µ-Opioid receptors negatively regulate striatal dopamine D1 receptor signaling through heteromerization

Abstract Integrating biophysical, computational, in situ, and in vivo approaches with selective heteromer-disrupting transmembrane peptides, this study demonstrates that µ-opioid receptors (MORs) and dopamine D1 receptors (D1Rs) assemble into functional heteromers. Furthermore, real-time in vivo fiber photometry using a genetically encoded cAMP biosensor directly demonstrates that MORs regulate striatal D1Rs via heteromerization in the intact mouse brain. Our findings support a tetrameric structure composed of Gi-coupled MOR and Gs-coupled D1R homodimers, which enables both canonical Gi-Gs antagonistic interaction at the level of adenylyl cyclase and receptor-level cross-antagonism, by which MOR agonists and antagonists counteract D1R agonist-induced signaling. Located predominantly within striosomal GABAergic projection neurons, MOR–D1R heteromers constitute mechanistically defined signaling units that integrate dopaminergic and opioid neurotransmission governing motor control and reward processing. Collectively, these findings establish MOR–D1R heteromers as a previously unrecognized therapeutic target for the treatment of opioid withdrawal-associated restlessness and restless legs syndrome.