Current address: Department of Anaesthesiology, University of Hirosaki School of Medicine, 5 Zaifu-Cho, Hirosaki 036-8562, Japan
Partial agonist behaviour depends upon the level of nociceptin/orphanin FQ receptor expression: studies using the ecdysone-inducible mammalian expression system
Article first published online: 30 JAN 2009
2003 British Pharmacological Society
British Journal of Pharmacology
Volume 140, Issue 1, pages 61–70, September 2003
How to Cite
McDonald, J., Barnes, T. A., Okawa, H., Williams, J., Calo', G., Rowbotham, D. J. and Lambert, D. G. (2003), Partial agonist behaviour depends upon the level of nociceptin/orphanin FQ receptor expression: studies using the ecdysone-inducible mammalian expression system. British Journal of Pharmacology, 140: 61–70. doi: 10.1038/sj.bjp.0705401
- Issue published online: 30 JAN 2009
- Article first published online: 30 JAN 2009
- (Received April 8, 2003, Revised May 27, 2003, Accepted June 3, 2003)
- Nociceptin/orphanin FQ;
- nociceptin receptor;
- ecdysone-inducible expression;
- partial agonists;
- GTPγ35S binding;
Partial agonism is primarily dependent upon receptor density and coupling efficiency. As these parameters are tissue/model dependent, intrinsic activity in different tissues can vary. We have utilised the ecdysone-inducible expression system containing the human nociceptin/orphanin FQ (N/OFQ) peptide receptor (hNOP) expressed in Chinese hamster ovary cells (CHOINDhNOP) to examine the activity of a range of partial agonists in receptor binding, GTPγ35S binding and inhibition of adenylyl cyclase studies.
Incubation of CHOINDhNOP cells with ponasterone A (PON) induced hNOP expression ([leucyl-3H]N/OFQ binding) of 24, 68, 191 and 1101 fmol mg−1 protein at 1, 2, 5 and 10 μM PON, respectively. At 191 fmol mg−1, protein hNOP pharmacology was identical to that reported for other traditional expression systems.
pEC50 values for GTPγ35S binding ranged from 7.23 to 7.72 (2–10 μM PON) for the partial agonist [Phe1ψ(CH2–NH)Gly2]N/OFQ(1–13)–NH2 ([F/G]N/OFQ(1–13)–NH2) and 8.12–8.60 (1–10 μM PON) for N/OFQ(1–13)–NH2 and Emax values (stimulation factor relative to basal) ranged from 1.51 to 3.21 (2–10 μM PON) for [F/G]N/OFQ(1–13)–NH2 and 1.28–6.95 (1–10 μM) for N/OFQ(1–13)–NH2. Intrinsic activity of [F/G]N/OFQ(1–13)–NH2 relative to N/OFQ(1–13)–NH2 was 0.3–0.5. [F/G]N/OFQ(1–13)–NH2 did not stimulate GTPγ35S binding at 1 μM PON, but competitively antagonised the effects of N/OFQ(1–13)–NH2 with a pKB=7.62.
pEC50 values for cAMP inhibition ranged from 8.26 to 8.32 (2–10 μM PON) for [F/G]N/OFQ(1–13)–NH2 and 9.42–10.35 for N/OFQ(1–13)–NH2 and Emax values (% inhibition) ranged from 19.6 to 83.2 for [F/G]N/OFQ(1–13)–NH2 and 40.9–86.0 for N/OFQ(1–13)–NH2. The intrinsic activity of [F/G]N/OFQ(1–13)–NH2 relative to N/OFQ(1–13)–NH2 was 0.48–0.97.
In the same cellular environment with receptor density as the only variable, we show that the profile of [F/G]N/OFQ(1–13)–NH2 can be manipulated to encompass full and partial agonism along with antagonism.
British Journal of Pharmacology (2003) 140, 61–70. doi:10.1038/sj.bjp.0705401