Modulation of morphine tolerance by Mas-related G protein-coupled receptor D signalling in the mouse dorsal root...

Opioids such as morphine are essential for the management of moderate to severe pain. However, their prolonged use leads to tolerance. Although the central mechanisms of morphine tolerance have been extensively studied, the peripheral molecular pathways remain poorly understood.

We performed time-course RNA sequencing of the dorsal root ganglia (DRG) from a morphine tolerance mouse model, followed by mechanistic studies using genetic and pharmacological manipulations combined with behavioural, molecular, and biochemical assays.

Acute morphine administration reduced Mas-related G protein-coupled receptor D (Mrgprd) expression in DRG neurones by ∼70% within 6-24 hours, with levels returning to baseline by day 4 of the tolerance model. Mrgprd knockdown delayed the onset of tolerance, whereas AAV-mediated Mrgprd overexpression accelerated its development. Morphine treatment also decreased c-Jun expression by ∼40% at 24 h, consistent with the observed suppression of Mrgprd. In vivo siJun (siRNA targeting Jun mRNA to silence c-Jun expression) suppressed Mrgprd expression in DRG, and ChIP-qPCR and luciferase assays confirmed c-Jun binding to the Mrgprd promoter. Mrgprd knockdown increased Oprm1 mRNA and MOR protein concentrations. Approximately 50% increase in DS-lncRNA after Mrgprd suppression was observed in parallel with decreased Ehmt2/G9a expression, thereby relieving repression of Oprm1/MOR. DS-lncRNA knockdown restored Ehmt2 expression and reduced Oprm1 concentrations, reinstating tolerance in Mrgprd knockout mice.

These findings reveal a peripheral pathway (c-Jun-Mrgprd-DS-lncRNA-Ehmt2/G9a-Oprm1/MOR) that modulates opioid responsiveness. Targeting this axis offers new potential therapeutic strategies to mitigate tolerance and improve the durability of opioid analgesia.