Inhibition of endothelin A receptor by a novel, selective receptor antagonist enhances morphine-induced analgesia: Possible functional interaction of dimerized endothelin A and μ-opioid receptors

Abstract

Background: The misuse of opioids has led to an epidemic in recent times. The endothelin A receptor (ETAR) has recently attracted attention as a novel therapeutic target to enhance opioid analgesia. We hypothesized that endothelin A receptors may affect pain mechanisms by heterodimerization with μ opioid receptors. We examined the mechanisms of ETAR-mediated pain and the potential therapeutic effects of an ETAR antagonist, Compound-E, as an agent for analgesia.

Methods: Real-time in vitro effect of Compound-E on morphine response was assessed in HEK293 cells expressing both endothelin A and μ opioid receptors through CellKey™ and cADDis cAMP assays. Endothelin A/μ opioid receptor dimerization was assessed by immunoprecipitation and live cell imaging. The in vivo effect of Compound-E was evaluated using a morphine analgesia mouse model that observed escape response behavior, body temperature, and locomotor activity.

Results: In CellKey™ and cAMP assays, pretreatment of cells with endothelin-1 attenuated morphine-induced responses. These responses were improved by Compound-E, but not by BQ-123 nor by bosentan, an ETAR and endothelin B receptor antagonist. Dimerization of ETARs and μ opioid receptors was confirmed by Western blot and total internal reflection fluorescence microscopy in live cells. In vivo, Compound-E potentiated and prolonged the analgesic effects of morphine, enhanced hypothermia, and increased locomotor activity compared to morphine alone.

Conclusion: The results suggest that attenuation by endothelin-1 of morphine analgesia may be caused by dimerization of Endothelin A/μ opioid receptors. The novel ETAR antagonist Compound-E could be an effective adjunct to reduce opioid use.

Keywords: Analgesic; BQ-123 sodium salt (PubChem CID: 52943236); BQ-788 sodium salt (PubChem CID: 23693553); Endothelin A receptor antagonist; G protein-coupled receptor; Morphine; Pain; Receptor heterodimerization; and Bosentan Monohydrate (PubChem CID: 185462); included endothelin-1 (PubChem CID: 16212950); morphine hydrochloride (PubChem CID: 5464110).