Pseudoirreversible binding characteristics of [D-Ala2,Glu4]deltorphin and its Cys4 substituted derivative to delta-opioid receptors.
Following the identification of [D-Ala2,Glu4]deltorphin as a selective delta 2-opioid receptor agonist in vivo, we synthesized the Cys4-substituted analogue as a potential ligand which might bind 'irreversibly' at this site through a proposed thiol-disulfide exchange mechanism. Previous studies showed that intracerebroventricular (i.c.v.) pretreatment with [D-Ala2,Cys4]deltorphin, 24 h prior to antinociceptive testing, produced a selective antagonism of [D-Ala2,Glu4]deltorphin-induced antinociception in mice. Surprisingly, however, the Ser4-analogue (synthesized as a control) and even the parent molecule, [D-Ala2,Glu4]deltorphin, had the same antagonistic effect following pretreatment in vivo, while pretreatment with an equiantinociceptive dose of [D-Ser2,Leu5,Thr6]-enkephalin, a structurally unrelated delta 2-opioid receptor agonist did not exhibit long-lasting antinociceptive actions. These data raised questions regarding the mechanism of the antagonism observed in vivo with the deltorphins; the present studies have attempted to explore these issues using radioligand binding techniques. The results demonstrate a decrease in the Bmax of [tyrosyl-3',5'-3H,D-Pen2,p-Cl-Phe4,D-Pen5]-enkephalin ([3H]p-Cl-DPDPE) (delta-opioid receptor ligand) following i.c.v. pretreatment of mice (at -24 h) with [D-Ala2,Cys4]deltorphin or [D-Ala2,Glu4]deltorphin, but not with [D-Ala2,Ser4]deltorphin, suggesting a difference in mechanism of antagonism seen in vivo with these compounds. Incubation of mouse whole brain homogenates in vitro with [D-Ala2,Cys4]deltorphin or with [D-Ala2,Glu4]deltorphin, also resulted in a decrease in the radioligand binding of [3H]p-Cl-DPDPE, but this effect was not prevented by coincubation with dithiothreitol, a thiol-reducing agent.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Pharmacology, University of Arizona, Tucson.
K D
KD
Wild
P J
PJ
Horan
A
A
Misicka
A
A
Lipkowski
R C
RC
Haaseth
T O
TO
Matsunaga
V J
VJ
Hruby
G
G
Toth
A
A
Borsodi
H I
HI
Yamamura
D000700:Analgesics / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001921:Brain / Q000187:drug effects / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D020881:Enkephalin, D-Penicillamine (2,5)-; D004745:Enkephalins / Q000378:metabolism; D066298:In Vitro Techniques; D007276:Injections, Intraventricular; D008297:Male; D008566:Membranes / Q000378:metabolism; D051379:Mice; D009842:Oligopeptides / Q000378:metabolism* / Q000494:pharmacology; D011869:Radioligand Assay; D017465:Receptors, Opioid, delta / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000700:Analgesics; D004745:Enkephalins; D009842:Oligopeptides; D017465:Receptors, Opioid, delta; C082680:deltorphin, Ala(2)-Cys(4)-; C060209:deltorphin II, Ala(2)-; D020881:Enkephalin, D-Penicillamine (2,5)-; D004229:Dithiothreitol