Endogenous Opioid-Masked Latent Pain Sensitization: Studies from Mouse to Human

Abstract

Following the resolution of a severe inflammatory injury in rodents, administration of mu-opioid receptor inverse agonists leads to reinstatement of pain hypersensitivity. The mechanisms underlying this form of latent pain sensitization (LS) likely contribute to the development of chronic pain, but LS has not yet been demonstrated in humans. Using a C57BL/6 mouse model of cutaneous mild heat injury (MHI) we demonstrated a dose-dependent reinstatement of pain sensitization, assessed as primary (P < 0.001) and secondary hyperalgesia (P < 0.001) by naloxone (0.3–10 mg/kg), 168 hrs after the induction of MHI. Forward-translating the dose data to a human MHI model (n = 12) we could show that LS does indeed occur after naloxone 2 mg/kg, 168 hrs after a MHI. Our previous unsuccessful efforts to demonstrate unmasking of LS in humans are thus likely explained by an insufficient naloxone dose (0.021 mg/kg). However, while LS was consistently demonstrated in 21/24 mice, LS was only seen in 4/12 subjects. This difference is likely due to selection bias since the C57BL/6 mouse strain exhibits markedly enhanced pain sensitivity in assays of acute thermal nociception. Future exploratory studies in humans should prioritize inclusion of “high-sensitizers” prone to develop LS and use post-surgical models to elucidate markers of vulnerability to chronic postsurgical pain.

Trial registration: EudraCT 2012-005663-27.

Fig 1. CONSORT flow-algorithm for human subjects.

Four subjects were allocated to a feasibility study. One subject was excluded from this part due to an unsuspected finding of an abnormal ultrasound cardiography. Fifteen subjects were included and randomized. Three subjects were excluded for reasons not related to the study. Twelve subjects completed the study and data were examined in an interim analysis by an independent statistician.

Fig 2. Study timeline in the human MHI-model.

Day 1/Day 3 include induction of mild heat injury (MHI) with baseline assessments (green rectangle), heat injury (red) and post-injury assessments (blue). Day 2/Day 4 include a pre-drug assessment (blue; post-injury 165 hrs), drug-infusions (naloxone or placebo; grey) and post-infusion assessments (magenta). Stars indicate assessments of mechanical pain thresholds (red star), thermal thresholds (yellow) and secondary hyperalgesia areas (green).

Fig 3. Hyperalgesia was assessed by a decrease in paw withdrawal threshold (PWT_50%) in a model of mild heat injury (MHI).

Line graphs illustrate response to von Frey (vF) stimulation at baseline (BL) and at 1, 2, 3, 72, and 168 hrs after MHI. Naloxone reinstated primary hyperalgesia (A-B) at a dose of 0.3 mg/kg (60 min, P = 0.0015), 3 mg/kg (30 min, P = 0.0011; 60 min, P = 0.0002), and 10 mg/kg (30 min, P < 0.0001; 60 min, P < 0.0001; 120 min, P < 0.0001) and secondary hyperalgesia (C-D) at a dose of 0.3 mg/kg (60 min, P = 0.0148), 3 mg/kg (30 min, P = 0.0004; 60 min, P = 0.0002), and 10 mg/kg (30 min, P < 0.0001; 60 min, P < 0.0001; 120 min, P = 0.0013). A,C: Line drawings. B,D: Histograms of the post-injection time points (plotted as the mean of the 30–120 min time points). Values represent mean ± 95% CI. * P < 0.05.

Fig 4. Heat pain thresholds and secondary hyperalgesia areas before and after a mild heat injury.

Heat pain thresholds (white circles; left y-axis) and secondary hyperalgesia areas (red circles; right y-axis) before and, 60, 120 and 180 min, after a mild heat injury (MHI). Values represent mean of Day 1 and Day 3 values (mean ± 95% CI). * P < 0.01; ** P < 0.005; **** P < 0.0005.

Fig 5. Individual trajectories of secondary hyperalgesia areas during the early and late phase after a mild heat injury.

Individual trajectories of secondary hyperalgesia areas (n = 12) during the early phase (0–3 hrs) and the late phase (165–169 hrs) after induction of the mild heat injury (MHI). Data are from naloxone (A) and placebo (B) sessions. The trajectories of the four naloxone responders, indicating presence of endogenous opioid masked sensitization, are delineated by red circles and the trajectories from the eight non-responders by blue filled circles. MHI and drug administration are indicated by vertical arrows. One (#11) of the responders demonstrated residual SHA at 165 hrs after the induction of MHI (indicated by oblique arrow; B), but not during placebo. Secondary hyperalgesia areas developed in 4/12 subjects (#2,6,7,11) after naloxone (A) and in 0/12 after placebo (B).