The Antitussive Activity of d-Opioid Receptor Stimulation in Guinea Pigs

In this study, the activity of the d-opioid receptor subtypeselective agonist, SB 227122, was investigated in a guinea pig model of citric acid-induced cough. Parenteral administration of selective agonists of the d-opioid receptor (SB 227122), m-opioid receptor (codeine and hydrocodone), and k-opioid receptor (BRL 52974) produced dose-related inhibition of citric acid-induced cough with ED50 values of 7.3, 5.2, 5.1, and 5.3 mg/kg, respectively. The nonselective opioid receptor antagonist, naloxone (3 mg/kg, i.m.), attenuated the antitussive effects of codeine or SB 227122, indicating that the antitussive activity of both compounds is opioid receptor-mediated. The d-receptor antagonist, SB 244525 (10 mg/kg, i.p.), inhibited the antitussive effect of SB 227122 (20 mg/kg, i.p.). In contrast, combined pretreatment with b-funaltrexamine (m-receptor antagonist; 20 mg/kg, s.c.) and norbinaltorphimine (k-receptor antagonist; 20 mg/kg, s.c.), at doses that inhibited the antitussive activity of mand k-receptor agonists, respectively, was without effect on the antitussive response of SB 227122 (20 mg/kg, i.p.). The s-receptor antagonist rimcazole (3 mg/kg, i.p.) inhibited the antitussive effect of dextromethorphan (30 mg/kg, i.p.), a s-receptor agonist, but not that of SB 227122. These studies provide compelling evidence that the antitussive effects of SB 227122 in this guinea pig cough model are mediated by agonist activity at the d-opioid receptor. Opioid receptor agonists are classified by their activity at three opioid receptor subtypes known as m (Wang et al., 1994), k (Mansson et al., 1994), and d (Knapp et al., 1994). Members of this class of compounds have been demonstrated to have antitussive effects. For example, compounds such as morphine and codeine, considered to be the most potent and effective antitussive drugs currently on the market, are categorized generally as agonists at the m-opioid receptor (Eddy et al., 1969; Karlsson et al., 1990). In addition, k-opioidselective agonists have been shown to inhibit cough in laboratory animals (Kamei et al., 1990). Although the involvement of d-opioid receptors in cough has been described (Kamei et al., 1992; Dondio et al., 1997), some of the data are conflicting. Some reports indicate that d-receptor antagonists produce an antitussive effect against capsaicin-induced cough in rats and mice (Kamei et al., 1993c, 1994b). Furthermore, evidence has been presented that d-opioid receptor agonists can either reduce (Kamei et al., 1991) or enhance (Kamei et al., 1993d) the antitussive effect of m-receptor agonists [e.g., D-Ala-Me-Phe-Gly-olenkephalin (DAMGO), morphine]. The reason(s) for these discrepancies is not known but may be linked to the limited selectivity of the compounds that were investigated. The goal of this study was to clarify the influence of dreceptors in a cough model. Specifically, we investigated the ability of the novel d-opioid receptor agonist, SB 227122 ([10R, 4bS-(4bb,9ab)]-7-diisopropylaminocarbonyl-8,14-dimethyl-4hydroxy-3-methoxy-4b,5,9,9a,10,11-hexahydro-(6H)-[2,3-h]-pyrrolo[10,4-b]iminoethenophenanthrene; Fig. 1), to inhibit citric acid-induced cough in the guinea pig. SB 227122 is a nonpeptide agonist with high affinity for the human d-receptor (Ki 5 6.9 nM from binding studies) and Ki . 2 mM versus mor k-receptors (Petrillo et al., 1998). In addition, we attempted to confirm the mechanism of the antitussive activity of SB 227122 by using subtype-selective opioid receptor antagonists. Materials and Methods Cell Lines. Stable expression of human d-opioid receptors (hDORs) and m-opioid receptors (h-MORs) in Chinese hamster ovary (CHO) cells and human k-opioid receptors (h-KORs) in human embryonic kidney 293 (HEK-293) cells were prepared in-house. The h-DOR was cloned by screening primers based on the public seReceived for publication July 30, 1999. ABBREVIATIONS: DAMGO, D-Ala-Me-Phe-Gly-ol-enkephalin; h-DOR, human d-opioid receptor; h-MOR, human m-opioid receptor; h-KOR, human k-opioid receptor; b-FNA, b-funaltrexamine; Nor-BNI, norbinaltorphimine; CHO, Chinese hamster ovary; HEK-293, human embryonic kidney 293; RT-PCR, reverse-transcription polymerase chain reaction; DADLE, D-Ala-D-Leu-enkephalin; S.A., specific activity; PLSD, protected least-squares difference. 0022-3565/00/2922-0803$03.00/0 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 292, No. 2 Copyright © 2000 by The American Society for Pharmacology and Experimental Therapeutics Printed in U.S.A. JPET 292:803–809, 2000 803 at A PE T Jornals on Sptem er 3, 2017 jpet.asjournals.org D ow nladed from