title: Affinity labeling of delta opioid receptors by an enkephalin-derivative alkylating agent, DSLET-Mal.
abstract: Opioid binding properties of Tyr-D-Ser-Gly-Phe-Leu-Thr-NH-NH-Gly-Mal (DSLET-Mal), a novel enkephalin-framed affinity label, was determined in rat brain membranes. In competition studies the ligand showed high affinity for the delta opioid sites, labelled by [(3)H][Ile(5,6)]deltorphin II (K(i) = 8 nM), whereas its binding to the mu ([(3)H]DAMGO) and kappa ([(3)H]EKC) sites was weaker. Preincubation of the rat brain membranes with DSLET-Mal at micromolar concentrations resulted in a wash-resistant and dose-dependent inhibition of the [(3)H][Ile(5,6)]deltorphin II binding sites (96% blocking at 10 microM concentration). Intracerebroventricular (ICV) administration of DSLET-Mal reduced the density of delta opioid receptors and had no effect on mu and kappa receptors, as determined by saturation binding studies. [Ile(5, 6)]deltorphin II-stimulated [(35)S]GTPgammaS binding was determined in membrane preparations of different brain areas of the ICV-treated animals. In both frontal cortex and hippocampus DSLET-Mal significantly decreased G protein activation by the delta agonist, having no effect on DAMGO stimulated [(35)S]GTPgammaS binding. DSLET-Mal had qualitatively similar effects on both receptor binding and G protein activation. These characteristics of the compound studied suggest that DSLET-Mal can serve as an affinity label for further studies of the delta-opioid receptors.
authors.affiliation: Institute of Biochemistry, Biological Research Center, Hungarian Academy of Sciences, Szeged, Hungary.
authors.forename: I
authors.initials: I
authors.lastname: Szatmári
authors.forename: G
authors.initials: G
authors.lastname: Orosz
authors.forename: K
authors.initials: K
authors.lastname: Medzihradszky
authors.forename: A
authors.initials: A
authors.lastname: Borsodi
mesh_terms: D000345:Affinity Labels; D000477:Alkylating Agents; D000818:Animals; D001667:Binding, Competitive; D001921:Brain / Q000378:metabolism; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D004743:Enkephalin, Leucine / Q000031:analogs & derivatives; D007700:Kinetics; D008297:Male; D009842:Oligopeptides; D011869:Radioligand Assay; D051381:Rats; D017208:Rats, Wistar; D017465:Receptors, Opioid, delta / Q000378:metabolism*; D017473:Receptors, Opioid, kappa / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000378:metabolism
chemical_list: D000345:Affinity Labels; D000477:Alkylating Agents; D009842:Oligopeptides; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; C401843:tyrosyl-seryl-glycyl-phenylalanyl--leucyl-threoninamidoamino-glycyl-maleic acid; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; C088242:deltorphin II, Ile(5,6)-; D004743:Enkephalin, Leucine; C034318:enkephalin, Ser(2), Leu(5), Thr(6)-