Human delta opioid receptor: functional studies on stably transfected Chinese hamster ovary cells after acute and chronic treatment with the selective nonpeptidic agonist SNC-80.
The SNC-80 series of nonpeptidic agonists for the delta-opioid receptor are being developed as potential analgesic drugs. It is important to understand their acute and chronic effects at human delta-opioid receptors. Thus, we measured the ability of SNC-80 and [D-Pen2,4'-Cl-Phe4,D-Pen5]enkephalin to inhibit forskolin-stimulated adenylyl cyclase activity in recombinant Chinese hamster ovary cells stably expressing the cloned human delta-opioid receptor. The calculated EC50 values for [D-Pen2,4'-Cl-Phe4,D-Pen5]enkephalin and SNC-80 were 0.6 +/- 0.1 nM and 6.3 +/- 0.1 nM, respectively. Pretreatment of these cells with SNC-80 (100 nM) for 24 hr produced 1) a time-dependent reduction of delta receptor density, as measured by radioligand binding studies with [3H]naltrindole; 2) a shift in the EC50 value of SNC-80 from 7.7 +/- 4.2 nM to 44.1 +/- 12 nM, as measured by the cyclic AMP assay; 3) a reduction in the maximum inhibition of adenylyl cyclase activity from 86% to 48%; 4) a marked increase in the forskolin stimulation of basal cyclic AMP accumulation by nearly 100% (from 442 pmol/mg of protein to 824 pmol/mg of protein); and 5) a 5-fold increase in forskolin-stimulated cyclic AMP accumulation after addition of naltrindole. These studies showed that SNC-80 produced desensitization and down-regulation of human delta-opioid receptors in recombinant Chinese hamster ovary cells after chronic treatment and that this effect was associated with an increase in adenylyl cyclase activity.
Malatynska
E
E
Department of Pharmacology, University of Arizona, Health Sciences Center, Tucson, USA.
Wang
Y
Y
Knapp
R J
RJ
Waite
S
S
Calderon
S
S
Rice
K
K
Hruby
V J
VJ
Yamamura
H I
HI
Roeske
W R
WR
D000818:Animals; D001549:Benzamides / Q000378:metabolism*; D001667:Binding, Competitive; D016466:CHO Cells; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D015536:Down-Regulation; D006801:Humans; D009271:Naltrexone / Q000031:analogs & derivatives / Q000378:metabolism; D010879:Piperazines / Q000378:metabolism*; D011869:Radioligand Assay; D017465:Receptors, Opioid, delta / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000502:physiology*; D011994:Recombinant Proteins; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014162:Transfection
D001549:Benzamides; D010879:Piperazines; D017465:Receptors, Opioid, delta; D011994:Recombinant Proteins; C088464:4-(alpha-(4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl)-N,N-diethylbenzamide; D009271:Naltrexone; D000242:Cyclic AMP; C055382:naltrindole