Functional differences between NPFF1 and NPFF2 receptor coupling: high intrinsic activities of RFamide-related peptides on stimulation of [35S]GTPgammaS binding.
By using an optimized [(35)S]GTPgammaS binding assay, the functional activities (potency and efficacy) of peptides belonging to three members of the RFamide family; Neuropeptide FF (NPFF), prolactin-releasing peptide (PrRP) and 26RFamide, were investigated on NPFF(1) and NPFF(2) receptors stably expressed in Chinese Hamster Ovary (CHO) cells. Despite their large differences in affinity and selectivity, all analogues tested behaved as agonists toward NPFF(1) and NPFF(2) receptors. High NaCl concentration in the assay strongly increased the efficacy toward NPFF(2) receptors and augmented differences among agonists. In low sodium conditions, whereas the potencies of agonists correlated with their affinities for NPFF(1) receptors, NPFF(2) receptors exhibited an extraordinary activity since all compounds tested displayed EC(50) values of GTPgammaS binding lower than their K(I) values. Comparisons of functional values between NPFF(1) and NPFF(2) receptors revealed unexpected potent selective NPFF(2) agonists especially for the PLRFamide and the VGRFamide sequences. By using blocker peptides, we also show that Galpha(i3) and Galpha(s) are the main transducers of NPFF(1) receptors while NPFF(2) are probably coupled with Galpha(i2), Galpha(i3), Galpha(o) and Galpha(s) proteins. Our data indicate that NPPF(1) and NPFF(2) receptors are differently coupled to G proteins in CHO cells.
Gouardères
C
C
Institut de Pharmacologie et de Biologie Structurale, CNRS UMR 5089, 205 Route de Narbonne, 31077 Toulouse Cedex, France.
Mazarguil
H
H
Mollereau
C
C
Chartrel
N
N
Leprince
J
J
Vaudry
H
H
Zajac
J-M
JM
D000818:Animals; D016466:CHO Cells; D002462:Cell Membrane / Q000000981:diagnostic imaging; D006224:Cricetinae; D003412:Cricetulus; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism*; D006801:Humans; D007554:Isotopes / Q000493:pharmacokinetics; D009479:Neuropeptides / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D011877:Radionuclide Imaging; D018013:Receptors, Neuropeptide / Q000378:metabolism*; D012503:Saponins / Q000494:pharmacology
D044385:GTP-Binding Protein alpha Subunits; D007554:Isotopes; D009479:Neuropeptides; C416987:RFamide peptide; D018013:Receptors, Neuropeptide; D012503:Saponins; C083929:neuropeptide FF receptor; D016244:Guanosine 5'-O-(3-Thiotriphosphate)