Partial agonist behaviour depends upon the level of nociceptin/orphanin FQ receptor expression: studies using the ecdysone-inducible mammalian expression system

Br J Pharmacol. 2003 Sep;140(1):61-70. doi: 10.1038/sj.bjp.0705401. Epub 2003 Aug 11.

Abstract

(1) 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, GTPgamma35S binding and inhibition of adenylyl cyclase studies. (2) 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 microm PON, respectively. At 191 fmol mg-1, protein hNOP pharmacology was identical to that reported for other traditional expression systems. (3) pEC50 values for GTPgamma35S binding ranged from 7.23 to 7.72 (2-10 microm PON) for the partial agonist [Phe1psi(CH2-NH)Gly2]N/OFQ(1-13)-NH2 ([F/G]N/OFQ(1-13)-NH2) and 8.12-8.60 (1-10 microm PON) for N/OFQ(1-13)-NH2 and Emax values (stimulation factor relative to basal) ranged from 1.51 to 3.21 (2-10 microm PON) for [F/G]N/OFQ(1-13)-NH2 and 1.28-6.95 (1-10 microm) 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 GTPgamma35S binding at 1 microm PON, but competitively antagonised the effects of N/OFQ(1-13)-NH2 with a pKB=7.62. (4) pEC50 values for cAMP inhibition ranged from 8.26 to 8.32 (2-10 microm 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. (5) 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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CHO Cells
  • Cell Count / methods
  • Cricetinae
  • Cyclic AMP / metabolism
  • Dose-Response Relationship, Drug
  • Gene Expression Regulation / drug effects
  • Gene Expression Regulation / physiology*
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism
  • Humans
  • Nociceptin Receptor
  • Protein Binding / drug effects
  • Protein Binding / physiology
  • Receptors, Opioid / agonists*
  • Receptors, Opioid / biosynthesis*
  • Receptors, Steroid / agonists
  • Receptors, Steroid / biosynthesis*

Substances

  • Receptors, Opioid
  • Receptors, Steroid
  • ecdysone receptor
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Cyclic AMP
  • Nociceptin Receptor