Modulation of neuropeptide FF receptors by guanine nucleotides and cations in membranes of rat brain and spinal cord.
Using a radioligand binding assay, we examined ionic modulation and G protein coupling of neuropeptide FF (NPFF) receptors in membranes of rat brain and spinal cord. We found that NaCl (but not KCl or LiCl) and MgCl2 increased specific 125I-YLFQPQRFamide (125I-Y8Fa) binding to NPFF receptors in both tissues in a dose-dependent manner, with optimal conditions being 60 mM NaCl and 1 mM MgCl2. Guanine nucleotides dose-dependently inhibited specific 125I-Y8Fa binding to rat brain and spinal cord membranes with maximal effects of 64 +/- 6 and 71 +/- 2%, respectively. The order of potency was nonhydrolyzable GTP analogues > GTP > or = GDP >> GMP, ATP. The guanine nucleotide inhibition was observed in the absence and presence of NaCl and MgCl2. The mechanism of inhibition in spinal cord membranes appeared to be a reduction in the number of NPFF receptors; in one experiment, control KD and Bmax values were 0.068 nM and 7.2 fmol/mg of protein, respectively, and with 0.1 microM guanylylimidodiphosphate the respective values were 0.081 nM and 4.9 fmol/mg, a 32% reduction in receptor number. Similar results were obtained with guanosine 5'-O-(3-thiotriphosphate). Our data suggest that 125I-Y8Fa binding sites in rat CNS are G protein-coupled NPFF receptors regulated by GTP and cations.
Laboratory of Biochemical Genetics, NIMH Neuroscience Center, St. Elizabeth's Hospital, Washington, D.C.
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Yang
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002412:Cations / Q000494:pharmacology*; D006150:Guanine Nucleotides / Q000494:pharmacology*; D008566:Membranes / Q000378:metabolism; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism*; D051381:Rats; D011956:Receptors, Cell Surface / Q000187:drug effects* / Q000378:metabolism; D013116:Spinal Cord / Q000378:metabolism*
D002412:Cations; D006150:Guanine Nucleotides; D009479:Neuropeptides; D009842:Oligopeptides; D011956:Receptors, Cell Surface; C080740:3-iodotyrosyl-leucyl-phenylalanyl-glutaminyl-prolyl-glutaminyl-arginyl-phenylalaninamide; C053651:phenylalanyl-leucyl-phenylalanyl-glutaminyl-prolyl-glutaminyl-arginyl-phenylalaninamide