Evidence for chemical differentiation of delta opioid receptor subtypes by the sulfhydryl reagent N-ethylmaleimide.
In this study, the delta receptor-selective nonequilibrium affinity ligands, 5'-NTII and DALCE, and the nonspecific sulfhydryl reagent NEM were evaluated over a range of concentrations and treatment conditions for their ability to selectively alter the binding properties of delta 1- or delta 2-preferring opioid radioligands in brain homogenate. Treatment of tissue preparations with DALCE (0-10,000 nM) or NTII (0-10,000 nM) resulted in an equivalent concentration-dependent loss of binding capacity for the delta 1 agonist 3H-DPDPE and the mu/delta 2 agonist 3H-DSLET. In contrast, treatment of tissue with NEM (0-8000 microM) resulted in greater loss of 3H-DPDPE binding. Scatchard analysis of the binding of 3H-DPDPE, 3H-DSLET, and 3H-NTI in 3 mM NEM-treated rat brain P2 preparation revealed an equivalent decrease in affinity for the agonist ligands, but a significantly greater decrease in Bmax for 3H-DPDPE compared with control tissue values. Comparison of the K(i) values for a series of delta-selective compounds against 3H-DSLET binding in control vs 3 mM NEM treated P2 fraction showed differential effects of NEM on affinity within the series that were consistent with a selective depletion of delta 1 sites. Overall, these results indicate that NEM treatment selectively reduced delta 1 receptor binding, resulting in a preparation that is enriched in delta 2 sites.
Tam
L
L
Neurological Diseases Research, G. D. Searle and Co., Skokie, IL 60077.
Rafferty
M F
MF
D000345:Affinity Labels; D000818:Animals; D001921:Brain / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D016308:Enkephalin, Leucine-2-Alanine / Q000031:analogs & derivatives / Q000494:pharmacology; D004745:Enkephalins / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology*; D017879:Isothiocyanates / Q000494:pharmacology; D008297:Male; D051379:Mice; D009271:Naltrexone / Q000031:analogs & derivatives / Q000494:pharmacology; D011869:Radioligand Assay / Q000379:methods*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017465:Receptors, Opioid, delta / Q000145:classification* / Q000187:drug effects / Q000378:metabolism
D000345:Affinity Labels; D004745:Enkephalins; D017879:Isothiocyanates; D017465:Receptors, Opioid, delta; C053821:enkephalin-Leu, Ala(2)-Cys(6)-; C064283:naltrindole 5'-isothiocyanate; D009271:Naltrexone; D016308:Enkephalin, Leucine-2-Alanine; D005033:Ethylmaleimide