Abstract
A new class of highly selective δ-opioid receptor antagonists has been recently developed, termed the TIP(P) peptides. Two prototypical compounds in this class are TIPP (H-Tyr-Tic-Phe-Phe-OH) and a derivative, TIPP-ψ (H-Tyr-Tic[CH2NH]-Phe-Phe-OH). Surprisingly, both TIPP and TIPP-ψ demonstrated inhibition of adenylyl cyclase activity in GH3 cells transfected with δ-opioid receptors (GH3DORT), an effect normally observed by agonists. The agonist activity was δ-selective, because no inhibition occurred in wild-type GH3 or GH3MOR (μ-opioid receptor) cells. Both TIPP and TIPP-ψ exhibited concentration-dependent inhibition of adenylyl cyclase activity; however, TIPP-ψ was found to be less potent (IC50 = 3.97 versus 0.162 nM) and less efficacious (Imax = 50% versus 70%) than TIPP. Pretreatment of cells with pertussis toxin attenuated the inhibition of maximally effective concentrations of TIPP and TIPP-ψ, indicating the involvement of Giα/Goα G-proteins. Other δ-antagonists, naltriben, naloxone, and ICI 174864, attenuated the inhibition of adenylyl cyclase activity mediated by TIPP. Coadministration of TIPP with the selective δ-agonist [d-Pen2,5]enkephalin resulted in an additive interaction. Both TIPP and TIPP-ψ exhibited significant inhibition of adenylyl cyclase activity in different GH3DORT clones expressing a 28-fold range of δ-opioid receptor densities, and in cell lines expressing endogenous (i.e., N1E115 and NG108-15) and transfected (i.e., Chinese hamster ovary-DOR and human embryonic kidney-DOR) δ-opioid receptors, with densities ranging from 0.12 to 6.67 pmol/mg. These results suggest that compounds previously thought to be purely δ-opioid receptor antagonists also demonstrate agonist activity in several in vitro models.
Footnotes
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This work was supported in part by the National Institute on Drug Abuse Grant DA10936 (to P.L.P.), by The American Heart Association-Heartland Affiliate (to N.A.M.), and by the University of Arkansas for Medical Sciences Graduate Student Research Fund (to N.A.M).
- Abbreviations:
- Aib
- α-aminoisobutyric acid
- TIPP
- H-Tyr-Tic-Phe-Phe-OH
- TIPP-ψ
- H-Tyr-Tic[CH2NH]-Phe-Phe-OH
- DMSO
- dimethyl sulfoxide
- DPDPE
- [d-Pen2,5]enkephalin
- DMEM
- Dulbecco's modified Eagle's medium
- MOR
- μ-opioid receptor
- MORDOR
- μ- and δ-opioid receptors
- CHO
- Chinese hamster ovary
- HEK
- human embryonic kidney
- DOR
- δ-opioid receptor
- DORT
- tagged δ-opioid receptor
- PTX
- pertussis toxin
- ANOVA
- analysis of variance
- Received December 29, 2000.
- Accepted April 3, 2001.
- The American Society for Pharmacology and Experimental Therapeutics
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