Cannabinoid CB2 receptors in primary sensory neurons are implicated in CB2 agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance

doi:10.1016/j.biopha.2024.116879

. 2024 Jul:176:116879.

doi: 10.1016/j.biopha.2024.116879. Epub 2024 Jun 7.

Cannabinoid CB₂ receptors in primary sensory neurons are implicated in CB₂ agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance

Kelsey G Guenther¹, Xiaoyan Lin¹, Zhili Xu¹, Alexandros Makriyannis², Julian Romero³, Cecilia J Hillard⁴, Ken Mackie⁵, Andrea G Hohmann⁶

Affiliations

¹ Program in Neuroscience, Indiana University, Bloomington, IN, United States; Department of Psychological and Brain Sciences, Indiana University, Bloomington, IN, United States.
² Center for Drug Discovery, Northeastern University, Boston, MA, United States.
³ Faculty of Experimental Sciences, Universidad Francisco de Vitoria, 28223 Pozuelo de Alarcón, Madrid, Spain.
⁴ Department of Pharmacology and Toxicology, Med. Col. Of Wisconsin, Milwaukee, WI, United States.
⁵ Program in Neuroscience, Indiana University, Bloomington, IN, United States; Department of Psychological and Brain Sciences, Indiana University, Bloomington, IN, United States; Gill Center for Biomolecular Science, Indiana University, Bloomington, IN, United States.
⁶ Program in Neuroscience, Indiana University, Bloomington, IN, United States; Department of Psychological and Brain Sciences, Indiana University, Bloomington, IN, United States; Gill Center for Biomolecular Science, Indiana University, Bloomington, IN, United States. Electronic address: hohmanna@indiana.edu.

PMID: 38850666
PMCID: PMC11209786
DOI: 10.1016/j.biopha.2024.116879

Cannabinoid CB₂ receptors in primary sensory neurons are implicated in CB₂ agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance

Kelsey G Guenther et al. Biomed Pharmacother. 2024 Jul.

. 2024 Jul:176:116879.

doi: 10.1016/j.biopha.2024.116879. Epub 2024 Jun 7.

Authors

Kelsey G Guenther¹, Xiaoyan Lin¹, Zhili Xu¹, Alexandros Makriyannis², Julian Romero³, Cecilia J Hillard⁴, Ken Mackie⁵, Andrea G Hohmann⁶

Affiliations

¹ Program in Neuroscience, Indiana University, Bloomington, IN, United States; Department of Psychological and Brain Sciences, Indiana University, Bloomington, IN, United States.
² Center for Drug Discovery, Northeastern University, Boston, MA, United States.
³ Faculty of Experimental Sciences, Universidad Francisco de Vitoria, 28223 Pozuelo de Alarcón, Madrid, Spain.
⁴ Department of Pharmacology and Toxicology, Med. Col. Of Wisconsin, Milwaukee, WI, United States.
⁵ Program in Neuroscience, Indiana University, Bloomington, IN, United States; Department of Psychological and Brain Sciences, Indiana University, Bloomington, IN, United States; Gill Center for Biomolecular Science, Indiana University, Bloomington, IN, United States.
⁶ Program in Neuroscience, Indiana University, Bloomington, IN, United States; Department of Psychological and Brain Sciences, Indiana University, Bloomington, IN, United States; Gill Center for Biomolecular Science, Indiana University, Bloomington, IN, United States. Electronic address: hohmanna@indiana.edu.

PMID: 38850666
PMCID: PMC11209786
DOI: 10.1016/j.biopha.2024.116879

Abstract

Cannabinoid CB₂ agonists show therapeutic efficacy without unwanted CB₁-mediated side effects. The G protein-biased CB₂ receptor agonist LY2828360 attenuates the maintenance of chemotherapy-induced neuropathic nociception in male mice and blocks development of morphine tolerance in this model. However, the cell types involved in this phenomenon are unknown and whether this therapeutic profile is observed in female mice has never been investigated. We used conditional deletion of CB₂ receptors to determine the cell population(s) mediating the anti-allodynic and morphine-sparing effects of CB₂ agonists. Anti-allodynic effects of structurally distinct CB₂ agonists (LY2828360 and AM1710) were present in paclitaxel-treated CB₂^f/f mice and in mice lacking CB₂ receptors in CX3CR1 expressing microglia/macrophages (CX3CR1^CRE/+; CB₂^f/f), but were absent in mice lacking CB₂ receptors in peripheral sensory neurons (Advillin^CRE/+; CB₂^f/f). The morphine-sparing effect of LY28282360 occurred in a sexually-dimorphic manner, being present in male, but not female, mice. LY2828360 treatment (3 mg/kg per day i.p. x 12 days) blocked the development of morphine tolerance in male CB₂^f/f and CX3CR1^CRE/+; CB₂^f/f mice with established paclitaxel-induced neuropathy but was absent in male (or female) Advillin^CRE/+; CB₂^f/f mice. Co-administration of morphine with a low dose of LY2828360 (0.1 mg/kg per day i.p. x 6 days) reversed morphine tolerance in paclitaxel-treated male CB₂^f/f mice, but not Advillin^CRE/+; CB₂^f/f mice of either sex. LY2828360 (3 mg/kg per day i.p. x 8 days) delayed, but did not prevent, the development of paclitaxel-induced mechanical or cold allodynia in either CB₂^f/f or CX3CR1^CRE/+; CB₂^f/f mice of either sex. Our findings have potential clinical implications.

Keywords: CB2; Cancer; Chemotherapy-induced peripheral neuropathy; Dorsal root ganglion; Opioid tolerance.

PubMed Disclaimer

Conflict of interest statement

Declaration of Competing Interest The authors have no financial/personal interest or belief that could affect the objectivity of this manuscript. Dr. Hohmann is a consultant for Anagin Inc.

Figures

**Fig. 1.**
Comparison of CB₂ mRNA (A, B, C, D) and GFP mRNA (E, F, G, H) expression in DRG (A, E), spinal cord (B, F), paw skin (C, G) and spleen (D, H) of male and female ${C B}_{2}^{f / f}$ (n = 8 male, 5 female), Advillin^CRE/+; ${C B}_{2}^{f / f}$ , (n = 5 female) and wild type (n = 5 male, 3 female) mice. Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice showed lower levels of CB₂ mRNA compared to both ${C B}_{2}^{f / f}$ and WT mice and lower levels of GFP compared to ${C B}_{2}^{f / f}$ mice. WT mice did not show any GFP mRNA expression. Mean (±SEM) 2^−ΔΔCT values for experiment 1 (unpaired two-tailed t-tests). Asterisk indicates a group difference; *p < 0.05, ***p < 0.001.

**Fig. 2.**
Mechanical (A) and cold (B) allodynia develop normally in ${C B}_{2}^{f / f}$ and Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice following paclitaxel treatment (n = 4–7 per group, mixed sex). Data expressed as mean ±SEM, *p < 0.05, **p < 0.005, ***p < 0.001 (two-way ANOVA, Sidak’s multiple comparisons test). Gray asterisk indicate difference between ${C B}_{2}^{f / f}$ groups; black asterisk indicate difference between Advillin^CRE/+; ${C B}_{2}^{f / f}$ groups.

**Fig. 3.**
LY2828360 dose dependently alleviated paclitaxel evoked mechanical (A) and cold (B) allodynia in ${C B}_{2}^{f / f}$ , but not Advillin^CRE/+; ${C B}_{2}^{f / f}$ male mice. LY2828360 did not alter mechanical (C) or cold (D) sensitivity in ${C B}_{2}^{f / f}$ or Advillin^CRE/+; ${C B}_{2}^{f / f}$ control male mice treated with cremophor vehicle (n = 5–6 per group). Data expressed as mean ±SEM, *p < 0.05, ***p < 0.001 (two-way ANOVA, Sidak’s multiple comparisons test).

**Fig. 4.**
AM1710 dose dependently alleviated paclitaxel-evoked sensitization to mechanical (A) and cold (B) stimulation in ${C B}_{2}^{f / f}$ , but not Advillin^CRE/+; ${C B}_{2}^{f / f}$ male mice. AM1710 did not alter mechanical (C) or cold (D) sensitivity in ${C B}_{2}^{f / f}$ or Advillin^CRE/+; ${C B}_{2}^{f / f}$ control male mice treated with cremophor vehicle (n = 5–6 per group). Data expressed as mean ±SEM, **p < 0.005, ***p < 0.001 (two-way ANOVA, Sidak’s multiple comparisons test).

**Fig. 5.**
History of chronic LY2828360 treatment blocked the development of morphine tolerance in male, but not female, ${C B}_{2}^{f / f}$ mice, whereas morphine tolerance developed in Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice of either sex. (A) Schematic shows the injection schedule used to evaluate the two-phase treatment during the maintenance phase of paclitaxel-induced neuropathic pain. Chronic LY2828360 treatment (3 mg/kg per day i.p. x 12 days) alleviated paclitaxel evoked mechanical (B) and cold (C) allodynia in male ${C B}_{2}^{f / f}$ , but not Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice and blocked the development of tolerance to the anti-allodynic effects of morphine (10 mg/kg per day i. p. x 12 days). LY2828360 had no effect on mechanical (D) or cold (E) allodynia in female ${C B}_{2}^{f / f}$ or Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice and did not block the development of tolerance to the anti-allodynic effects of morphine (n = 8–9 per group). Data expressed as mean ±SEM, ***p < 0.001 (two-way ANOVA, Sidak’s multiple comparisons test).

**Fig. 6.**
Impact of a history of chronic LY2828360 (3 mg/kg per day i.p. x 12 days) treatment on naloxone-precipitated morphine withdrawal in male and female ${C B}_{2}^{f / f}$ and Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice used in Experiment 4. Following chronic treatment with morphine (10 mg/kg per day i.p. x 12 days) male mice show fewer withdrawal jumps compared to female mice regardless of genotype (n = 8–9 per group). Data expressed as mean ±SEM, *p < 0.05 (two-way ANOVA).

**Fig. 7.**
Co-administration with LY2828360 reverses morphine tolerance in male, but not female ${C B}_{2}^{f / f}$ mice, and had no effect on morphine tolerance in Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice of either sex. (A) Schematic shows the injection schedule used to evaluate the two-phase treatment during the maintenance phase of paclitaxel-induced neuropathic pain. Chronic administration of morphine (10 mg/kg per day i.p. x 6 days) initially alleviated paclitaxel-evoked mechanical (B, D) and cold (C, E) allodynia in both male and female ${C B}_{2}^{f / f}$ and Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice, but later resulted in tolerance. Co-administration of morphine with low-dose LY2828360 (0.1 mg/kg per day i.p. x 6 days) caused a reversal of tolerance to morphine’s mechanical (B) and cold (C) anti-allodynic effects in male ${C B}_{2}^{f / f}$ , but not male Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice. LY2828360 had no effect on mechanical (D) or cold (E) allodynia in female ${C B}_{2}^{f / f}$ or female Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice and did not block the development of tolerance to the anti-allodynic effects of morphine (n = 6–8 per group). Data expressed as mean ±SEM, ***p < 0.001 (two-way ANOVA, Sidak’s multiple comparisons test).

**Fig. 8.**
Impact of co-administration of low-dose LY2828360 with morphine (0.1 and 10 mg/kg per day i.p. x 6 days, respectively) on naloxone-precipitated morphine withdrawal in male and female ${C B}_{2}^{f / f}$ and Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice used in Experiment 5. ${C B}_{2}^{f / f}$ mice treated with morphine + vehicle instead of LY2828360 showed an increased number of jumps compared to the LY2828360 + morphine group. No difference in withdrawal jumps was observed between groups in Advillin^CRE/+; ${C B}_{2}^{f / f}$ mice (n = 6–8 per group). Data expressed as mean ±SEM, *p < 0.05 (two-way ANOVA).

**Fig. 9.**
LY2828360 dose-dependently alleviated paclitaxel evoked mechanical (A) and cold (B) allodynia in both ${C B}_{2}^{f / f}$ and CX3CR1^CRE/+; ${C B}_{2}^{f / f}$ mice. LY2828360 did not alter mechanical (C) or cold (D) sensitivity in ${C B}_{2}^{f / f}$ and CX3CR1^CRE/+; ${C B}_{2}^{f / f}$ control mice treated with cremophor vehicle (n = 6–16 per group, mixed sex). Data expressed as mean ±SEM (two-way ANOVA).

**Fig. 10.**
History of chronic LY2828360 treatment blocked the development of morphine tolerance in male, but not female ${C B}_{2}^{f / f}$ and CX3CR1^CRE/+; ${C B}_{2}^{f / f}$ mice. (A) Schematic shows the injection schedule used to evaluate the two-phase treatment during the maintenance phase of paclitaxel-induced neuropathic pain. Chronic LY2828360 treatment (3 mg/kg per day i.p. x 12 days) alleviated paclitaxel evoked mechanical (B, D) and cold (C, E) allodynia and blocked the development of tolerance to the anti-allodynic effects of morphine (10 mg/kg per day i.p. x 12 days) in male (B, C), but not female (D, E) ${C B}_{2}^{f / f}$ and CX3CR1^CRE/+; ${C B}_{2}^{f / f}$ mice (n = 6–8 per group). Data expressed as mean ±SEM (two-way ANOVA).

**Fig. 11.**
Impact of a history of chronic LY2828360 (3 mg/kg per day i.p. x 12 days) on naloxone-precipitated morphine withdrawal in male and female ${C B}_{2}^{f / f}$ and CX3CR1^CRE/+; ${C B}_{2}^{f / f}$ mice used in Experiment 7 (n = 6–8 per group). No differences were found between genotype or sex. Data expressed as mean ±SEM, *p < 0.05 (two-way ANOVA).

**Fig. 12.**
LY2828360 (3 mg/kg per day i.p. x 8 days) delayed the onset of paclitaxel-induced mechanical (A) and cold (B) allodynia but did not prevent their development. Anti-allodynic effects of LY2828360 were no longer observed 4–7 days following termination of dosing (n = 5 per group, mixed sex). Data expressed as mean ± SEM, **p < 0.005, ***p < 0.001 vs. ${C B}_{2}^{f / f}$ comparator groups (two-way ANOVA, Sidak’s multiple comparisons test).

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Update of

Cannabinoid CB ₂ receptors in primary sensory neurons are implicated in CB ₂ agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance.
Guenther KG, Lin X, Xu Z, Makriyannis A, Romero J, Hillard CJ, Mackie K, Hohmann AG. Guenther KG, et al. bioRxiv [Preprint]. 2024 Mar 10:2024.03.05.583426. doi: 10.1101/2024.03.05.583426. bioRxiv. 2024. Update in: Biomed Pharmacother. 2024 Jul;176:116879. doi: 10.1016/j.biopha.2024.116879 PMID: 38496640 Free PMC article. Updated. Preprint.

Cited by

Chemotherapy-Induced Peripheral Neuropathy: A Recent Update on Pathophysiology and Treatment.
Mattar M, Umutoni F, Hassan MA, Wamburu MW, Turner R, Patton JS, Chen X, Lei W. Mattar M, et al. Life (Basel). 2024 Aug 9;14(8):991. doi: 10.3390/life14080991. Life (Basel). 2024. PMID: 39202733 Free PMC article. Review.

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[1] Yekkirala AS, Roberson DP, Bean BP, Woolf CJ, Breaking barriers to novel analgesic drug development, Nat. Rev. Drug Discov 16 (8) (2017) 545–564. - PMC - PubMed

[2] Yekkirala AS, Roberson DP, Bean BP, Woolf CJ, Breaking barriers to novel analgesic drug development, Nat. Rev. Drug Discov 16 (8) (2017) 545–564. - PMC - PubMed

[3] Coluzzi F, Rullo L, Scerpa MS, Losapio LM, Rocco M, Billeci D, Candeletti S, Romualdi P, Current and Future Therapeutic Options in Pain Management: Multi-mechanistic Opioids Involving Both MOR and NOP Receptor Activation, CNS Drugs 36 (6) (2022) 617–632. - PMC - PubMed

[4] Coluzzi F, Rullo L, Scerpa MS, Losapio LM, Rocco M, Billeci D, Candeletti S, Romualdi P, Current and Future Therapeutic Options in Pain Management: Multi-mechanistic Opioids Involving Both MOR and NOP Receptor Activation, CNS Drugs 36 (6) (2022) 617–632. - PMC - PubMed

[5] Habibi-Asl B, Vaez H, Najafi M, Bidaghi A, Ghanbarzadeh S, Attenuation of morphine-induced dependence and tolerance by ceftriaxone and amitriptyline in mice, Acta Anaesthesiol. Taiwan 52 (2014) 163–168. - PubMed

[6] Habibi-Asl B, Vaez H, Najafi M, Bidaghi A, Ghanbarzadeh S, Attenuation of morphine-induced dependence and tolerance by ceftriaxone and amitriptyline in mice, Acta Anaesthesiol. Taiwan 52 (2014) 163–168. - PubMed

[7] Mansouri MT, Khodayar MJ, Tabatabaee A, Ghorbanzadeh B, Naghizadeh B, Modulation of morphine antinociceptive tolerance and physical dependence by co-administration of simvastatin, Pharmacol. Biochem. Behav 137 (2015) 38–43. - PubMed

[8] Mansouri MT, Khodayar MJ, Tabatabaee A, Ghorbanzadeh B, Naghizadeh B, Modulation of morphine antinociceptive tolerance and physical dependence by co-administration of simvastatin, Pharmacol. Biochem. Behav 137 (2015) 38–43. - PubMed

[9] Hassanipour M, Amini-Khoei H, Shafaroodi H, Shirzadian A, Rahimi N, Imran-Khan M, Rezayat SM, Dehpour A, Atorvastatin attenuates the antinociceptive tolerance of morphine via nitric oxide dependent pathway in male mice, Brain. Res. Bull 125 (2016) 173–180. - PubMed

[10] Hassanipour M, Amini-Khoei H, Shafaroodi H, Shirzadian A, Rahimi N, Imran-Khan M, Rezayat SM, Dehpour A, Atorvastatin attenuates the antinociceptive tolerance of morphine via nitric oxide dependent pathway in male mice, Brain. Res. Bull 125 (2016) 173–180. - PubMed

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Cannabinoid CB₂ receptors in primary sensory neurons are implicated in CB₂ agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance

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Cannabinoid CB₂ receptors in primary sensory neurons are implicated in CB₂ agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance

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Abstract

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Abstract

Conflict of interest statement

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