Tissue injury produces a delicate balance between latent pain sensitization (LS) and compensatory endogenous opioid receptor analgesia that continues for months, even after re-establishment of normal pain thresholds. To evaluate the contribution of mu (MOR), delta (DOR), and/or kappa (KOR) opioid receptors to the silencing of chronic postoperative pain, we performed plantar incision at the hindpaw, waited 21 days for the resolution of hyperalgesia, and then intrathecally injected subtype-selective ligands. We found that the MOR-selective inhibitor CTOP (1-1000ng) dose-dependently reinstated mechanical hyperalgesia. Two DOR-selective inhibitors naltrindole (1-10 µg) and TIPP[Ψ] (1-20µg) reinstated mechanical hyperalgesia, but only at the highest dose that also produced itching, licking, and tail biting. Both the prototypical KOR-selective inhibitors nor-BNI (0.1-10µg) and the newer KOR inhibitor with more canonical pharmocodynamic effects, LY2456302 (0.1-10µg), reinstated mechanical hyperalgesia. Furthermore, LY2456302 (10µg) increased the expression of phosphorylated signalregulated kinase (pERK), a marker of central sensitization, in dorsal horn neurons but not glia. Sex studies revealed that LY2456302 (0.3 µg) reinstated hyperalgesia and pERK expression to a greater degree in female as compared to male mice. Our results suggest that spinal MOR and KOR, but not DOR, maintain LS within a state of remission to reduce the intensity and duration of postoperative pain, and that endogenous KOR but not MOR analgesia is greater in female mice.

Latent sensitization (LS) of pain engages pronociceptive signaling pathways in the dorsal horn that include NMDA receptor (NMDAR)fiadenylyl cyclase-1 (AC1)fiprotein kinase A (PKA), and exchange proteins directly activated by cyclic AMP (Epacs). To determine whether these pathways operate similarly between males and females or are under the inhibitory control of spinal j opioid receptors (KOR), we allowed hyperalgesia to resolve after plantar incision and then blocked KOR with intrathecal administration of LY2456302, which reinstated hyperalgesia and facilitated touch-evoked immunoreactivity of phosphorylated extracellular signal-regulated kinase (pERK) in neurons (NeuN) but not astrocytes (GFAPs) nor microglia (Iba1). LY2456302 reinstated hyperalgesia even when administered 13 months later, indicating that chronic postoperative pain vulnerability persists for over a year in a latent state of remission. In both sexes, intrathecal MK-801 (an NMDAR competitive antagonist) prevented LY2456302-evoked reinstatement of hyperalgesia as did AC1 gene deletion or the AC1 inhibitor NB001. NB001 also prevented stimulus-evoked pERK. In both sexes, the Epac inhibitor ESI-09 prevented reinstatement, whereas the Epac activator 8-CPT reinstated hyperalgesia. By contrast, the PKA inhibitor H89 prevented reinstatement only in male mice, whereas the PKA activator 6-bnz-cAMP itself evoked reinstatement at all doses tested (3-30 nmol, i.t.). In neither sex did incision change gene expression of KOR, GluN1, PKA, or Epac1 in dorsal horn. We conclude that sustained KOR signaling inhibits spinal PKA-dependent mechanisms that drive postoperative LS in a sex-dependent manner. Our findings support the development of AC1, PKA, and Epac inhibitors toward a new pharmacotherapy for chronic postoperative pain.

nervous system. However, opioid receptors also mediate analgesia on the peripheral terminals of primary sensory afferents. Despite more than a decade of research directed at developing peripherally-restricted opioids, no analgesic to date has delivered on the promise of effective pain control without the risk for addiction and diversion. Therefore, we studied the ability of loperamide (Lo), a highly efficacious MOR agonist that is excluded from the CNS, and oxymorphindole (OMI), a DOR agonist that was shown to synergize with morphine spinally, to mediate peripheral analgesia when administered alone, or in combination. We studied this combination in na€ ıve, inflamed and nerve-injured mice. Our initial efficacy studies-using intraplantar, systemic, or transdermal applications-show that when combined in a 1:1 dose ratio OMI-Lo produces a robust analgesic synergy. This synergy is blocked by the peripherally restricted opioid antagonist, naloxone methiodide, reinforcing the peripheral localization of the effect. The synergistic interaction is also completely ablated when G protein-coupled, inwardly rectifying potassium channels (GIRKs) are blocked or knocked out. We concluded that MOR agonists significantly synergize with DOR agonists at peripheral sites of action, providing strong evidence in support of peripherally restricted opioid combination therapy. The systemic and transdermal efficacy of the combination, along with loperamide's extremely low abuse liability, suggests that this combination therapy might be therapeutically useful to control inflammatory pain in the clinic.