Characterization of specific opioid binding sites in neural membranes from the myenteric plexus of porcine small intestine.
Delta- and kappa-opioid receptors (OPRs), but not micro-OPRs, are expressed in the myenteric plexus of the porcine distal small intestine. In a subpopulation of myenteric neurons, delta- and kappa-OPRs seem to be colocalized and may functionally interact. In this study, radioligand binding was used to characterize myenteric OPR populations in detail. The nonselective OPR antagonist [3H]diprenorphine bound to a single, high-affinity site in myenteric neural membrane homogenates. Naloxone displaced 65 and 59% of [3H]diprenorphine binding from this site in Na(+)-free Tris and Krebs-HEPES buffers, respectively. Naltrexone-derived delta- and kappa-OPR antagonists, including naltriben, 7-benzylidenenaltrexone, nor-binaltorphimine, and 5'-guanidinonaltrindole, displaced [3H]diprenorphine from two distinct binding sites to levels similar to that of naloxone. The selective delta-OPR ligands Tyr-1,2,3,4-tetrahydroisoquinoline-Phe-Phe-OH (TIPP), [D-Pen2,D-Pen5]enkephalin (DPDPE), [D-Ala2, Glu4]deltorphin II, and (+)-4-[(alphaR)-alpha((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl-3-methoxybenzyl)-N,N-diethylbenzamide (SNC-80) and the kappa-OPR agonist (D-(5alpha,7alpha,8beta)-(-)-N-methyl-N-(7-(1-pyrrolidinyl)-1-oxoaspiro-(4,5)dec-8-yl) benzeneacetamide (U-69,593) displaced [3H]diprenorphine from three independent binding sites; these included high-affinity delta- and kappa-OPR sites, and a residual binding site. Residual [3H]diprenorphine binding was displaced by the selective kappa-OPR antagonist nor-binaltorphimine after saturation of delta and kappa sites, respectively, with DPDPE and U-69,593. The residual binding site displayed low affinity for delta- and kappa-OPR agonists and TIPP, as well as moderate affinity for naltrexone-derived ligands, properties reminiscent of delta-/kappa-OPR heterodimers.
Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Minnesota, St. Paul, Minnesota 55108-6010, USA.
DeWayne
D
Townsend
Philip S
PS
Portoghese
David R
DR
Brown
D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004174:Diprenorphine / Q000494:pharmacology; D004347:Drug Interactions; D020881:Enkephalin, D-Penicillamine (2,5)- / Q000494:pharmacology; D005260:Female; D006531:HEPES / Q000378:metabolism; D007421:Intestine, Small / Q000166:cytology*; D008297:Male; D009197:Myenteric Plexus / Q000187:drug effects* / Q000378:metabolism; D009292:Narcotic Antagonists / Q000494:pharmacology; D009294:Narcotics / Q000494:pharmacology*; D017465:Receptors, Opioid, delta / Q000819:agonists / Q000378:metabolism*; D017473:Receptors, Opioid, kappa / Q000819:agonists / Q000378:metabolism*; D017450:Receptors, Opioid, mu / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D013552:Swine; D014316:Tritium
D009292:Narcotic Antagonists; D009294:Narcotics; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D014316:Tritium; D004174:Diprenorphine; D020881:Enkephalin, D-Penicillamine (2,5)-; D012964:Sodium; D006531:HEPES