Role of G protein-coupled receptor kinases 2 and 3 in μ-opioid receptor desensitization and internalization

Lowe, Janet D.; Sanderson, Helen S.; Cooke, Alexandra E.; Ostovar, Mehrnoosh; Tsisanova, Elena; Withey, Sarah L.; Chavkin, Charles; Husbands, Stephen M.; Kelly, Eamonn; Henderson, Graeme; Bailey, Chris P.

Abstract

There is ongoing debate about the role of G protein-coupled receptor kinases (GRKs) in agonist-induced desensitization of the μ-opioid receptor (MOPr) in brain neurons. In the present paper, we have used a novel membrane-permeable, small-molecule inhibitor of GRK2 and GRK3, Takeda compound 101 (Cmpd101; 3-[[[4-methyl-5-(4-pyridyl)-4H-1,2,4-triazole-3-yl] methyl] amino]-N-[2-(trifuoromethyl) benzyl] benzamidehydrochloride), to study the involvement of GRK2/3 in acute agonist-induced MOPr desensitization. We observed that Cmpd101 inhibits the desensitization of the G protein-activated inwardly-rectifying potassium current evoked by receptor-saturating concentrations of methionine-enkephalin (Met-Enk), [D-Ala2, N-MePhe4, Gly-ol5]-enkephalin (DAMGO), endomorphin-2, and morphine in rat and mouse locus coeruleus (LC) neurons. In LC neurons from GRK3 knockout mice, Met-Enk-induced desensitization was unaffected, implying a role for GRK2 in MOPr desensitization. Quantitative analysis of the loss of functional MOPrs following acute agonist exposure revealed that Cmpd101 only partially reversed MOPr desensitization. Inhibition of extracellular signal-regulated kinase 1/2, protein kinase C, c-Jun N-terminal kinase, or GRK5 did not inhibit the Cmpd101-insensitive component of desensitization. In HEK 293 cells, Cmpd101 produced almost complete inhibition of DAMGO-induced MOPr phosphorylation at Ser375, arrestin translocation, and MOPr internalization. Our data demonstrate a role for GRK2 (and potentially also GRK3) in agonist-induced MOPr desensitization in the LC, but leave open the possibility that another, as yet unidentified, mechanism of desensitization also exists.

• Journal Article Type: Article
• Acceptance Date: Feb 2, 2015
• Online Publication Date: May 26, 2015
• Publication Date: Aug 1, 2015
• Deposit Date: Jun 22, 2021
• Journal: Molecular Pharmacology
• Print ISSN: 0026-895X
• Electronic ISSN: 1521-0111
• Publisher: American Society for Pharmacology and Experimental Therapeutics
• Peer Reviewed: Peer Reviewed
• Volume: 88
• Issue: 2
• Pages: 347-356
• DOI: https://doi.org/10.1124/mol.115.098293
• Public URL: https://uwe-repository.worktribe.com/output/7483584