Abstract
Nucleus raphe magnus (NRM) sends the projection to spinal dorsal horn and inhibits nociceptive transmission. Analgesic effect produced by mu-opioid receptor agonists including morphine partially results from activating the NRM-spinal cord pathway. It is generally believed that mu-opioid receptor agonists disinhibit spinally projecting neurons of the NRM and produce analgesia by hyperpolarizing GABAergic interneurons. In the present study, whole-cell patch-clamp recordings combined with single-cell RT-PCR analysis were used to test the hypothesis that DAMGO ([D-Ala(2),N-methyl-Phe(4),Gly-ol(5)]enkephalin), a specific mu-opioid receptor agonist, selectively hyperpolarizes NRM neurons expressing mRNA of glutamate decarboxylase (GAD(67)). Homologous desensitization of mu-opioid receptors in NRM neurons could result in the development of morphine-induced tolerance. G protein-coupled receptor kinase (GRK) is believed to mediate mu-opioid receptor desensitization in vivo. Therefore, we also investigated the involvement of GRK in mediating homologous desensitization of DAMAMGO-induced electrophysiological effects on NRM neurons by using two experimental strategies. First, single-cell RT-PCR assay was used to study the expression of GRK2 and GRK3 mRNAs in individual DAMGO-responsive NRM neurons. Whole-cell recording was also performed with an internal solution containing the synthetic peptide, which corresponds to G(betagamma)-binding domain of GRK and inhibits G(betagamma) activation of GRK. Our results suggest that DAMGO selectively hyperpolarizes NRM GABAergic neurons by opening inwardly rectifying K(+) channels and that GRK2 mediates short-term homologous desensitization of mu-opioid receptors in NRM GABAergic neurons.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Analgesics, Opioid / pharmacology
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Animals
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Binding Sites
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Cyclic AMP-Dependent Protein Kinases / biosynthesis
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Cyclic AMP-Dependent Protein Kinases / genetics
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Cyclic AMP-Dependent Protein Kinases / physiology*
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Drug Tolerance / physiology
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Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / pharmacology
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G-Protein-Coupled Receptor Kinase 2
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G-Protein-Coupled Receptor Kinase 3
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Glutamate Decarboxylase / biosynthesis
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Ion Transport / drug effects
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Membrane Potentials / drug effects
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Molecular Sequence Data
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Nerve Tissue Proteins / biosynthesis
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Nerve Tissue Proteins / genetics
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Nerve Tissue Proteins / physiology*
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Neurons / drug effects
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Neurons / physiology
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Oligopeptides / chemical synthesis
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Oligopeptides / pharmacology
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Pain / physiopathology
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Patch-Clamp Techniques
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Phosphorylation
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Potassium / metabolism
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Potassium Channels / metabolism
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Potassium Channels, Inwardly Rectifying*
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Protein Processing, Post-Translational
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Protein Serine-Threonine Kinases / biosynthesis
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Protein Serine-Threonine Kinases / genetics
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Protein Structure, Tertiary
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Protein Transport / drug effects
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RNA, Messenger / biosynthesis
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RNA, Messenger / genetics
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Raphe Nuclei / drug effects
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Raphe Nuclei / enzymology
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Raphe Nuclei / physiology*
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Raphe Nuclei / physiopathology
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Rats
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Rats, Sprague-Dawley
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Receptors, Opioid, mu / agonists
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Receptors, Opioid, mu / physiology*
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Reverse Transcriptase Polymerase Chain Reaction
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Signal Transduction / drug effects
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Spinal Cord / physiopathology
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beta-Adrenergic Receptor Kinases
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gamma-Aminobutyric Acid / physiology*
Substances
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Analgesics, Opioid
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Nerve Tissue Proteins
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Oligopeptides
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Potassium Channels
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Potassium Channels, Inwardly Rectifying
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RNA, Messenger
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Receptors, Opioid, mu
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Enkephalin, Ala(2)-MePhe(4)-Gly(5)-
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gamma-Aminobutyric Acid
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Protein Serine-Threonine Kinases
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Cyclic AMP-Dependent Protein Kinases
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G-Protein-Coupled Receptor Kinase 3
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Grk2 protein, rat
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Grk3 protein, rat
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beta-Adrenergic Receptor Kinases
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G-Protein-Coupled Receptor Kinase 2
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Glutamate Decarboxylase
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Potassium