Class A G protein-coupled receptors (GPCRs) normally function as monomers, although evidence from heterologous expression systems suggests they may sometimes form homodimers and/or heterodimers. This study aims to evaluate possible functional interplay of endogenous µ- and d-Opioid receptors (MORs and DORs) in mouse neurons. Detecting GPCR dimers in native tissues however has been challenging. Previously, MORs and DORs co-expressed in transfected cells have been reported to form heterodimers, and their possible co-localization in neurons has been studied in knock-in mice expressing genetically engineered receptors fused to fluorescent proteins. Here we find that single cholinergic neurons in the mouse striatum endogenously express both MORs and DORs. The receptors on neurons from live brain slices were fluorescently labeled in live brain slices with a ligand-directed labeling reagent, NAI-A594. The selective activation of MORs and DORs, with DAMGO (µ-agonist) and deltorphin (d-agonist) inhibited spontaneous firing in all cells examined. In the continued presence of agonist, the firing rate returned to baseline as the result of receptor desensitization with the application of deltorphin but was less observed with the application of DAMGO. In addition, agonist-induced internalization of DORs but not MORs was detected. When MORs and DORs were activated simultaneously with [Met5]-enkephalin, desensitization of MORs was facilitated but internalization was not increased. Together, these results indicate that while MORs and DORs are expressed in single striatal cholinergic interneurons, the two receptors function independently.
All data generated or analyzed during this study are included in the manuscript.
- Seksiri Arttamangkul
- David Farrens
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: All animal uses were conducted in accordance with the National Institutes of Health guidelines and with approval from the Institutional Animal Care and Use Committee (IACUC) protocol #IP00000160 of the Oregon Health & Science University. Rats and mice were anesthetized with isofluorane before euthanized with minimal suffering.
- Julie A Kauer, Stanford University, United States
© 2021, Arttamangkul et al.
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