Physical Association between Neuropeptide FF and μ-Opioid Receptors as a Possible Molecular Basis for Anti-opioid Activity

Abstract

Neuropeptide FF (NPFF) modulates the opioid system by exerting functional anti-opioid activity on neurons, the mechanism of which is unknown. By using a model of SH-SY5Y cells, we recently postulated that anti-opioid activity likely takes place upstream from the signaling cascade, suggesting that NPFF receptors could block opioid receptors by physical interaction. In the present study, fluorescence techniques were used to monitor the physical association and the dynamic of NPFF 2 and -opioid (

Heterologous Regulation of Mu-Opioid (MOP) Receptor Mobility in the Membrane of SH-SY5Y Cells

“…The specific MOP receptor radioligand [ (Table 1), revealing a specific dynamic confinement for each receptor. As previously observed (18,46), the MOP-YFP receptor was found to diffuse …”

“…For cells transfected with the human NPFF 2 receptor (SH2-D9 cells), the medium was supplemented with 400 g/ml G418 (Invitrogen); for cells expressing the human NPFF 2 receptor in addition to the YFP tagged MOP receptor ((SH 2 -D9)MOP-YFP cells), 2 g/ml blasticidin (Cayla, France) was further added to maintain selection (18). Cells expressing the NPFF 2 -YFP receptor were cultured in the presence of 5 g/ml blasticidin.…”

“…The heteromerization of the MOP receptor with other opioid receptors, DOP, KOP, and NOP (9 -12) or with non-opioid receptors such as ␣ 2 -adrenergic (13)(14)(15), somatostatin SST 2A (16) or substance P NK1 (17) receptors has been reported. By using FRET and immunoprecipitation methods, we have also provided evidence for a molecular interaction between the MOP receptor and the receptor for the opioid-modulating peptide neuropeptide FF (NPFF) (18). NPFF receptor activation not only promotes MOP/NPFF receptor interaction as revealed by an increased FRET signal but also induces MOP receptor desensitization by a new mechanism involving a G protein-coupled receptor kinase GRK2-dependent transphosphorylation of the human MOP receptor Ser 377 (19).…”

“…In particular, SPT and FRAP experiments performed at variable radius (vrFRAP) have revealed a confined mode of diffusion for the MOP receptor in microdomains with a radius size of ϳ1 m (18,38,45,46). The domain size and/or diffusion parameters of the MOP receptor are modulated upon agonist or antagonist administration (45)(46)(47), suggesting that a selective pharmacological response could be associated with a specific dynamic organization of the receptors in the membrane.…”

“…These parameters are also influenced by changes in membrane composition such as cholesterol depletion or caveolae enrichment (47,48) or by co-expression of another receptor such as the DOP receptor (49). Interestingly, we have previously reported that activation of the NPFF 2 receptor suppresses the confinement of the MOP receptor and increases its lateral mobility (18). This loss of constraint was suspected to impair interaction of the MOP receptor with signaling partners, contributing to the reduced opioid response in the presence of the anti-opioid peptide.…”