesia induced by TAN-67 was attenuated during withdrawal in alcohol-gavaged mice. DORs appear to play a protective role in the establishment of AWiMA. Our current results indicate that DORs could be targeted to prevent or reduce the development of AWiMA during alcohol use; however, DORs may be a less suitable target to treat AWiMA during active withdrawal. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  12. Sex differences in the effects of adolescent social deprivation on alcohol consumption in μ-opioid receptor knockout mice.

    Science.gov (United States)

    Moriya, Yuki; Kasahara, Yoshiyuki; Hall, F Scott; Sakakibara, Yasufumi; Uhl, George R; Tomita, Hiroaki; Sora, Ichiro

    2015-04-01

    Evidence based on clinical and experimental animal studies indicates that adolescent social deprivation influences alcohol consumption in a sex-dependent manner, perhaps by influencing stress responses. However, the mechanisms underlying the interaction between these phenomena remain to be elucidated. Since the μ-opioid receptor (MOP) has been reported to have key roles in social stress responses as well as the reinforcing/addictive effects of ethanol, MOP is a candidate molecule that may link adolescent social deprivation and subsequent alterations in alcohol consumption. To evaluate the involvement of MOP and social isolation-induced changes in alcohol consumption, as well as the effect of sex differences on responses to social isolation, alcohol consumption was assessed using a two-bottle home-cage consumption procedure (8 % ethanol vs. water) in MOP knockout (MOP-KO) and wild type (WT) mice of both sexes exposed to adolescent social deprivation or reared socially. Isolation rearing had no effects upon alcohol consumption of WT mice, whereas it significantly altered alcohol consumption in both male and female MOP-KO mice. Interestingly, social isolation affected ethanol consumption differently in male and female mice. Ethanol consumption was increased in male MOP-KO mice, but decreased in female MOP-KO mice, by isolation rearing. These results indicate that disturbances of MOP function influence the effects of isolation rearing on ethanol consumption in a sex-dependent manner. Consequently, this suggests the possibility that genetic variation that influences MOP function may have differential roles in alcoholism in men and women, and alcoholism treatments that target MOP function may be differentially effective in males and females.

  13. Flow Cytometry-Based Bead-Binding Assay for Measuring Receptor Ligand Specificity

    NARCIS (Netherlands)

    Sprokholt, Joris K.; Hertoghs, Nina; Geijtenbeek, Teunis B. H.

    2016-01-01

    In this chapter we describe a fluorescent bead-binding assay, which is an efficient and feasible method to measure interaction between ligands and receptors on cells. In principle, any ligand can be coated on fluorescent beads either directly or via antibodies. Binding between ligand-coated beads

  14. Discovery and mapping of an intracellular antagonist binding site at the chemokine receptor CCR2

    DEFF Research Database (Denmark)

    Zweemer, Annelien J M; Bunnik, Julia; Veenhuizen, Margo

    2014-01-01

    be divided into two groups with most likely two topographically distinct binding sites. The aim of the current study was to identify the binding site of one such group of ligands, exemplified by three allosteric antagonists, CCR2-RA-[R], JNJ-27141491, and SD-24. We first used a chimeric CCR2/CCR5 receptor...

  15. Apparent non-statistical binding in a ditopic receptor for guanosine

    NARCIS (Netherlands)

    Likhitsup, Asawin; Deeth, Robert J.; Otto, Sijbren; Marsh, Andrew

    2009-01-01

    Analysis of stepwise association constants for guests binding to more than one site in a receptor is expected to give a ratio of the first association constant to the second of about 4 : 1 on statistical grounds (since a second guest should have an equal chance of binding to a different site on the

  16. A non-multimacrocyclic heteroditopic receptor that cooperatively binds and effectively extracts KAcO salt.

    Science.gov (United States)

    Zakrzewski, Maciej; Kwietniewska, Natalia; Walczak, Wojciech; Piątek, Piotr

    2018-06-06

    Prepared in only three synthetic steps, a non-multimacrocyclic heteroditopic receptor binds potassium salts of halide