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Opioid peptide receptor studies. 17. Attenuation of chronic morphine effects after antisense oligodeoxynucleotide knock-down of RGS9 protein in cells expressing the cloned Mu opioid receptor.
Synapse. 2004 Jun 01; 52(3):209-17.S

Abstract

RGS proteins are a recently described class of regulators that influence G-protein-mediated signaling pathways. We have shown previously that chronic morphine results in functional uncoupling of the mu opioid receptor from its G protein in CHO cells expressing cloned human mu opioid receptors. In the present study, we examined the effects of morphine treatment (1 microM, 20 h) on DAMGO-stimulated high-affinity [35S]GTP-gamma-S binding and DAMGO-mediated inhibition of forskolin-stimulated cAMP accumulation in HN9.10 cells stably expressing the cloned rat mu opioid receptor, in the absence and presence of the RGS9 protein knock-down condition (confirmed by Western blot analysis). RGS9 protein expression was reduced by blocking its mRNA with an antisense oligodeoxynucleotide (AS-114). Binding surface analysis resolved two [35S]GTP-gamma-S binding sites (high affinity and low affinity sites). In sense-treated control cells, DAMGO-stimulated [35S]GTP-gamma-S binding by increasing the B(max) of the high-affinity site. In sense-treated morphine-treated cells, DAMGO-stimulated [35S]GTP-gamma-S binding by decreasing the high-affinity Kd without changing the B(max). AS-114 significantly inhibited chronic morphine-induced upregulation of adenylate cyclase activity and partially reversed chronic morphine effects as measured by DAMGO-stimulated [35S]GTP-gamma-S binding. Morphine treatment increased the EC50 (6.2-fold) for DAMGO-mediated inhibition of forskolin-stimulated cAMP activity in control cells but not in cells treated with AS-114 to knock-down RGS9. These results provide additional evidence for involvement of RGS9 protein in modulating opioid signaling, which may contribute to the development of morphine tolerance and dependence.

Authors+Show Affiliations

Clinical Psychopharmacology Section, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Baltimore, Maryland 21224, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

15065220

Citation

Xu, Heng, et al. "Opioid Peptide Receptor Studies. 17. Attenuation of Chronic Morphine Effects After Antisense Oligodeoxynucleotide Knock-down of RGS9 Protein in Cells Expressing the Cloned Mu Opioid Receptor." Synapse (New York, N.Y.), vol. 52, no. 3, 2004, pp. 209-17.
Xu H, Wang X, Wang J, et al. Opioid peptide receptor studies. 17. Attenuation of chronic morphine effects after antisense oligodeoxynucleotide knock-down of RGS9 protein in cells expressing the cloned Mu opioid receptor. Synapse. 2004;52(3):209-17.
Xu, H., Wang, X., Wang, J., & Rothman, R. B. (2004). Opioid peptide receptor studies. 17. Attenuation of chronic morphine effects after antisense oligodeoxynucleotide knock-down of RGS9 protein in cells expressing the cloned Mu opioid receptor. Synapse (New York, N.Y.), 52(3), 209-17.
Xu H, et al. Opioid Peptide Receptor Studies. 17. Attenuation of Chronic Morphine Effects After Antisense Oligodeoxynucleotide Knock-down of RGS9 Protein in Cells Expressing the Cloned Mu Opioid Receptor. Synapse. 2004 Jun 1;52(3):209-17. PubMed PMID: 15065220.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Opioid peptide receptor studies. 17. Attenuation of chronic morphine effects after antisense oligodeoxynucleotide knock-down of RGS9 protein in cells expressing the cloned Mu opioid receptor. AU - Xu,Heng, AU - Wang,Xiaoying, AU - Wang,Jun, AU - Rothman,Richard B, PY - 2004/4/6/pubmed PY - 2004/6/18/medline PY - 2004/4/6/entrez SP - 209 EP - 17 JF - Synapse (New York, N.Y.) JO - Synapse VL - 52 IS - 3 N2 - RGS proteins are a recently described class of regulators that influence G-protein-mediated signaling pathways. We have shown previously that chronic morphine results in functional uncoupling of the mu opioid receptor from its G protein in CHO cells expressing cloned human mu opioid receptors. In the present study, we examined the effects of morphine treatment (1 microM, 20 h) on DAMGO-stimulated high-affinity [35S]GTP-gamma-S binding and DAMGO-mediated inhibition of forskolin-stimulated cAMP accumulation in HN9.10 cells stably expressing the cloned rat mu opioid receptor, in the absence and presence of the RGS9 protein knock-down condition (confirmed by Western blot analysis). RGS9 protein expression was reduced by blocking its mRNA with an antisense oligodeoxynucleotide (AS-114). Binding surface analysis resolved two [35S]GTP-gamma-S binding sites (high affinity and low affinity sites). In sense-treated control cells, DAMGO-stimulated [35S]GTP-gamma-S binding by increasing the B(max) of the high-affinity site. In sense-treated morphine-treated cells, DAMGO-stimulated [35S]GTP-gamma-S binding by decreasing the high-affinity Kd without changing the B(max). AS-114 significantly inhibited chronic morphine-induced upregulation of adenylate cyclase activity and partially reversed chronic morphine effects as measured by DAMGO-stimulated [35S]GTP-gamma-S binding. Morphine treatment increased the EC50 (6.2-fold) for DAMGO-mediated inhibition of forskolin-stimulated cAMP activity in control cells but not in cells treated with AS-114 to knock-down RGS9. These results provide additional evidence for involvement of RGS9 protein in modulating opioid signaling, which may contribute to the development of morphine tolerance and dependence. SN - 0887-4476 UR - https://www.unboundmedicine.com/medline/citation/15065220/Opioid_peptide_receptor_studies__17__Attenuation_of_chronic_morphine_effects_after_antisense_oligodeoxynucleotide_knock_down_of_RGS9_protein_in_cells_expressing_the_cloned_Mu_opioid_receptor_ L2 - https://doi.org/10.1002/syn.20019 DB - PRIME DP - Unbound Medicine ER -