title: Guanine nucleotide-mediated inhibition of opioid agonist binding. Modulatory effects of ions and of receptor occupancy.
abstract: We have analysed the potency of GTP, GDP and their analogues in reducing [3H]DADLE binding to opioid receptors in NG 108-15 cell membranes. Under conditions where non-specific hydrolysis and transphosphorylation is inhibited, the following rank order of potency was found: GDP greater than or equal to GTP gamma S greater than GTP greater than GDP beta S greater than or equal to GDPNH2 greater than GppNHp much greater than GMP. Remarkably, the slopes for the inhibition curves of GTP, GDP and their thiosubstituted analogues, but not of GDPNH2 and GppNHp, were extremely shallow, indicating either negative cooperativity or the existence of two states for the guanine nucleotide binding proteins, that both can mediate the effect of nucleotides on agonist receptor binding. The potencies of the different guanine nucleotide analogues, except that of GppNHp, were increased by the presence of sodium or chloride ions in the assay medium. Magnesium also affected GTP-mediated inhibition of opioid agonist binding since it decreased the IC50 of the nucleotide and steepened the slope of the inhibition curve. The IC50s of nucleotides and the slopes of their inhibition curves were also dependent on the extent of receptor occupancy by the agonist. From these data we conclude that (1) either diphospho- or triphosphonucleotides can regulate agonist binding. (2) Magnesium, sodium and chloride, by acting at different components of the receptor/G protein complex produce similar effects on nucleotide mediated regulation of agonist binding. (3) A mutual influence exists between receptor occupancy by agonists and G protein-mediated guanine nucleotide effect on the receptor.
authors.affiliation: Department of Neuropharmacology, Max-Planck-Institute for Psychiatry, Planegg-Martinsried, Federal Republic of Germany.
authors.forename: S
authors.initials: S
authors.lastname: Ott
authors.forename: T
authors.initials: T
authors.lastname: Costa
mesh_terms: D002460:Cell Line; D002712:Chlorides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004723:Endorphins / Q000378:metabolism; D004743:Enkephalin, Leucine / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors / Q000378:metabolism; D016308:Enkephalin, Leucine-2-Alanine; D019204:GTP-Binding Proteins / Q000502:physiology; D006150:Guanine Nucleotides / Q000502:physiology*; D006868:Hydrolysis; D008274:Magnesium / Q000494:pharmacology; D010766:Phosphorylation; D011957:Receptors, Opioid / Q000187:drug effects / Q000502:physiology*; D012964:Sodium / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D002712:Chlorides; D004723:Endorphins; D006150:Guanine Nucleotides; D011957:Receptors, Opioid; D010566:Virulence Factors, Bordetella; D004743:Enkephalin, Leucine; D016308:Enkephalin, Leucine-2-Alanine; D012964:Sodium; D019204:GTP-Binding Proteins; D008274:Magnesium