Morphine Tolerance Gated through EZH2‐Mediated Suppression of <i>Trpc5</i> in Spinal GABAergic Interneurons in Male Mice

A major unresolved issue in managing severe pain is tolerance caused by repeated treatment of opioid analgesics. Here, it is demonstrated that tolerance-inducing treatment with morphine results in the persistent downregulation of transient receptor potential canonical 5 (TRPC5), impairing the Ca²⁺ homeostasis in GABAergic interneurons of the spinal dorsal horn (SDH) and consequently reducing GABA release. Spinal activation of TRPC5 by riluzole (RLZ) or lentiviral-mediated TRPC5 overexpression in GABAergic interneurons produces a long-lasting enhancement of morphine's analgesic effect. In contrast, pharmacological inhibition of TRPC5 and mice lacking TRPC5 accelerates the development of morphine tolerance. Mechanistically, it is found that transcriptional suppression of Trpc5 results from enhancer of zeste homolog 2 (EZH2)-mediated epigenetic modifications at the Trpc5 gene promoter. Morphine decreases the enrichment of RNA polymerase II at the Trpc5 promoter. Moreover, exposure to morphine increases EZH2 binding to the Trpc5 promoter, leading to the enrichment of histone H3 lysine-27 trimethylation (H3K27me3). Pharmacological blockade of EZH2 by EPZ6438 or genetic silencing in GABAergic interneurons reverses morphine tolerance. Thus, it is proposed that the clinical translation of these findings may help reduce the suffering of individuals with intractable pain.