Abstract
Morphine tolerance in vivo is reduced following blockade of the orphanin FQ/nociceptin (OFQ/N)/opioid receptor-like 1 (ORL1) receptor system, suggesting that OFQ/N contributes to the development of morphine tolerance. We previously reported that a 60-min activation of ORL1 receptors natively expressed in BE(2)-C cells desensitized both μ and ORL1 receptor-mediated inhibition of cAMP. Investigating the mechanism(s) of OFQ/N-mediated μ and ORL1 receptor cross-talk, we found that pretreatment with the protein kinase C inhibitor, chelerythrine chloride (1 μM), blocked OFQ/N-mediated homologous desensitization of ORL1 and heterologous desensitization of μ opioid receptors. Furthermore, depletion of PKC by 12-O-tetradecanoylphorbol-13-acetate exposure (48 h, 1 μM) also prevented OFQ/N-mediated μ and ORL1 desensitization. OFQ/N pretreatment resulted in translocation of PKC-α, G protein-coupled receptor kinase 2 (GRK2) and GRK3 from the cytosol to the membrane, and this translocation was also blocked by chelerythrine. Reduction of GRK2 and GRK3 levels by antisense, but not sense DNA treatment blocks ORL1 and μ receptor desensitization. This suggests that PKC-α is required for GRK2 and GRK3 translocation to the membrane, where GRK can inactivate ORL1 and μ opioid receptors upon rechallenge with the appropriate agonist. Our results demonstrate for the first time the involvement of conventional PKC isozymes in OFQ/N-induced μ-ORL1 cross-talk, and represent a possible mechanism for OFQ/N-induced anti-opioid actions.
Footnotes
-
This work was supported in part by work sponsored by a National Research Service Award (DA14171) to C.D.M. and grants from the Department of Health and Human Services (DA10738) and the Texas Advanced Research Program (003652-0114-1999) to K.M.S.
-
DOI: 10.1124/jpet.102.033159
- Abbreviations:
- OFQ/N
- orphanin FQ/nociceptin
- ORL1
- opioid receptor-like 1
- DAMGO
- [d-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin
- ERK1/2
- extracellular-regulated kinases 1 and 2
- PKC
- protein kinase C
- MEK-1
- mitogen-activated protein kinase kinase
- GRK
- G-protein receptor kinase
- TPA
- 12-O-tetradecanoylphorbol-13-acetate
- 4αPDD
- 4α-phorbol-12,13-didecanoate
- HBSS
- Hanks' balanced salt solution
- PAGE
- polyacrylamide gel electrophoresis
- PBS
- phosphate-buffered saline
- BSA
- bovine serum albumin
- PMSF
- phenylmethylsulfonyl fluoride
- ODN
- oligodeoxynucleotide
- TBS/T
- Tris-buffered saline/Tween 20
- GAPDH
- glyceraldehyde-3-phosphate dehydrogenase
- ANOVA
- analysis of variance
- DAG
- diacylglycerol
- PKA
- protein kinase A
- PD98059
- 4H-1-benzopyran-4-one 2-(2-amino-3-methoxy phenyl)
- H-89
- N-[2-((p-bromocinnamyl)amino)ethyl]-5-isoquinoline sulfonamide
- Received January 16, 2002.
- Accepted April 23, 2002.
- The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|