Characterization of opioid binding sites in rat spinal cord

Paola Petrillo, Jan Kowalski, Massimo Sbacchi, Alessandra Tavani

Research output: Contribution to journalArticlepeer-review


Binding sites were characterized in rat whole spinal cord crude membrane preparations using selective labelling techniques with multiple methods of mathematical analysis of experimental curves. Mathematical analysis of single [3H]-[D-Ala2, MePhe4, Gly-ol5] enkephalin (DAGO) saturation curves suggested binding of the [3H]-ligand at one site, while displacement curves of low concentrations of [3H]-DAGO with selective μligands indicated the presence of high- and low-affinity sites. All the [3H]-DAGO curves processed simultaneously by LIGAND analysis showed the presence of high (27% and low (73% affinity components, with a total Bmax of 3.19 pmol/g tissue. Eighty percent of [3H]-[D-Ala2, D-Leu5] enkephalin (DADLE) binding was displaced by DAGO with high affinity, indicating that a high percentage of [3H]-DADLE binding was at μsites. Saturation curves of [3H]-DADLE after inhibition of μsites by unlabelled DAGO (∂sites) were monophasic with non-linear fitting analysis and the Bmax was 0.90 pmol/g tissue. Most mathematical analysis of single saturation curves of [3H]-(-)-bremazocine indicated binding at more than one site. DAGO, DADLE, U-69,593 and PD 117302 displaced 0.15 nM of [3H]-(-)-bremazocine biphasically: the percentages of displacement calculated with the non-linear fitting program were respectively 50 (μsites), 64 (μplus;part;sites), 18 and 25 (kappa-sites). Haloperidol displaced [3H]-(-)-bremazocine only at μM concentrations, suggesting no binding at sigma-sites. In the presence of 225 nM of DAGO, DADLE displaced only 21% of [3H]-(-)-bremazocine 0.15 nM binding (∂sites). Most mathematical analysis of saturation curves of [3H]-(-)-bremazocine, after inhibition of binding at μ and ∂sites by DAGO and DADLE, still indicated binding at more

Original languageEnglish
Pages (from-to)39-57
Number of pages19
JournalJournal of Receptors and Signal Transduction
Issue number1
Publication statusPublished - 1992

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Pharmacology


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