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. 2023 Jun 1;164(6):1375-1387.
doi: 10.1097/j.pain.0000000000002828. Epub 2022 Dec 6.

Neuroendocrine mechanisms in oxaliplatin-induced hyperalgesic priming

Larissa Staurengo-Ferrari  1 , Dionéia Araldi  1 , Paul G Green  1   2 , Jon D Levine  1   2   3
Affiliations

Neuroendocrine mechanisms in oxaliplatin-induced hyperalgesic priming

Larissa Staurengo-Ferrari et al. Pain. .

Abstract

Stress plays a major role in the symptom burden of oncology patients and can exacerbate cancer chemotherapy-induced peripheral neuropathy (CIPN), a major adverse effect of many classes of chemotherapy. We explored the role of stress in the persistent phase of the pain induced by oxaliplatin. Oxaliplatin induced hyperalgesic priming, a model of the transition to chronic pain, as indicated by prolongation of hyperalgesia produced by prostaglandin E 2 , in male rats, which was markedly attenuated in adrenalectomized rats. A neonatal handling protocol that induces stress resilience in adult rats prevented oxaliplatin-induced hyperalgesic priming. To elucidate the role of the hypothalamic-pituitary-adrenal and sympathoadrenal neuroendocrine stress axes in oxaliplatin CIPN, we used intrathecally administered antisense oligodeoxynucleotides (ODNs) directed against mRNA for receptors mediating the effects of catecholamines and glucocorticoids, and their second messengers, to reduce their expression in nociceptors. Although oxaliplatin-induced hyperalgesic priming was attenuated by intrathecal administration of β 2 -adrenergic and glucocorticoid receptor antisense ODNs, oxaliplatin-induced hyperalgesia was only attenuated by β 2 -adrenergic receptor antisense. Administration of pertussis toxin, a nonselective inhibitor of Gα i/o proteins, attenuated hyperalgesic priming. Antisense ODNs for Gα i 1 and Gα o also attenuated hyperalgesic priming. Furthermore, antisense for protein kinase C epsilon, a second messenger involved in type I hyperalgesic priming, also attenuated oxaliplatin-induced hyperalgesic priming. Inhibitors of second messengers involved in the maintenance of type I (cordycepin) and type II (SSU6656 and U0126) hyperalgesic priming both attenuated hyperalgesic priming. These experiments support a role for neuroendocrine stress axes in hyperalgesic priming, in male rats with oxaliplatin CIPN.

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Conflict of interest statement

The authors have no conflict of interest to declare.

Figures

Figure 1.
Figure 1.
Oxaliplatin induces long-lasting mechanical hyperalgesia and hyperalgesic priming in male rats. Male rats received a single intravenous injection of vehicle (0.9% saline) or oxaliplatin (2 mg/kg), and mechanical nociceptive threshold was evaluated before their administration (day 0) and on days 1, 7, 14, 21, 28, 42, and 60 after administration. Oxaliplatin-induced hyperalgesic priming was evaluated on days 21, 42, and 60. (A) The experimental protocol, with timing of treatments and nociceptive threshold measurements. (B) Oxaliplatin-induced mechanical hyperalgesia that lasted 42 days; *P<0.05, **P<0.01. (C) PGE2 (100 ng/5 μL, i.d.) was injected on the dorsum of 1 hind paw, and the mechanical nociceptive threshold was evaluated 30 minutes and 4 hours later to test for hyperalgesic priming. Hyperalgesic priming was evaluated before oxaliplatin and again 21, 42, and 60 days after oxaliplatin administration. PGE2-induced hyperalgesia at 30 minutes, and before administration of oxaliplatin, PGE2-induced hyperalgesia was no longer present at the fourth hour (30 minutes vs 4 hours, P = 0.0005, Student paired 2-tailed t test, t5 = 8.134). However, after treatment with oxaliplatin, the hyperalgesia induced by PGE2 was still present at the fourth hour 21, 42, and 60 days after oxaliplatin (for each time point, 30 minutes vs 4 hours, P = NS). Data are shown as mean ± SEM. PGE2, prostaglandin E2.
Figure 2.
Figure 2.
Role of neuroendocrine stress axes in oxaliplatin-induced hyperalgesia and priming. Male rats were submitted to sham bilateral adrenalectomy (adrenal intact animals), bilateral adrenalectomy only, or bilateral adrenalectomy plus stress hormone (epinephrine and corticosterone) replacements. Mechanical nociceptive threshold was evaluated before oxaliplatin administration (day 0) and again on day 21 after oxaliplatin administration. On day 21, after measurement of mechanical threshold, PGE2 (100 ng/paw) was injected intradermally to assess for hyperalgesic priming (ie, prolongation of PGE2-induced hyperalgesia). Mechanical nociceptive threshold was evaluated 30 minutes and 4 hours after PGE2. (A) The experimental protocol, with timing of treatments and nociceptive threshold measurements. (B) When the magnitude of oxaliplatin-induced hyperalgesia was evaluated on day 21, it was markedly attenuated in the oxaliplatin-treated adrenalectomized rats, compared with the adrenal intact group. However, in the adrenalectomized rats replaced with stress hormones, oxaliplatin-induced hyperalgesia was of similar magnitude to that observed in adrenal intact rats, supporting a contribution of stress hormones to oxaliplatin-induced hyperalgesia. Data are shown as mean ± SEM, treatment F(2,15) = 14.45; **P < 0.01: sham adrenalectomy group vs adrenalectomy group, ***P < 0.001: adrenalectomy group vs adrenalectomy plus stress hormone replacement (n = 6, 1-way repeated-measures ANOVA followed by Bonferroni post hoc test). (C) PGE2-induced hyperalgesia at 30 minutes, in all groups, that was no longer present at the fourth hour, in adrenalectomized rats. However, PGE2-induced hyperalgesia was still present 4 hours after its injection in the sham adrenalectomized group (adrenal intact animals) and in the adrenalectomized group that received replacement stress hormones, in support of a contribution of stress hormones to oxaliplatin-induced hyperalgesic priming. Data are shown as mean ± SEM, treatment F(2,15) = 14.45; **P < 0.01: sham adrenalectomy group vs adrenalectomized group, ***P < 0.001: adrenalectomy group vs adrenalectomy plus stress hormone replacement (n = 6, 1-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM, time F(1,30) = 5.922; treatment F(2,30) = 6.024; interaction F(2,30) = 4.143, **P < 0.01: sham adrenalectomy group vs adrenalectomy group, *P < 0.05: adrenalectomized group vs adrenalectomy plus stress hormones group (n = 6 rats/group), 2-way repeated-measures ANOVA followed by Bonferroni post hoc test. Adx, adrenalectomized; ANOVA, analysis of variance; PGE2, prostaglandin E2.
Figure 3.
Figure 3.
Role of the β2-adrenergic and glucocorticoid receptor in oxaliplatin-induced priming. Male rats received a single intravenous injection of oxaliplatin (2 mg/kg) on day 0, and starting on day 18, MM-ODNs or AS-ODNs (both 120 μg/20 μL, gray bar) to ADRB2 or GR mRNA were injected intrathecally for 3 consecutive days. On day 21 after administration of oxaliplatin, ∼17 hours after the last ODN injection, mechanical nociceptive threshold was measured, and PGE2 (100 ng/5 μL) was injected intradermally. Thirty minutes and 4 hours after injection of PGE2, the mechanical nociceptive threshold was again measured to assess for hyperalgesic priming. (A) The experimental protocol providing timing of treatments and nociceptive threshold measurements. (B) When the magnitude of oxaliplatin-induced hyperalgesia was evaluated on day 21, an inhibition in oxaliplatin-induced hyperalgesia (t10 = 2.321, *P = 0.0427: ADRB2 AS-ODN–treated group vs ADRB2 MM-ODN–treated group, unpaired Student t test) and the prolongation of PGE2 hyperalgesia were observed in ADRB2 AS-ODN–treated group (time F(1,20) = 9.624; treatment F(1,20) = 2.274; interaction F(1,20) = 9.768, **P < 0.01: ADRB2 AS-ODN–treated group vs ADRB2 MM-ODN–treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n = 6 rats/group). These findings support the suggestion that oxaliplatin-induced hyperalgesia and hyperalgesic priming are β2-adrenergic receptor-dependent. (C) When the magnitude of oxaliplatin-induced hyperalgesia was evaluated on day 21, no differences between the GR AS-ODN–treated and MM-ODN–treated groups were observed (t10 = 1.794, P = 0.1100: GR AS-ODN–treated group vs GR MM-ODN–treated group, unpaired Student t test). However, an inhibition of the prolongation of PGE2-induced hyperalgesia was observed in the GR AS-ODN–treated group, when compared with the GR MM-ODN–treated group (time F(1,20) = 6.420; treatment F(1,20) = 5.427; interaction F(1,20) = 10.10, ***P < 0.002: GR AS-ODN–treated group vs GR MM-ODN–treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n = 6 rats/group). These findings indicate that although oxaliplatin-induced hyperalgesia is glucocorticoid receptor-independent, in oxaliplatin-induced priming, this receptor plays a significant role. ADRB2, β2-adrenergic receptor; ANOVA, analysis of variance; AS, antisense; BL, antisense; GR, glucocorticoid receptor; ODN, oligodeoxynucleotide; ; MM, mismatch; PGE2, prostaglandin E2.
Figure 4.
Figure 4.
Neonatal handling-induced stress resilience attenuates oxaliplatin-induced hyperalgesia and hyperalgesic priming in adult rats. Male rats were submitted to neonatal handling and as adults received oxaliplatin. (A) The experimental protocol providing timing of treatments and nociceptive threshold measurements. (B) Neonatal handling prevented oxaliplatin-induced hyperalgesia, when compared with control adult rats that received oxaliplatinneonatal handling, oxaliplatin vs oxaliplatin, ****P2, prostaglandin E2.
Figure 5.
Figure 5.
Role of GPCR Gi/o subunits in oxaliplatin-induced hyperalgesia and hyperalgesic priming. Male rats received a single intravenous injection of oxaliplatin (2 mg/kg), and the mechanical nociceptive threshold was evaluated before oxaliplatin administration (day 0) and again on day 20 after its administration. On day 20, after measurement of mechanical nociceptive threshold, vehicle (saline) or pertussis toxin, which inactivates diverse Gαi/o subunits, was injected intradermally (1 µg/5 µL). On day 21, ∼17 hours after pertussis toxin injection, mechanical nociceptive threshold was evaluated, and PGE2 (100 ng/paw) was then injected intradermally. In these experiments, mechanical nociceptive threshold was evaluated 30 minutes and 4 hours after injection of PGE2 to assess for hyperalgesic priming. (A) The experimental protocol providing timing of treatments and nociceptive threshold measurements. (B) When the magnitude of oxaliplatin-induced hyperalgesia was evaluated on day 21, no difference between the pertussis toxin–treated and vehicle-treated groups was observed (t10 = 0.3922, P = 0.6817: pertussis toxin–treated group vs vehicle-treated group, unpaired Student t test). (C) However, inhibition of the prolongation of PGE2 was observed in the pertussis toxin–treated group, when compared with the vehicle-treated group (time F(1,20) = 14; treatment F(1,20) = 10.38; interaction F(1,20) = 6.415, **P < 0.01: for pertussis toxin–treated group vs vehicle-treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n = 6 rats/group). These findings indicate that although oxaliplatin-induced hyperalgesia is Gαi/o-GPCR subunit-independent, they contribute to oxaliplatin-induced hyperalgesic priming. ANOVA, analysis of variance; BL, baseline; GPCR, G protein-coupled receptor; Gα/o, G-protein subunit α/o; PGE2, prostaglandin E2.
Figure 6.
Figure 6.
Role of GPCR Gi/o subunits in oxaliplatin-induced priming. Male rats received a single intravenous injection of oxaliplatin (2 mg/kg), and on day 18 after its administration, SE-ODNs (120 μg/20 μL, black bar) or AS-ODNs (120 μg/20 μL, gray bar) against Gαi1, Gαi2, Gαi3, or Gαo, mRNA was injected intrathecally for 3 consecutive days. On day 21, ∼17 hours after the last ODN injection, mechanical nociceptive threshold was measured, and then PGE2 (100 ng/5 μL) was injected intradermally. Thirty minutes and 4 hours after PGE2, the mechanical nociceptive threshold was again measured to assess for hyperalgesic priming. (A) The experimental protocol providing timing of treatments and nociceptive threshold measurements. (B) An inhibition of the prolongation of PGE2 effect was observed in the Gαi1 AS-ODN–treated group, when compared with the Gαi1 SE-ODN group (time F(1,20) = 26.36; treatment F(1,20) = 16.25; interaction F(1,20) = 8.369, **P < 0.01: Giα1 AS-ODN–treated group vs Gαi1 SE-ODN–treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n = 6 rats/group). These findings indicate that oxaliplatin-induced hyperalgesic priming is Gαi1-dependent. (C) An inhibition of the prolongation of PGE2 hyperalgesia was observed in the Goα AS-ODN–treated group, when compared with the Goα SE-ODN–treated group (time F(1,20) = 46.27; treatment F(1,20) = 58.32; interaction F(1,20) = 42.99, **P < 0.0001: Goα AS-ODN–treated group vs Gαo SE-ODN–treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n= 6 rats/group). These findings indicate that oxaliplatin-induced hyperalgesic priming is Gαo-dependent. Data are shown as mean ± SEM (n = 6 rats/group). (D and E) The prolongation of PGE2 was present in both Gαi3 (C), Gαi2 (D) AS-ODNs, and their respective SE-ODN–treated groups, with no statistical differences between these 2 groups (C: time F(1,20) = 5.067; treatment F(1,20) = 0.1166; interaction F(1,20) = 0.6648, P = 0.4245 Gαi3 AS-ODN–treated group and SE-treated group; D: time F(1,20) = 1.024; treatment F(1,20) = 0.3675; interaction F(1,20) = 0.9293, P = 0.5601: Gαi2 AS-ODN–treated group and SE-treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n = 6 rats/group). These findings support the suggestion that oxaliplatin-induced hyperalgesic priming is Gαi3- and Gαi2-independent. ANOVA, analysis of variance; AS, antisense; BL, baseline; GPCR, G protein-coupled receptor; Gα/o, G-protein subunit α/o; ODN, oligodeoxynucleotide; PGE2, prostaglandin E2.
Figure 7.
Figure 7.
Oxaliplatin induces type I and II hyperalgesic priming. Male rats received a single intravenous injection of oxaliplatin (2 mg/kg). Twenty-one days later, the protein translation inhibitor, cordycepin (1 µg/20 µL, i.d.) or the combination of an Src kinase inhibitor (SU6656, 1 µg/2 µL, i.d.) and an MAPK inhibitor (U0126, 1 µg/2 µL, i.d.) was administered, on the dorsum of the hind paw at the site of nociceptive threshold testing, followed 15 minutes later by PGE2 (100 ng/5 µL, i.d.) at the same site. In the group that received only oxaliplatin, nociceptive threshold was evaluated before and 21 days after oxaliplatin and again after administration of inhibitors alone or with PGE2. (A) The experimental protocol providing timing of treatments and nociceptive threshold measurements. (B) Both cordycepin and the combination of SU6656 and U0126 attenuate oxaliplatin-induced hyperalgesia measured on day 21 after administration of oxaliplatin (time F(2, 30) = 8.909, treatment F(1, 30) = 7.235, **P < 0.01: oxaliplatin, vehicle group vs oxaliplatin, cordycepin at 45′, *P < 0.01: oxaliplatin, vehicle group vs oxaliplatin, cordycepin at 4 hours or *P < 0.01: oxaliplatin, vehicle group vs oxaliplatin, SU6656 + U0126 at 45, 1-way repeated-measures ANOVA followed by Bonferroni post hoc test). No significant differences between the vehicle-treated group and the group treated with SU6656 plus U0126 were detected at the fourth hour, indicating the transitory effect of this combination of inhibitors on oxaliplatin-induced hyperalgesia. (C) Hyperalgesic priming, induced by oxaliplatin, is robustly attenuated by cordycepin or by a combination of SU6656 and U0126 because the prolongation of the PGE2 hyperalgesia was observed only in the vehicle-treated group. Data are shown as mean ± SEM (n = 6 rats/group). These findings support the suggestion that protein translation in nociceptors plays a role in oxaliplatin-induced hyperalgesia and oxaliplatin-induced hyperalgesic priming. ANOVA, analysis of variance; AS, antisense; BL, baseline; MM, mismatch; PKCε, protein kinase C ε; PGE2, prostaglandin E2.
Figure 8.
Figure 8.
Role of PKCε in oxaliplatin-induced hyperalgesia and priming. Male rats received a single intravenous injection of oxaliplatin (2 mg/kg), and on day 18 after administration, MM-ODNs (120 μg/20 μL, black bar) or AS-ODNs (120 μg/20 μL, gray bar) against PKCε mRNA were injected for 3 consecutive days. On day 21, ∼17 hours after the last ODN injection, mechanical nociceptive threshold was measured, and PGE2 (100 ng/5 μL) was injected intradermally. Thirty minutes and 4 hours after PGE2, the mechanical nociceptive threshold was again measured to assess for hyperalgesic priming. (A) The experimental protocol providing timing of treatments and nociceptive threshold measurement. (B) When the magnitude of oxaliplatin-induced hyperalgesia was evaluated on day 21, no differences between PKCε AS-ODN– and MM-ODN–treated groups were observed (t10 = 1.839, P = 0.4203: PKCε AS-ODN–treated group vs PKCε MM-ODN–treated group, unpaired Student t test). (C) PGE2 induced significant hyperalgesia at 30 minutes in PKCε AS-ODN– and MM-ODN–treated group that was no longer present at the fourth hour in the PKCε AS-ODN–treated group (time F(1,20) = 19.73; treatment F(1,20) = 8.417; interaction F(1,20) = 10.22, ***P < 0.01: PKCε AS-ODN–treated group vs PKCε AS-ODN MM-ODN–treated group, 2-way repeated-measures ANOVA followed by Bonferroni post hoc test). Data are shown as mean ± SEM (n= 6 rats/group). These findings support the suggestion that although oxaliplatin-induced hyperalgesia is PKCε-independent, PKCε plays a role in oxaliplatin-induced hyperalgesic priming. ANOVA, analysis of variance; ODN, oligodeoxynucleotide; PGE2, prostaglandin E2.
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