Characterization of Specific Opioid Binding Sites in Neural Membranes from the Myenteric Plexus of Porcine Small Intestine

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

δ- and κ-Opioid receptors (OPRs), but not μ-OPRs, are expressed in the myenteric plexus of the porcine distal small intestine. In a subpopulation of myenteric neurons, δ- and κ-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 δ- and κ-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 δ-OPR ligands Tyr-1,2,3,4-tetrahydroisoquinoline-Phe-Phe-OH (TIPP), [D-Pen 2,D-Pen5]enkephalin (DPDPE), [D-Ala2, Glu 4]deltorphin II, and (+)-4-[(αR)-α ((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl-3-methoxybenzyl)-N, N-diethylbenzamide (SNC-80) and the κ-OPR agonist (D-(5α,7α ,8β)-(-)-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 δ- and κ-OPR sites, and a residual binding site. Residual [ 3H]diprenorphine binding was displaced by the selective κ-OPR antagonist nor-binaltorphimine after saturation of δ and κ sites, respectively, with DPDPE and U-69,593. The residual binding site displayed low affinity for δ- and κ-OPR agonists and TIPP, as well as moderate affinity for naltrexone-derived ligands, properties reminiscent of δ-/κ-OPR heterodimers.

Original languageEnglish (US)
Pages (from-to)385-393
Number of pages9
JournalJournal of Pharmacology and Experimental Therapeutics
Volume308
Issue number1
DOIs
StatePublished - Jan 1 2004

Fingerprint

Myenteric Plexus
Opioid Receptors
Opioid Analgesics
Small Intestine
Diprenorphine
Swine
Binding Sites
Membranes
Narcotic Antagonists
phenylalanylphenylalanine
Naltrexone
Naloxone
Benzeneacetamides
Tetrahydroisoquinolines
Ligands
HEPES
Buffers
Neurons

Cite this

@article{45197a35b05a4605b1ccc346daff694d,
title = "Characterization of Specific Opioid Binding Sites in Neural Membranes from the Myenteric Plexus of Porcine Small Intestine",
abstract = "δ- and κ-Opioid receptors (OPRs), but not μ-OPRs, are expressed in the myenteric plexus of the porcine distal small intestine. In a subpopulation of myenteric neurons, δ- and κ-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 δ- and κ-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 δ-OPR ligands Tyr-1,2,3,4-tetrahydroisoquinoline-Phe-Phe-OH (TIPP), [D-Pen 2,D-Pen5]enkephalin (DPDPE), [D-Ala2, Glu 4]deltorphin II, and (+)-4-[(αR)-α ((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl-3-methoxybenzyl)-N, N-diethylbenzamide (SNC-80) and the κ-OPR agonist (D-(5α,7α ,8β)-(-)-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 δ- and κ-OPR sites, and a residual binding site. Residual [ 3H]diprenorphine binding was displaced by the selective κ-OPR antagonist nor-binaltorphimine after saturation of δ and κ sites, respectively, with DPDPE and U-69,593. The residual binding site displayed low affinity for δ- and κ-OPR agonists and TIPP, as well as moderate affinity for naltrexone-derived ligands, properties reminiscent of δ-/κ-OPR heterodimers.",
author = "DeWayne Townsend and Portoghese, {Philip S} and Brown, {David R}",
year = "2004",
month = "1",
day = "1",
doi = "10.1124/jpet.103.058016",
language = "English (US)",
volume = "308",
pages = "385--393",
journal = "Journal of Pharmacology and Experimental Therapeutics",
issn = "0022-3565",
publisher = "American Society for Pharmacology and Experimental Therapeutics",
number = "1",

}

TY - JOUR

T1 - Characterization of Specific Opioid Binding Sites in Neural Membranes from the Myenteric Plexus of Porcine Small Intestine

AU - Townsend, DeWayne

AU - Portoghese, Philip S

AU - Brown, David R

PY - 2004/1/1

Y1 - 2004/1/1

N2 - δ- and κ-Opioid receptors (OPRs), but not μ-OPRs, are expressed in the myenteric plexus of the porcine distal small intestine. In a subpopulation of myenteric neurons, δ- and κ-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 δ- and κ-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 δ-OPR ligands Tyr-1,2,3,4-tetrahydroisoquinoline-Phe-Phe-OH (TIPP), [D-Pen 2,D-Pen5]enkephalin (DPDPE), [D-Ala2, Glu 4]deltorphin II, and (+)-4-[(αR)-α ((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl-3-methoxybenzyl)-N, N-diethylbenzamide (SNC-80) and the κ-OPR agonist (D-(5α,7α ,8β)-(-)-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 δ- and κ-OPR sites, and a residual binding site. Residual [ 3H]diprenorphine binding was displaced by the selective κ-OPR antagonist nor-binaltorphimine after saturation of δ and κ sites, respectively, with DPDPE and U-69,593. The residual binding site displayed low affinity for δ- and κ-OPR agonists and TIPP, as well as moderate affinity for naltrexone-derived ligands, properties reminiscent of δ-/κ-OPR heterodimers.

AB - δ- and κ-Opioid receptors (OPRs), but not μ-OPRs, are expressed in the myenteric plexus of the porcine distal small intestine. In a subpopulation of myenteric neurons, δ- and κ-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 δ- and κ-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 δ-OPR ligands Tyr-1,2,3,4-tetrahydroisoquinoline-Phe-Phe-OH (TIPP), [D-Pen 2,D-Pen5]enkephalin (DPDPE), [D-Ala2, Glu 4]deltorphin II, and (+)-4-[(αR)-α ((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl-3-methoxybenzyl)-N, N-diethylbenzamide (SNC-80) and the κ-OPR agonist (D-(5α,7α ,8β)-(-)-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 δ- and κ-OPR sites, and a residual binding site. Residual [ 3H]diprenorphine binding was displaced by the selective κ-OPR antagonist nor-binaltorphimine after saturation of δ and κ sites, respectively, with DPDPE and U-69,593. The residual binding site displayed low affinity for δ- and κ-OPR agonists and TIPP, as well as moderate affinity for naltrexone-derived ligands, properties reminiscent of δ-/κ-OPR heterodimers.

UR - http://www.scopus.com/inward/record.url?scp=0345866766&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0345866766&partnerID=8YFLogxK

U2 - 10.1124/jpet.103.058016

DO - 10.1124/jpet.103.058016

M3 - Article

VL - 308

SP - 385

EP - 393

JO - Journal of Pharmacology and Experimental Therapeutics

JF - Journal of Pharmacology and Experimental Therapeutics

SN - 0022-3565

IS - 1

ER -