The δ Opioid Receptor Agonist SNC80 Selectively Activates Heteromeric μ–δ Opioid Receptors

Abstract

Coexpressed and colocalized μ-and δ-opioid receptors have been established to exist as heteromers in cultured cells and in vivo. However the biological significance of opioid receptor heteromer activation is less clear. To explore this significance, the efficacy of selective activation of opioid receptors by SNC80 was assessed in vitro in cells singly and coexpressing opioid receptors using a chimeric G-proteinmediated calcium fluorescence assay, SNC80 produced a substantially more robust response in cells expressing μ−δ heteromers than in all other cell lines. Intrathecal SNC80 administration in μ-and δ-opioid receptor knockout mice produced diminished antinociceptive activity compared with wild type. The combined in vivo and in vitro results suggest that SNC80 selectively activates μ−δ heteromers to produce maximal antinociception. These data contrast with the current view that SNC80 selectively activates δ-opioid receptor homomers to produce antinociception. Thus, the data suggest that heteromeric μ−δ receptors should be considered as a target when SNC80 is employed as a pharmacological tool in vivo. KEYWORDS: μ opioid receptor, δ opioid receptor, μ−δ heteromer, SNC80, knockout, antinociception C ompound SNC80 (Figure 1) has been employed as the prototypic nonpeptide, selective δ-opioid receptor agonist for nearly 20 years. The experimental techniques employed to define the δ-selectivity of SNC80 included radioligand binding, smooth muscle assays, and in vivo antagonism studies employing selective δ-antagonists. 1−4 SNC80 has found broad recognition and adoption as a selective δ-agonist standard, occupying the previously vacant nonpeptide selective δ-agonist niche in the opioid pharmacological toolbox. 3 However, in view of the discovery of opioid receptor heteromers as targets for ligands over the past decade, it is possible that opioid receptor heteromers may be activated by SNC80. In this regard, three heteromers, μ−δ, 5 μ−κ, 6 and κ−δ, 7 have been established in heterologous cell lines, 8 and selective ligands that target these heteromers have been identified for each of the above heteromers. 9−11 Moreover, heteromeric μ−δ receptors have been detected using selective antibodies in both cultured cells and DRG neurons. 12 Thus, δ-receptors organized as heteromers could be relevant in the action of SNC80.In this regard, SNC80-treated μ-opioid receptor knockout (μ-KO) mice have been found to be without effect in a visceral antiwrithing test. 13 Also, SNC80 produced an antihyperalgesic effect in WT mice but lacked this effect in μ-KO mice, 14 an effect similar to a separate study in δ-KO mice. 15 Additionally, in vitro studies on the effects of SNC80 on opioid receptors suggested the involvement of μ-opioid receptors on the activity of SNC80. These included the finding that colocalized μ-and δ-opioid receptors in large and small dorsal root ganglion (DRG) neurons 16 are cointernalized into the same subcellular compartment for lysosomal degradation upon treatment with

Heteromers of μ‐δ opioid receptors: new pharmacology and novel therapeutic possibilities

“…Studies using cells co‐expressing a chimeric G protein with either opioid receptor heteromers or individual receptor homomers found that SNC80 elicited a robust release in intracellular calcium only in cells expressing MOP‐DOP receptor heteromers (Metcalf et al ., ). In addition, behavioural studies showed that the antinociceptive effect of SNC80 was abolished in animals lacking MOP receptors (Sora et al ., ; Metcalf et al ., ). Furthermore, the antinociceptive efficacy of SNC80 was right‐shifted by approximately threefold and sixfold in MOP or DOP receptor knockout mice, respectively (Metcalf et al ., ).…”

“…In addition, behavioural studies showed that the antinociceptive effect of SNC80 was abolished in animals lacking MOP receptors (Sora et al ., ; Metcalf et al ., ). Furthermore, the antinociceptive efficacy of SNC80 was right‐shifted by approximately threefold and sixfold in MOP or DOP receptor knockout mice, respectively (Metcalf et al ., ). Taken together, these results indicate that the presence of both MOP and DOP receptors are necessary for the antinociceptive activity of SNC80 (Table ).…”

“…The involvement of MOP‐DOP receptor heteromers in the antinociceptive effects of SNC80, a DOP receptor‐selective agonist, has also been investigated. Studies using cells co‐expressing a chimeric G protein with either opioid receptor heteromers or individual receptor homomers found that SNC80 elicited a robust release in intracellular calcium only in cells expressing MOP‐DOP receptor heteromers (Metcalf et al ., ). In addition, behavioural studies showed that the antinociceptive effect of SNC80 was abolished in animals lacking MOP receptors (Sora et al ., ; Metcalf et al ., ).…”

Novel GPCR paradigms at the μ‐opioid receptor

“…This is in contrast with the several descriptions of ‘allosteric interactions’ across opioid receptor dimers, whether homodimers or heterodimers (Jordan and Devi, ; Wang, ; Milligan, ; Yekkirala et al ., ). With regards to the MOR subtype, the crystal structure of the MOR already suggests an oligomeric arrangement of this receptor (Manglik et al ., ), and allosteric interactions have been described between MOR and mGluR5, CB 1 , DOR and κ–opioid receptors (Yekkirala et al ., ; Metcalf et al ., ; Akgün et al ., ; Le Naour et al ., ).…”

Novel diazabicycloalkane delta opioid agonists

“…53 According to previously reported procedures, 54 in this work we investigated SNC80 d agonist profile by analysing up-regulation of P-ERK 1/2 expression in neuroblastoma  glioma NG108-15 hybrid cells (Fig. 5a).…”