Opioid efficacy in a C6 glioma cell line stably expressing the delta opioid receptor.
A C6 glioma cell line stably transfected with the rat delta opioid receptor (C6delta) was used to characterize receptor binding and G protein activation by both peptide and nonpeptide delta opioid ligands. The ligand binding affinities for [3H]naltrindole and [3H]pCl-[D-Pen2,D-Pen5]enkephalin (DPDPE) were similar to those observed in monkey brain membranes. The nonpeptide agonists, BW373U86 and SNC80, as well as peptide agonist [D-Ser2, L-Leu5]enkephalyl-Thr maximally stimulated [35S]GTPgammaS binding by 640, 654 and 576%, respectively, over basal. The peptide agonists, DPDPE and deltorphin II, both stimulated [35S]GTPgammaS binding by 375%. Etorphine, diprenorphine, oxymorphindole and 7-spiroindanyloxymorphone were also partial agonists in this assay, although they were less efficacious than deltorphin II. Stimulation of [35S]GTPgammaS binding by agonists was blocked completely by pertussis toxin pretreatment. Both delta-1 and delta-2 selective antagonists 7-benzylidenenaltrexone and a benzofuran analog of naltrindole displayed high affinity for the cloned receptor (0.04 and 0.08 nM) and antagonized the stimulation of [35S]GTPgammaS binding by BW373U86 and DPDPE with similar potencies. Other evidence suggesting the lack of receptor subtypes includes the finding that stimulation of [35S]GTPgammaS binding by receptor subtype selective ligands DPDPE and deltorphin II was not additive. BW373U86, SNC80 and DPDPE maximally inhibited forskolin-stimulated adenylyl cyclase. These cells highly express a homogeneous population of delta opioid receptor that couple to inhibitory Go/Gi proteins. Ligand affinity for the delta opioid receptor correlates with ligand EC50 values for stimulation of [35S]GTPgammaS binding.
Clark
M J
MJ
Department of Pharmacology,University of Michigan, Ann Arbor, Michigan 48109-0632, USA.
Emmerson
P J
PJ
Mansour
A
A
Akil
H
H
Woods
J H
JH
Portoghese
P S
PS
Remmers
A E
AE
Medzihradsky
F
F
D037361:Adenylate Cyclase Toxin; D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D002772:Cholera Toxin / Q000494:pharmacology; D020881:Enkephalin, D-Penicillamine (2,5)-; D004745:Enkephalins / Q000378:metabolism / Q000494:pharmacology; D005910:Glioma / Q000378:metabolism* / Q000473:pathology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D009294:Narcotics / Q000378:metabolism / Q000494:pharmacology*; D037342:Pertussis Toxin; D051381:Rats; D017465:Receptors, Opioid, delta / Q000145:classification / Q000187:drug effects* / Q000378:metabolism; D014407:Tumor Cells, Cultured; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D000067956:Adenylyl Cyclase Inhibitors; D004745:Enkephalins; D009294:Narcotics; D017465:Receptors, Opioid, delta; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D020881:Enkephalin, D-Penicillamine (2,5)-; D002772:Cholera Toxin; D037342:Pertussis Toxin