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. 2020 Apr 28:13:54.
doi: 10.3389/fnmol.2020.00054. eCollection 2020.

Positive Allosteric Modulation of CB1 Cannabinoid Receptor Signaling Enhances Morphine Antinociception and Attenuates Morphine Tolerance Without Enhancing Morphine- Induced Dependence or Reward

Richard A Slivicki  1   2 , Vishakh Iyer  1   2 , Sonali S Mali  2 , Sumanta Garai  3 , Ganesh A Thakur  3 , Jonathon D Crystal  1   2 , Andrea G Hohmann  1   2   4
Affiliations

Positive Allosteric Modulation of CB1 Cannabinoid Receptor Signaling Enhances Morphine Antinociception and Attenuates Morphine Tolerance Without Enhancing Morphine- Induced Dependence or Reward

Richard A Slivicki et al. Front Mol Neurosci. .

Abstract

Opioid analgesics represent a critical treatment for chronic pain in the analgesic ladder of the World Health Organization. However, their use can result in a number of unwanted side-effects including incomplete efficacy, constipation, physical dependence, and overdose liability. Cannabinoids enhance the pain-relieving effects of opioids in preclinical studies and dampen unwanted side-effects resulting from excessive opioid intake. We recently reported that a CB1 positive allosteric modulator (PAM) exhibits antinociceptive efficacy in models of pathological pain and lacks the adverse side effects of direct CB1 receptor activation. In the present study, we evaluated whether a CB1 PAM would enhance morphine's therapeutic efficacy in an animal model of chemotherapy-induced neuropathic pain and characterized its impact on unwanted side-effects associated with chronic opioid administration. In paclitaxel-treated mice, both the CB1 PAM GAT211 and the opioid analgesic morphine reduced paclitaxel-induced behavioral hypersensitivities to mechanical and cold stimulation in a dose-dependent manner. Isobolographic analysis revealed that combinations of GAT211 and morphine resulted in anti-allodynic synergism. In paclitaxel-treated mice, a sub-threshold dose of GAT211 prevented the development of tolerance to the anti-allodynic effects of morphine over 20 days of once daily dosing. However, GAT211 did not reliably alter somatic withdrawal signs (i.e., jumps, paw tremors) in morphine-dependent neuropathic mice challenged with naloxone. In otherwise naïve mice, GAT211 also prolonged antinociceptive efficacy of morphine in the tail-flick test and reduced the overall right-ward shift in the ED50 for morphine to produce antinociception in the tail-flick test, consistent with attenuation of morphine tolerance. Pretreatment with GAT211 did not alter somatic signs of μ opioid receptor dependence in mice rendered dependent upon morphine via subcutaneous implantation of a morphine pellet. Moreover, GAT211 did not reliably alter μ-opioid receptor-mediated reward as measured by conditioned place preference to morphine. Our results suggest that a CB1 PAM may be beneficial in enhancing and prolonging the therapeutic properties of opioids while potentially sparing unwanted side-effects (e.g., tolerance) that occur with repeated opioid treatment.

Keywords: allosteric modulator; endocannabinoid; isobologram; neuropathic pain; opioid; withdrawal.

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Figures

FIGURE 1
FIGURE 1
The CB1 PAM GAT211 and the opioid analgesic morphine reduce paclitaxel-induced allodynia in a dose-dependent manner. GAT211 (0.1, 1, 2.5, 5, 10, 20, 30 mg/kg i.p.; previously published in Slivicki et al., 2018b) and morphine (1, 3, 5, 10, 20, 30 mg/kg i.p.; previously published in Slivicki et al., 2018a) both dose-dependently reduced paclitaxel-induced behavioral hypersensitivities to (A) mechanical and (B) cold stimulation. Values were converted to % maximal possible effect (MPE) for (C) mechanical and (D) cold modalities. Data are expressed as mean ± SEM (n = 5–6 per group) #P < 0.05 vs. pre-paclitaxel values, two-way ANOVA followed by Bonferroni post hoc test.
FIGURE 2
FIGURE 2
GAT211 synergizes with the opioid analgesic morphine in suppressing paclitaxel-induced allodynia. Co-administration of GAT211 produces a leftward shift in the dose-response curves of morphine to reduce paclitaxel-induced mechanical (A,C) and cold (B,D) allodynia. Isobolographic analysis revealed a synergistic interaction of GAT211 with morphine in suppressing paclitaxel-induced hypersensitivities to both mechanical (E) and cold (F) stimulation when administered in a 1:1 ratio. Data are expressed as mean ± SEM (n = 6 per group) *P < 0.05 two-tailed t-test vs. theoretical additive values. #P < 0.05 vs. pre-paclitaxel values, two-way ANOVA followed by Bonferroni post hoc test.
FIGURE 3
FIGURE 3
A sub-threshold dose of GAT211 prevents tolerance to the anti-allodynic effects of morphine without exacerbating morphine dependence. (A) Schematic shows timing of experimental treatments. The gray vertical arrows show timing of injections (i.p.) of paclitaxel or cremophor-based vehicle. The black vertical arrows show the timing of behavioral testing for assessing responsiveness to mechanical (Von Frey) and cold (acetone) stimulation. The black horizontal arrow shows the duration of once daily chronic dosing. On day 21 naloxone was injected (i.p.) to precipitate opioid withdrawal. GAT211 (5 mg/kg i.p. × 20 days), administered at a sub-threshold dose for reducing paclitaxel-induced allodynia, enhanced efficacy of morphine in reducing hypersensitivities to mechanical (B) and cold (C) stimulation without the develop of tolerance over a 20-day dosing period. By contrast, tolerance developed to the anti-allodynic efficacy of morphine following repeated dosing. Challenge with naloxone (2 mg/kg i.p.) elicited jumping (D) and paw tremor bouts (E) in mice treated with morphine (10 mg/kg i.p.) and morphine (10 mg/kg i.p.) co-administered with GAT211 (5 mg/kg i.p.). Co-administration of GAT211 with morphine did not reliably enhance or reduce these behaviors relative to morphine alone. (B,C) #P < 0.05 vs. pre-paclitaxel thresholds, *P < 0.05 GAT211 + Morphine vs. all other groups, +P < 0.05 vs. vehicle and GAT211 + morphine (Two-way ANOVA followed by Bonferroni post hoc test. Data are expressed as mean ± SEM (n = 5–6 per group) (D,E) *P < 0.05 overall effect of treatment one-way ANOVA. +P < 0.05 one-way ANOVA followed by Bonferroni post hoc. Data are expressed mean ± SEM (n = 5–6 per group).
FIGURE 4
FIGURE 4
Co-treatment of GAT211 with morphine reduces tolerance to morphine antinociception in the tail-flick test. Schematic shows timing of experimental procedures (A); vertical arrows show time of assessment of tail flick latencies, which were measured 30 min following drug administration (i.p.) on days 2, 4, and 6 of repeated dosing (A). Ascending doses of morphine (0, 1, 3, 10, 30, 100 mg/kg i.p.) produced dose-dependent increases in tail-flick antinociception. Repeated injections of vehicle or GAT211 (20 mg/kg i.p. × 7 days) did not reliably shift the morphine dose response curve (B,C). Repeated injection of morphine (10 mg/kg i.p. × 7 days) produced a right-ward shift in the dose-response curve of morphine in producing antinociception in the tail-immersion test (D,E). The combination of GAT211 (20 mg/kg i.p.) + morphine (10 mg/kg i.p.) also produced a right-ward shift in the dose-response curve for morphine to produce tail-flick antinociception albeit to a lesser degree (D,E). Tolerance to morphine-induced antinociception in the tail immersion test was delayed by co-treatment with GAT211 (20 mg/kg i.p.) (F,G). GAT211+ morphine cotreatment produced heightened antinociception on day 4 but not on day 6 of repeated injections compared to all other groups (F,G). Vehicle and GAT211 do not elicit tail-flick antinociception when administered alone (F,G). Data are expressed as tail-flick latencies in seconds (B,D,F) and values transformed to % MPE (C,E,G) values. XP < 0.05 morphine vs. Vehicle and GAT211, +P < 0.05 GAT211 + morphine vs. Vehicle and GAT211 *P < 0.05 GAT211 + morphine vs. all other groups, two-way ANOVA followed by Bonferroni post-hoc. Mean ± SEM (n = 6 per group).
FIGURE 5
FIGURE 5
GAT211 does not alter somatic expression of morphine dependence in morphine-pelleted mice. Timeline of the behavioral protocol (A). In mice implanted subcutaneously with a 75 mg morphine pellet, naloxone (1 mg/kg i.p.) challenge did not alter the number of jumps (B) or bouts of paw tremors (C) following pretreatment with either GAT211 (20 mg/kg i.p.) or vehicle. Naloxone was injected 72 h following morphine pellet implantation. Data are expressed mean ± SEM (n = 6 per group).
FIGURE 6
FIGURE 6
GAT211 does not alter conditioned place preference to morphine. Morphine (8 mg/kg i.p. × 4 pairings) increases the time spent in the drug-paired chamber relative to the vehicle-paired chamber on the test day (A). The combination of GAT211 (20 mg/kg i.p.) and morphine (8 mg/kg i.p.) produces CPP relative to the vehicle-paired chamber on the test day (B). No difference in chamber preference times were observed pre-conditioning (baseline) in any study (A–C). Vehicle-vehicle pairings do not result in preference for any chamber (C). Chamber preference scores did not differ reliably in mice receiving morphine alone or morphine in combination with GAT211 (two-tailed t-test using Welch’s correction) (D). Testing for conditioned place preference/aversion was performed on day 12 in a drug-free state. Data are expressed as mean ± SEM (n = 10 per group). **P < 0.01, ****P < 0.001 vs. vehicle-paired chamber, two-way ANOVA followed by Bonferroni’s post hoc test. n.s., non-significant.
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