Library

X
feed icon rss

Your email was sent successfully. Check your inbox.

An error occurred while sending the email. Please try again.

Proceed reservation?

Export
  • 1
    Electronic Resource
    Electronic Resource
    Regulation of Cyclic AMP by the μ-Opioid Receptor in Human Neuroblastoma SH-SY5Y Cells (1990)
    Oxford, UK : Blackwell Publishing Ltd
    Journal of neurochemistry 55 (1990), S. 0 
    Details
    ISSN: 1471-4159
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Abstract: The human neuroblastoma clonal cell line SH-SY5Y expresses both μ- and δ-opioid receptors (ratio ∼4.5:1). Differentiation with retinoic acid (RA) was previously shown to enhance the inhibition of adenylyl cyclase (AC) by μ-opioid agonists. We tested here the inhibition of cyclic AMP (cAMP) accumulation by morphine under a variety of conditions: after stimulation with prostaglandin E1 (PGE,), forskolin, and vasoactive intestinal peptide (VIP), both in the presence and in the absence of the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX). Morphine inhibition of the forskolin cAMP response (∼65%) was largely unaffected by the presence of IBMX. In contrast, deletion of IBMX enhanced morphine's inhibition of the PGE, and VIP cAMP response from ∼50 to ∼80%. The use of highly μ- and δ-selective agents confirmed previous results that inhibition of cAMP accumulation by opioids is mostly μ, and not δ, receptor mediated in SH-SY5Y cells, regardless of the presence or absence of IBMX. Because of the large morphine inhibition and the high cAMP levels even in the absence of IBMX, PGE,-stimulated, RA-differentiated SHSY5Y cells were subsequently used to study narcotic analgesic tolerance and dependence in vitro. Upon pretreatment with morphine over 12h, a fourfold shift of the PGE,-morphine dose-response curve was observed, whether or not IBMX was added. However, μ-opioid receptor number and affinity to the μ-selective [D-Ala2, N-Me-Phe4, Gly5-ol]enkephalin were largely unaffected, and Na+- and guanyl nucleotide-induced shifts of morphine-[3H]naloxone competition curves were unchanged. Further, morphine pretreatment elicited an AC rebound effect, causing higher cAMP accumulation after the drug was removed from the medium or acutely antagonized by naloxone. These results document biochemical correlates of μ-opiate tolerance and dependence in SH-SY5Y cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1471-4159.1990.tb03151.x
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 2
    Electronic Resource
    Electronic Resource
    Phosphorylation and Agonist-Specific Intracellular Trafficking of an Epitope-Tagged μ-Opioid Receptor Expressed in HEK 293 Cells (1995)
    Oxford, UK : Blackwell Science Ltd
    Journal of neurochemistry 65 (1995), S. 0 
    Details
    ISSN: 1471-4159
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Abstract: We expressed the cloned μ-opioid receptor (μR) in high abundance (5.5 × 106 sites/cell) with an amino-terminal epitope tag (EYMPME) in human embryonic kidney 293 cells. The epitope-tagged receptor (EE-μR) was similar to the untagged μR in ligand binding and agonist-dependent inhibition of cyclic AMP accumulation. By confocal microscopy, the labeled receptor was shown to be largely confined to the plasma membrane. Pretreatment with morphine failed to affect the cellular distribution of the receptor as judged by immunofluorescence and tracer binding studies. In contrast, exposure to the μ-specific peptide agonist [d-Ala2,MePhe4,Glyol5]enkephalin (DAMGO) caused strong labeling of endocytic vesicles, indicating extensive agonist-induced cellular redistribution of EE-μR. Tracer binding studies suggested partial net internalization and a small degree of down-regulation caused by DAMGO. EE-μR-containing membranes were solubilized in detergent [3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate] and immunoprecipitated by an anti-epitope monoclonal antibody. Immunoblotting revealed a prominent band at ∼70 kDa with weaker bands at ∼65 kDa. EE-μR was labeled with [γ-32P]ATP in permeabilized cells, immunoprecipitated, and analyzed by polyacrylamide gel electrophoresis autoradiography. A prominent band at 65–70 kDa indicated the presence of basal receptor phosphorylation occurring in the absence of agonist, which was enhanced ∼1.8-fold with the addition of morphine. In conclusion, intracellular trafficking of the μR appears to depend on the agonist, with morphine and DAMGO having markedly different effects. Unlike other G protein-coupled receptors, basal phosphorylation is substantial, even in the absence of agonist.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1471-4159.1995.65041636.x
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 3
    Electronic Resource
    Electronic Resource
    A Constitutively Internalizing and Recycling Mutant of the μ-Opioid Receptor (1997)
    Oxford, UK : Blackwell Science Ltd
    Journal of neurochemistry 68 (1997), S. 0 
    Details
    ISSN: 1471-4159
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Abstract: Internalization and recycling of G protein-coupled receptors (GPCRs), such as the μ-opioid receptor, largely depend on agonist stimulation, whereas certain other receptor types recycle constitutively, e.g., the transferrin receptor. To investigate structural domains involved in μ-opioid receptor internalization, we constructed two truncation mutants bracketing a Ser/Thr-rich domain (354ThrSerSerThrIleGluGlnGlnAsn362) unique to the C-terminus of the μ-opioid receptor (mutants Trunc354 and Trunc363). Ligand binding did not differ substantially, and G protein coupling was slightly lower for these μ-receptor constructs, in particular for Trunc363. To permit localization of the receptor by immunocytochemistry, an epitope tag was added to the N-terminus of the wildtype and mutant receptors. Both the wild-type μ-opioid receptor and Trunc363 resided largely at the plasma membrane and internalized into vesicles upon stimulation with the agonist [d-Ala2,N-Me-Phe4,Gly-ol5]-enkephalin. Internalization occurred into vesicles that contain transferrin receptors, as shown previously, as well as clathrin, but not caveolin. In contrast, even without any agonist present, Trunc354 colocalized in intracellular vesicles with clathrin and transferrin receptors, but not caveolin. On blocking internalization by hyperosmolar sucrose or acid treatment, Trunc354 translocated to the plasma membrane, indicating that the mutant internalized into clathrin-coated vesicles and recycled constitutively. Despite agonist-independent internalization of Trunc354, basal G protein coupling was not elevated, suggesting distinct mechanisms for coupling and internalization. Furthermore, a portion of the C-terminus, particularly the Ser/Thr domain, appears to suppress μ-receptor internalization, which can be overcome by agonist stimulation. These results demonstrate that a mutant GPCR can be constructed such that internalization, normally an agonist-dependent process, can occur spontaneously without concomitant G protein activation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1471-4159.1997.68062395.x
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 4
    Electronic Resource
    Electronic Resource
    Residues Specifically Involved in Down-Regulation but Not Internalization of the m1 Muscarinic Acetylcholine Receptor (1997)
    Oxford, UK : Blackwell Science Ltd
    Journal of neurochemistry 68 (1997), S. 0 
    Details
    ISSN: 1471-4159
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Abstract: Human m1 muscarinic acetylcholine receptor mutants were screened to determine receptor domains and cellular pathways relevant to down-regulation. Mutations in the second intracellular loop and the junctions of the third intracellular loop of the receptor, where a role for receptor activation or internalization had been previously demonstrated in HEK293 cells, were selected for this study. To assess receptor down-regulation, the m1 receptor mutants were transfected into Chinese hamster ovary cells. Because receptor internalization is expected to precede down-regulation, mutants displaying intact internalization were selected to permit interpretation of mutational effects on down-regulation alone. Four mutations were identified that specifically impaired down-regulation without altering receptor internalization: V127A, I211A, E360A, and K362A. The results define new receptor domains in the second intracellular loop and the junctions of the third intracellular loop that are involved in down-regulation. These same four mutants were also defective in signaling via the phospholipase C and the adenylyl cyclase pathways and in G protein activation, as measured by [35S]GTPγS binding. However, the level of second messenger stimulation correlated poorly with the extent of down-regulation. In summary, several mutations of the m1 receptor selectively affect down-regulation, demonstrating that internalization and down-regulation represent distinct events driven by different cellular mechanisms.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1471-4159.1997.68020601.x
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 5
    Electronic Resource
    Electronic Resource
    Antibody to Epitope Tag Induces Internalization of Human Muscarinic Subtype 1 Receptor (1998)
    Oxford, UK : Blackwell Science Ltd
    Journal of neurochemistry 70 (1998), S. 0 
    Details
    ISSN: 1471-4159
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Abstract: We have studied the effect of an antibody against the epitope EYMPME on the internalization of the human muscarinic cholinergic receptor hm1 tagged with the epitope at the amino terminus. The antibody to the tag induces internalization of the hm1 receptor within minutes after exposure of human embryonic kidney 293 cells transfected with the tagged receptor. This antibody-induced internalization is reversible following removal of the antibody. In contrast to hm1 internalization induced by the agonist carbachol, internalization induced by antibody is not blocked by the muscarinic antagonist atropine. The mechanism of antibody-mediated internalization does not appear to involve receptor dimerization by the antibody, as Fab fragments derived from the antibody also induce internalization. The pathway of antibody-induced internalization, similar to the agonist-induced process, is mediated by clathrin-coated vesicles. Furthermore, antibody treatment does not result in any second messenger production, as measured by phosphoinositide accumulation. Our data show that internalization of a G protein-coupled receptor can be triggered by interaction of the amino terminus of the receptor with an exogenous ligand and can occur independently of second messenger production. This result suggests that the receptor can exist in multiple conformations, each mediating distinct downstream events.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1471-4159.1998.70010113.x
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 6
    Electronic Resource
    Electronic Resource
    Mapping the Ligand Binding Pocket of the Human Muscarinic Cholinergic Receptor Hml: Contribution of Tyrosine-82 (1992)
    Springer
    Pharmaceutical research 9 (1992), S. 1644-1647 
    Details
    ISSN: 1573-904X
    Keywords: muscarinic cholinergic Hm1 receptor ; site-directed mutagenesis ; receptor binding pocket
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The ligand binding pocket of many G protein-coupled receptors is thought to be located within the core formed by their seven transmembrane domains (TMDs). Previous results suggested that muscarinic antagonists bind to a pocket located toward the extracellular region of the TMDs, primarily at TMDs 2, 3, 6, and 7. Tyrosine-82 (Y82) is located in TMD2 only one helical turn from the presumed membrane surface of Hm1, whereas a phenylalanine (F124) is found in the equivalent position of the closely related Hm3. In order to determine the contribution of Y82 to Hm1 ligand binding and selectivity versus Hm3, we constructed the point mutation Y82 F of Hm1 and measured binding affinities of various ligands, with 3H-N-methylscopolamine (3H-NMS) as the tracer. The Hm1 wild-type receptor and the Y82F mutant were transfected into human embryonic kidney U293 cells. Whereas the affinities of NMS, carbachol, and atropine were either unchanged (carbachol) or enhanced by less than twofold (atropine and NMS), the affinity of the Hm1-selective piren-zepine was reduced threefold by the Y82 F mutation. These changes parallel affinity differences of Hm1 and Hm3, indicating that the Y82 F mutation affects the binding pocket and that Y82 contributes to the binding selectivity among closely related muscarinic receptors.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1015885029612
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 7
    Electronic Resource
    Electronic Resource
    Morphine Impurity with Opioid Activity Is Identified as 10α-Hydroxymorphine (1990)
    Springer
    Pharmaceutical research 7 (1990), S. 1205-1207 
    Details
    ISSN: 1573-904X
    Keywords: morphine, hydroxylated ; 10α-hydroxymorphine ; morphine, impurity ; morphine, hydroxylated, activity in guinea pig ileum ; opioid receptor binding, morphine impurity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1015957031449
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 8
    Electronic Resource
    Electronic Resource
    Structure and Function of G Protein Coupled Receptors (1990)
    Springer
    Pharmaceutical research 7 (1990), S. 1213-1221 
    Details
    ISSN: 1573-904X
    Keywords: G protein coupled receptors ; neurotransmitter receptors ; hormone receptors ; adrenergic receptors ; muscarinic cholinergic receptors ; second messengers
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The G protein coupled receptors (GPC-Rs) comprise a large superfamily of genes encoding numerous receptors which all show common structural features, e.g., seven putative membrane spanning domains. Their biological functions are extremely diverse, ranging from vision and olfaction to neuronal and endocrine signaling. The GPC-Rs couple via multiple G proteins to a growing number of recognized second messenger pathway, e.g., cAMP and phosphatidyl inositol turnover. This review summarizes our current knowledge of the molecular mechanisms by which the GPC-Rs activate second messenger systems, and it addresses their regulation and structure.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1015969301407
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
Close ⊗
This website uses cookies and the analysis tool Matomo. More information can be found here...