title: Opioid binding sites in the guinea pig and rat kidney: radioligand homogenate binding and autoradiography.
abstract: The specific binding of the selective mu-, delta-, and kappa-opioid ligands [3H][D-Ala2,MePhe4,Gly-ol5]enkephalin ([3H] DAGOL), [3H][D-Pen2,D-Pen5]enkephalin ([3H]DPDPE), and [3H]U69593, respectively, to crude membranes of the guinea pig and rat whole kidney, kidney cortex, and kidney medulla was investigated. In addition, the distribution of specific 3H-opioid binding sites in the guinea pig and rat kidney was visualized by autoradiography. Homogenate binding and autoradiography demonstrated the absence of mu- and kappa-opioid binding sites in the guinea pig kidney. No opioid binding sites were demonstrable in the rat kidney. In the guinea pig whole kidney, cortex, and medulla, saturation studies demonstrated that [3H]DPDPE bound with high affinity (KD = 2.6-3.5 nM) to an apparently homogeneous population of binding sites (Bmax = 8.4-30 fmol/mg of protein). Competition studies using several opioid compounds confirmed the nature of the delta-opioid binding site. Autoradiography experiments demonstrated that specific [3H]DPDPE binding sites were distributed radially in regions of the inner and outer medulla and at the corticomedullary junction of the guinea pig kidney. Computer-assisted image analysis of saturation data yielded KD values (4.5-5.0 nM) that were in good agreement with those obtained from the homogenate binding studies. Further investigation of the delta-opioid binding site in medulla homogenates, using agonist ([3H]DPDPE) and antagonist ([3H]diprenorphine) binding in the presence of Na+, Mg2+, and nucleotides, suggested that the delta-opioid site is linked to a second messenger system via a GTP-binding protein. Further studies are required to establish the precise localization of the delta binding site in the guinea pig kidney and to determine the nature of the second messenger linked to the GTP-binding protein in the medulla.
authors.affiliation: Parke-Davis Research Unit, Addenbrookes Hospital Site, Cambridge, UK.
authors.forename: V U
authors.initials: VU
authors.lastname: Dissanayake
authors.forename: J
authors.initials: J
authors.lastname: Hughes
authors.forename: J C
authors.initials: JC
authors.lastname: Hunter
mesh_terms: D000700:Analgesics / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001345:Autoradiography; D044482:Benzeneacetamides; D001667:Binding, Competitive; D002412:Cations; D004174:Diprenorphine / Q000494:pharmacology; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D020881:Enkephalin, D-Penicillamine (2,5)-; D004745:Enkephalins / Q000378:metabolism / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006168:Guinea Pigs; D007668:Kidney / Q000378:metabolism*; D007700:Kinetics; D009292:Narcotic Antagonists; D009711:Nucleotides / Q000494:pharmacology; D011759:Pyrrolidines / Q000378:metabolism; D011869:Radioligand Assay; D051381:Rats; D011957:Receptors, Opioid / Q000032:analysis / Q000378:metabolism*; D017465:Receptors, Opioid, delta; D014316:Tritium
chemical_list: D000700:Analgesics; D044482:Benzeneacetamides; D002412:Cations; D004745:Enkephalins; D009292:Narcotic Antagonists; D009711:Nucleotides; D011759:Pyrrolidines; D011957:Receptors, Opioid; D017465:Receptors, Opioid, delta; D014316:Tritium; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D004174:Diprenorphine; D020881:Enkephalin, D-Penicillamine (2,5)-; D019204:GTP-Binding Proteins; C045444:U 69593