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Background and Purpose: Mitochondrial dysfunction contributes to the pathogenesis and maintenance of chemotherapy-induced peripheral neuropathy (CIPN), a significant limitation of cancer chemotherapy. Recently, the stimulation of mitophagy, a pivotal process for mitochondrial homeostasis, has emerged as a promising treatment strategy for neurodegenerative diseases, but its therapeutic effect on CIPN has not been explored. Here, we assessed the mitophagy-inducing activity of 3,5-dibromo-2-(2',4'-dibromophenoxy)-phenol (PDE701), a diphenyl ether derivative isolated from the marine sponge Dysidea sp., and investigated its therapeutic effect on a CIPN model., Experimental Approach: Mitophagy activity was determined by a previously established mitophagy assay using mitochondrial Keima (mt-Keima). Mitophagy induction was further verified by western blotting, immunofluorescence, and electron microscopy. Mitochondrial dysfunction was analysed by measuring mitochondrial superoxide levels in SH-SY5Y cells and Drosophila larvae. A thermal nociception assay was used to evaluate the therapeutic effect of PDE701 on the paclitaxel-induced thermal hyperalgesia phenotype in Drosophila larvae., Key Results: PDE701 specifically induced mitophagy but was not toxic to mitochondria. PDE701 ameliorated paclitaxel-induced mitochondrial dysfunction in both SH-SY5Y cells and Drosophila larvae. Importantly, PDE701 also significantly ameliorated paclitaxel-induced thermal hyperalgesia in Drosophila larvae. Knockdown of ATG5 or ATG7 abolished the effect of PDE701 on thermal hyperalgesia, suggesting that PDE701 exerts its therapeutic effect through mitophagy induction., Conclusion and Implications: This study identified PDE701 as a novel mitophagy inducer and a potential therapeutic compound for CIPN. Our results suggest that mitophagy stimulation is a promising strategy for the treatment of CIPN and that marine organisms are a potential source of mitophagy-inducing compounds., (© 2024 The Author(s). British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Background and Purpose: We extend the characterization of the TRPM8 antagonist VBJ103 with tests of selectivity, specificity and distribution, therapeutic efficacy of systemic administration against oxaliplatin-induced cold hyperalgesia and the impact of systemic administration on core body temperature (CBT)., Experimental Approach: Selectivity at human TRPA1 and TRPV1 as well as in vitro safety profiling was determined. Effects of systemic administration of VBJ103 were evaluated in a model of oxaliplatin-induced cold hyperalgesia. Both peripheral and centrally mediated effects of VBJ103 on CBT were assessed with radiotelemetry., Key Results: VBJ103 had no antagonist activity at TRPV1 and TRPA1, but low potency TRPA1 activation. The only safety liability detected was partial inhibition of the dopamine transporter (DAT). VBJ103 delivered subcutaneously dose-dependently attenuated cold hypersensitivity in oxaliplatin-treated mice at 3, 10 and 30 mg·kg -1 (n = 7, P < 0.05). VBJ103 (30 mg·kg -1 ) antinociception was influenced by neither the TRPA1 antagonist HC-030031 nor the DAT antagonist GBR12909. Subcutaneous administration of VBJ103 (3, 10 and 30 mg·kg -1 , but not 100 or 300 mg·kg -1 , n = 7) decreased CBT (2°C). Intraperitoneal (i.p.) administration of VBJ103 (3, 10 and 30 mg·kg -1 ) dose-dependently decreased CBT to an extent larger than that detected with subcutaneous administration. Intracerebroventricular (i.c.v.) administration (306 nmol/1 μL; n = 5) did not alter CBT., Conclusions and Implications: We achieve therapeutic efficacy with subcutaneous administration of a novel TRPM8 antagonist that attenuates deleterious influences on CBT, a side effect that has largely prevented the translation of TRPM8 as a target., (© 2024 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Background and Purpose: The chemotherapy agent oxaliplatin can give rise to oxaliplatin-induced peripheral neuropathy (OIPN). Here, we investigated whether T-type calcium channels (Ca v 3) contribute to OIPN., Experimental Approach: We chronically treated mice with oxaliplatin and assessed pain responses and changes in expression of Ca v 3.2 calcium channels. We also tested the effects of T-type channel blockers on cold sensitivity in wild-type and Ca v 3.2 null mice., Key Results: Oxaliplatin treatment led to mechanical and cold hypersensitivity in male and female mice. Mechanical hypersensitivity persisted in Ca v 3.2 null mice of both sexes. Intraperitoneal or intrathecal delivery of pan T-type channel inhibitors attenuated mechanical hypersensitivity in wild-type but not Ca v 3.2 null mice. Remarkably cold hypersensitivity occurred in female but not male Ca v 3.2 null mice even without oxaliplatin treatment. Unexpectedly, intrathecal, intraplantar or intraperitoneal delivery of T-type channel inhibitors Z944 or TTA-P2 transiently induced cold hypersensitivity in both male and female wild-type mice. Acute knockdown of specific Ca v 3 isoforms revealed that the depletion of Ca v 3.1 in males and depletion of either Ca v 3.1 or Ca v 3.2 in females triggered cold hypersensitivity. Finally, reducing Ca v 3.2 expression by disrupting the interactions between Ca v 3.2 and the deubiquitinase USP5 with the small organic molecule II-2 reversed oxaliplatin-induced mechanical and cold hypersensitivity and importantly did not trigger cold allodynia., Conclusion and Implications: Altogether, our data indicate that T-type channels differentially contribute to the regulation of cold and mechanical hypersensitivity, and raise the possibility that T-type channel blockers could promote cold allodynia., (© 2024 The Author(s). British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Background: Social interactions with subjects experiencing pain can increase nociceptive sensitivity in observers, even without direct physical contact. In previous experiments, extended indirect exposure to soiled bedding from mice with alcohol withdrawal-related hyperalgesia enhanced nociception in their conspecifics. This finding suggested that olfactory cues could be sufficient for nociceptive hypersensitivity in otherwise untreated animals (also known as "bystanders")., Aim: The current study addressed this possibility using an inflammation-based hyperalgesia model and long- and short-term exposure paradigms in C57BL/6J mice., Materials & Method: Adult male and female mice received intraplantar injection of complete Freund's adjuvant (CFA) and were used as stimulus animals to otherwise naïve same-sex bystander mice (BS). Another group of untreated mice (OLF) was simultaneously exposed to the bedding of the stimulus mice., Results: In the long-term, 15-day exposure paradigm, the presence of CFA mice or their bedding resulted in reduced von Frey threshold but not Hargreaves paw withdrawal latency in BS or OLF mice. In the short-term paradigm, 1-hr interaction with CFA conspecifics or 1-hr exposure to their bedding induced mechanical hypersensitivity in BS and OLF mice lasting for 3 hrs. Chemical ablation of the main olfactory epithelium prevented bedding-induced and stimulus mice-induced mechanical hypersensitivity. Gas chromatography-mass spectrometry (GC-MS) analysis of the volatile compounds in the bedding of experimental mice revealed that CFA-treated mice released an increased number of compounds indicative of disease states., Discussion and Conclusion: These results demonstrate that CFA-induced inflammatory pain can modulate nociception in bystander mice via an olfactory mechanism involving dynamic changes in volatile compounds detectable in the rodent bedding., Significance: Social context can influence nociceptive sensitivity. Recent studies suggested involvement of olfaction in this influence. In agreement with this idea, the present study shows that the presence of mice with inflammatory pain produces nociceptive hypersensitivity in nearby conspecifics. This enhanced nociception occurs via olfactory cues present in the mouse bedding. Analysis of the bedding from mice with inflammatory pain identifies a number of compounds indicative of disease states. These findings demonstrate the importance of olfactory system in influencing pain states., (© 2023 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC ®.)

Objective: The aim of this study was to test whether a combination of sumatriptan with dual enkephalinase inhibitor PL37 would result in an additive or a synergistic effect., Background: Combination treatment is frequently used to improve the therapeutic efficacy of drugs. The co-administration of two drugs may result in efficacy at lower doses than those needed for either drug alone, thus minimizing side effects. Here, we tested the effect of the co-administration of two drugs on cutaneous mechanical hypersensitivity (MH), a symptom often affecting cephalic regions in patients with migraine: dual enkephalinase inhibitor PL37, a small molecule that protects enkephalins from rapid degradation, and sumatriptan, a serotonin 5-HT 1B/1D receptor agonist., Methods: We investigated the effects of oral administrations of sumatriptan, PL37, or their combination on changes in cutaneous mechanical sensitivity induced by a single intraperitoneal administration of the nitric oxide donor, isosorbide dinitrate (ISDN) in male rats. Mechanical sensitivity was assessed using von Frey filaments applied to the face of animals to determine pain thresholds. Isobolographic analysis was performed to determine the nature of the interaction between sumatriptan and PL37., Results: Sumatriptan as well as PL37 each produced a dose-dependent inhibition of ISDN-induced cephalic MH. Median effective dose (ED 50 ) values were 0.3 and 1.1 mg/kg for sumatriptan and PL37, respectively. An isobolographic analysis of the effect of combined doses of sumatriptan and PL37 based on their calculated ED 50 values demonstrated a synergistic effect of the combination on cephalic MH, with an interaction index of 0.14 ± 0.04., Conclusion: These results suggest that PL37 acts synergistically with sumatriptan to produce an anti-allodynic effect in a rat model of migraine. Thus, combining PL37 and sumatriptan may be a useful therapeutic strategy in the management of migraine., Plain Language Summary: There have been many advances in migraine treatment, but we still need more options that are effective and have few side effects. Sumatriptan is one available drug for acute treatment of migraine, but it does not work for every patient and is not suitable for some people. We tested a new drug called PL37 (that blocks enkephalinases) together with sumatriptan and the combination minimized side effects and allowed lower doses of the drugs for effective migraine treatment in an animal model., (© 2024 American Headache Society.)

Background: The compound NS5806 attenuates neuropathic pain via inhibiting extracellular signal-regulated kinase (ERK) activation in neuronal somata located at the dorsal root ganglion (DRG) and superficial spinal dorsal horn. NS5806 also reduces the expansion of DRG macrophages and spinal microglia several days after peripheral nerve injury, implying an anti-inflammatory effect., Methods: To test whether NS5806 inhibits inflammation, as a model we intraplantarly injected carrageenan into a hind paw of the rat. To examine whether NS5806 reduces carrageenan-evoked mechanical allodynia, thermal hyperalgesia, and edema, as well as ERK activation in the nerve fibres, mast cells, and macrophages in the hind paw skin, we used behavioural, immunohistochemical, and cytological methods., Results: NS5806 did not impair motor function, affect basal nociception, or cause edema in naive rats. Six hours after carrageenan injection, mechanical allodynia, thermal hyperalgesia, and edema appeared in the rat's ipsilateral hind paw, and all were reduced by intraplantar co-injection of NS5806. NS5806 suppressed carrageenan-evoked ERK activation in the peripheral axons and somata of L4 DRG neurons, as well as mast cells and macrophages in the paw skin. NS5806 also reduced carrageenan-evoked mast cell degranulation and macrophage proliferation. NS5806 and the ERK pathway inhibitor PD98059 had a similar effect in inhibiting the proliferation of cultured RAW264.7 macrophages. Furthermore, all the in vivo anti-inflammatory effects of NS5806 were similar to those of PD98059., Conclusions: Acting like an ERK pathway inhibitor, NS5806 reduces inflammation-evoked mechanical allodynia, thermal hyperalgesia, and edema by suppressing ERK activation in primary sensory neurons, mast cells, and macrophages., Significance: Previous studies show that NS5806 only acts on neurons. This report unveils that NS5806 also acts on immune cells in the skin to exert its anti-inflammatory effects. Since NS5806 is lipid soluble for skin penetration, it suggests that NS5806 could also be developed into an anti-inflammatory drug for external use., (© 2023 European Pain Federation - EFIC ®.)

Objective: To decipher the underlying mechanisms of nitroglycerin (NTG)-induced migraine electrophysiologically., Background: Migraine is a recurrent primary headache disorder with moderate to severe disability; however, the pathophysiology is not fully understood. Consequently, safe and effective therapies to alleviate migraine headaches are limited. Local field potential (LFP) recording, as a neurophysiological tool, has been widely utilized to investigate combined neuronal activity., Methods: We recorded LFP changes simultaneously from the anterior cingulate cortex, posterior nucleus of the thalamus, trigeminal ganglion, and primary visual cortex after NTG injection in both anesthetized and freely moving rats. Additionally, brain coherence was processed, and light-aversive behavior measurements were implemented., Results: Significant elevations of LFP powers with various response patterns for the delta, theta, alpha, beta, and gamma bands following NTG injection were detected in both anesthetized and freely moving rats; however, a surge of coherence alternations was exclusively observed in freely moving rats after NTG injection., Conclusion: The multi-region LFP signatures and brain coherence alternations in response to NTG-induced migraine attacks were determined. Furthermore, the results of behavior measurements in the freely moving group indicated that NTG induced the phenomenon of photophobia in our study. All these findings offer novel insights into the interpretation of migraine mechanisms and related treatments., (© 2023 American Headache Society.)

Background: The exact mechanism and site of action of triptans in aborting migraine attacks remain under debate. We hypothesized that the clinical efficacy of triptans lies in aborting central sensitization and focused on the question of why triptans are headache specific, that is highly effective in migraine and cluster headache and ineffective in extracephalic pain., Methods: Forty healthy participants were enrolled in this double-blinded, randomized, placebo-controlled study. The effect of sumatriptan (n = 20) versus placebo (n = 20) was investigated in a trigeminal (V1) versus an extracephalic dermatome (forearm) using a topical capsaicin sensitization model. Capsaicin-induced primary and secondary hyperalgesia were evaluated using quantitative sensory testing., Results: After capsaicin application, primary hyperalgesia developed in both the sumatriptan and placebo groups in both dermatomes. However, sumatriptan exclusively prevented secondary hyperalgesia in the V1 dermatome but not on the forearm. Placebo exerted no effects on secondary hyperalgesia in both trigeminal and extracephalic dermatomes. Additionally, sumatriptan reduced the flare size exclusively in the V1 dermatome., Conclusions: Our data suggest that sumatriptan reduces central sensitization (secondary hyperalgesia) without modulating peripheral sensitization (primary hyperalgesia) in a human pain model of capsaicin-induced sensitization. Moreover, despite a systemic administration of sumatriptan, the modulatory effects are trigeminal specific, echoing the clinical effect of triptans in aborting headaches, but not extracephalic pain., Significance: Our data suggest that triptans exert their efficacy by suppressing central sensitization. By revealing a dermatome-specific modulation, our study demonstrates a previously unrecognized interaction between the pharmacodynamics of triptans and the trigeminal nociceptive system that provides new insight into how triptans may work in aborting headache attacks., (© 2022 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC ®.)

Background: Bladder pain syndrome/interstitial cystitis (BPS/IC) is a refractory disease accompanied by bladder-related pain and hyperactivity. Studies have shown that the translocator protein (TSPO) modulates neuroinflammation and central sensitisation associated with pain. Moreover, we previously demonstrated that brain-derived neurotrophic factor (BDNF) regulates neuroinflammation and mechanical allodynia in cyclophosphamide (CYP)-induced cystitis through activation of glial cells. Here, we aimed to explore whether activation of TSPO attenuates mechanical allodynia and bladder dysfunction by regulating BDNF induced neuroinflammation in a CYP-induced cystitis model., Methods: Injection of CYP was performed to form a rat model of BPS/IC. The expression of TSPO was regulated by intrathecal injection of the TSPO agonist Ro5-4864. The von Frey filament test was applied to evaluate suprapubic allodynia. Bladder function was assessed using filling cystometry. Western blotting was used to detect the expression of TSPO, BDNF, GFAP, Iba-1, p-p38, p-JNK, TNF-α, and IL-1β, and double immunofluorescence was performed to localise TSPO in the L6-S1 spinal dorsal horn (SDH)., Results: TSPO was activated in the SDH after CYP injection and was primarily colocalised with astrocytes. Ro5-4864 reversed mechanical allodynia and bladder dysfunction induced by CYP. Moreover, the upregulation of BDNF and activation of astrocytes and microglia was suppressed by Ro5-4864, resulting in downregulation of p-p38, p-JNK, TNF-α, and IL-1β., Conclusions: Ro5-4864 alleviated mechanical allodynia and bladder dysfunction in the CYP model, possibly by inhibiting the elevation of BDNF and consequent activation of astrocytes and microglia induced neuroinflammation. TSPO may be a potential target for the treatment of BPS/IC., Significance: This study examined the mechanism underlying the ability of the translocator protein to modulate bladder pain syndrome/interstitial cystitis., (© 2022 European Pain Federation - EFIC®.)

Objective: The aim of this study was to determine if prolactin signaling modulates stress-induced behavioral responses in a preclinical migraine model., Background: Migraine is one of the most complex and prevalent disorders. The involvement of sex-selective hormones in migraine pathology is highly likely as migraine is more common in women and its frequency correlates with reproductive stages. Prolactin has been shown to be a worsening factor for migraine. Normally prolactin levels are low; however levels can surge during stress. Dopamine receptor agonists, which suppress pituitary prolactin release, are an effective migraine treatment in a subset of patients. Previously, we showed that administration of prolactin onto the dura mater induces female-specific behavioral responses, suggesting that prolactin may play a sex-specific role in migraine., Methods: The effects of prolactin signaling were assessed using a preclinical migraine model we published recently in which behavioral sensitization is induced by repeated stress. Plasma prolactin levels were assessed in naïve and stressed CD-1 mice (n = 3-5/group) and transgenic mice with conditional deletion of the Prlr in Nav1.8-positive sensory neurons (Prlr conditional knock-out [CKO]; n = 3/group). To assess the contribution of prolactin release during stress, naïve or stressed male and female CD-1 mice were treated with the prolactin release inhibitor bromocriptine (2 mg/kg; n = 7-12/group) or vehicle for 5 days (8-12/group) and tested for facial hypersensitivity following stress. Additionally, the contribution of ovarian hormones in regulating the prolactin-induced responses was assessed in ovariectomized female CD-1 mice (n = 6-10/group). Furthermore, the contribution of Prlr activation on Nav1.8-positive sensory neurons was assessed. Naïve or stressed male and female Prlr CKO mice and their control littermates were tested for facial hypersensitivity (n = 8-9/group). Immunohistochemistry was used to confirm loss of Prlr in Nav1.8-positive neurons in Prlr CKO mice. The total sample size is n = 245; the full analysis sample size is n = 221., Results: Stress significantly increased prolactin levels in vehicle-treated female mice (39.70 ± 2.77; p < 0.0001). Bromocriptine significantly reduced serum prolactin levels in stressed female mice compared to vehicle-treated mice (-44.85 ± 3.1; p < 0.0001). Additionally, no difference was detected between female stressed mice that received bromocriptine compared to naïve mice treated with bromocriptine (-0.70 ± 2.9; p = 0.995). Stress also significantly increased serum prolactin levels in male mice, although to a much smaller extent than in females (0.61 ± 0.08; p < 0.001). Bromocriptine significantly reduced serum prolactin levels in stressed males compared to those treated with vehicle (-0.49 ± 0.08; p = 0.002). Furthermore, bromocriptine attenuated stress-induced behavioral responses in female mice compared to those treated with vehicle (maximum effect observed on day 4 post stress [0.21 ± 0.08; p = 0.03]). Bromocriptine did not attenuate stress-induced behavior in males at any timepoint compared to those treated with vehicle. Moreover, loss of ovarian hormones did not affect the ability of bromocriptine to attenuate stress responses compared to vehicle-treated ovariectomy mice that were stressed (maximum effect observed on day 4 post stress [0.29 ± 0.078; p = 0.013]). Similar to CD-1 mice, stress increased serum prolactin levels in both Prlr CKO female mice (27.74 ± 9.96; p = 0.047) and control littermates (28.68 ± 9.9; p = 0.041) compared to their naïve counterparts. There was no significant increase in serum prolactin levels detected in male Prlr CKO mice or control littermates. Finally, conditional deletion of Prlr from Nav1.8-positive sensory neurons led to a female-specific attenuation of stress-induced behavioral responses (maximum effect observed on day 7 post stress [0.32 ± 0.08; p = 0.007]) compared to control littermates., Conclusion: These data demonstrate that prolactin plays a female-specific role in stress-induced behavioral responses in this preclinical migraine model through activation of Prlr on sensory neurons. They also support a role for prolactin in migraine mechanisms in females and suggest that modulation of prolactin signaling may be an effective therapeutic strategy in some cases., (© 2021 American Headache Society.)

Background and Purpose: P2Y 12 receptors regulate different forms of pain and inflammation. In this study, we investigated the participation of P2Y 12 receptors in an animal model of migraine., Experimental Approach: We tested the effect of the centrally administered selective P2Y 12 antagonist PSB-0739 and P2Y 12 receptor gene (P2ry12 -/- ) deficiency in acute nitroglycerin-treated mice. Additionally, platelet depletion was used to investigate the role of platelet P2Y 12 receptors during migraine-like pain., Key Results: Nitroglycerin induced sensory hypersensitivity of C57BL/6 wild-type (P2ry12 +/+ ) mice accompanied by an increase in c-fos and CGRP expression in the upper cervical spinal cord (C1-C2) and trigeminal nucleus caudalis. Similar changes were also observed in P2Y 12 gene-deficient (P2ry12 -/- ) mice. Prophylactic intrathecal application of PSB-0739 reversed thermal hyperalgesia and head grooming time in wild-type mice but had no effect in P2ry12 -/- mice. Furthermore, PSB-0739 was also effective when applied as a post-treatment. PSB-0739 administration suppressed the expression of c-fos in C1-C2 and trigeminal nucleus caudalis, and decreased the levels of dopamine and 5-hydroxytryptamine in C1-C2 in wild-type mice. Nitroglycerin treatment itself did not change adenosine diphosphate (ADP)-induced platelet activation measured by CD62P up-regulation in wild-type mice. Platelet depletion by anti-mouse CD41 antibody and clopidogrel attenuated nitroglycerin-induced thermal hypersensitivity and head grooming time in mice., Conclusion and Implications: Our findings show that acute inhibition of P2Y 12 receptors alleviates migraine-like pain in mice by modulating the expression of c-fos and that platelet P2Y 12 receptors might contribute to this effect. Thus the blockade of P2Y 12 receptors may have therapeutic potential against migraine., (© 2021 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

The present study aimed to determine the antioxidant effect of Citrus unshiu Markovich (CUM) extract in neuronal cell lines under oxidative stress and to investigate the effect of chemotherapy-induced peripheral neuropathy (CIPN) on the nociceptive response in a preclinical mice model. We tested the inhibition of H 2 O 2 in Neuro2A cells treated with CUM. Experimental animals were treated with oxaliplatin to induce CINP, and then administered oral CUM for 4 weeks in order to observe the effect of CUM. Animals were evaluated weekly for thermal hyperalgesia and digital motor nerve conduction velocity (NCV). Lumbar dorsal root ganglia (DRG) isolated from each animal were evaluated through immunochemical and western blot analysis for nerve damage, inflammatory response, and expression of redox signaling factors. The main mechanisms were determined to be decreased inducible nitric oxide synthase (iNOS) production due to the inhibition of NADPH oxidase 2 (NOX2). To determine the functional role of NOX2 in CINP, we administrated CUM into NOX2-deficient mice with neuropathic pain. Therefore, we suggest that CUM controls the expression levels of inflammatory factors in CINP via NOX2 inactivation. This study demonstrated that a complementary medicine such as CUM might be a potential novel therapeutic agent for the treatment of CINP., (© 2021 John Wiley & Sons Ltd.)

Background: Mechanical hyperalgesia and allodynia incidence varies considerably amongst neuropathic pain patients. This study explored whether sensory or psychological factors associate with mechanical hyperalgesia and brush allodynia in a human experimental model., Methods: Sixty-six healthy volunteers (29 male) completed psychological questionnaires and participated in two quantitative sensory testing (QST) sessions. Warmth detection threshold (WDT), heat pain threshold (HPT) and suprathreshold mechanical pain (STMP) ratings were measured before exposure to a capsaicin-heat pain model (C-HP). After C-HP exposure, brush allodynia and STMP were measured in one session, whilst mechanical hyperalgesia was measured in another session., Results: WDT and HPT measured in sessions separated by 1 month demonstrated significant but moderate levels of reliability (WDT: ICC = 0.5, 95%CI [0.28, 0.77]; HPT: ICC = 0.62, 95%CI [0.40, 0.77]). Brush allodynia associated with lower WDT (z = -3.06, p = 0.002; ϕ = 0.27). Those with allodynia showed greater hyperalgesia intensity (F = 7.044, p = 0.010, η p 2  = 0.107) and area (F = 9.319, p = 0.004, η p 2  = 0.163) than those without allodynia. No psychological self-report measures were significantly different between allodynic and nonallodynic groups. Intensity of hyperalgesia in response to lighter mechanical stimuli was associated with lower HPT, higher STMP ratings and higher Pain Sensitivity Questionnaire scores at baseline. Hyperalgesia to heavier probe stimuli associated with state anxiety and to a lesser extent somatic awareness. Hyperalgesic area associated with lower baseline HPT and higher STMP ratings. Hyperalgesic area was not correlated with allodynic area across individuals., Conclusions: These findings support research in neuropathic pain patients and human experimental models that peripheral sensory input and individual sensibility are related to development of mechanical allodynia and hyperalgesia during central sensitization, whilst psychological factors play a lesser role., Significance: We evaluated differential relationships of psychological and perceptual sensitivity to the development of capsaicin-induced mechanical allodynia and hyperalgesia. Fifty percent of healthy volunteers failed to develop mechanical allodynia. Baseline pain sensitivity was greater in those developing allodynia and was related to the magnitude and area of hyperalgesia. State psychological factors, whilst unrelated to allodynia, were related to mechanical hyperalgesia. This supports that the intensity of peripheral sensory input and individual sensibility are related to development of mechanical allodynia and hyperalgesia during central sensitization, whilst psychological factors play a lesser role., (© 2021 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC ®.)

Objective: The aim of this study was to determine if the non-convulsant delta-opioid receptor (DOR) agonist, KNT-127, could inhibit migraine-associated endpoints., Background: The DOR has been identified as a therapeutic target for migraine. However, the development of delta agonists is limited as some ligands have seizurogenic properties, which may be related to their ability to induce receptor internalization. While both pro- and non-convulsant delta agonists can reduce migraine-associated allodynia, only the proconvulsant agonist, SNC80, has been shown to decrease cortical spreading depression (CSD). It is unclear if the ability of delta agonists to modulate cortical activity is related to the same signaling mechanisms that produce proconvulsant effects., Methods: The effects of the non-convulsant delta agonist, KNT-127, were examined. Repetitive CSD was induced in female C57BL6/J (n = 6/group) mice by continuous application of KCl and the effect of KNT-127/vehicle (Veh) on both local field potentials and optical intrinsic signals was determined. To assess the effect of KNT-127 on established chronic migraine-associated pain, male and female C57BL6/J mice were treated with nitroglycerin (NTG; 10 mg/kg, ip) every other day for 9 days and tested with KNT-127 (5 mg/kg, sc) or Veh on day 10 (n = 6/group). DOR-enhanced green fluorescent protein mice (n = 4/group) were used to confirm the internalization properties of KNT-127 in the trigeminal ganglia, trigeminal nucleus caudalis, and somatosensory cortex., Results: KNT-127 inhibited CSD events (t (10)  = 3.570, p = 0.0051). In addition, this delta agonist also reversed established cephalic allodynia in the NTG model of chronic migraine (F (1, 20)  = 12.80, p < 0.01). Furthermore, KNT-127 caused limited internalization of DOR in key migraine processing regions., Conclusions: This study shows that the antimigraine effects of DOR agonists can be separated from their proconvulsant effects. This data provides valuable information for the continued development of delta agonists for the treatment of migraine., (© 2020 American Headache Society.)

Background: Unilateral injection of Complete Freund's Adjuvant (CFA) into the intra-plantar surface of the rodent hindpaw elicits chronic inflammation and hyperalgesia in the ipsilateral hindlimb. Mechanisms contributing to this hyperalgesia may act over multiple time courses and can include changes in ion channel expression and post-translational SUMOylation. Hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels mediate the hyperpolarization-activated current, I h . An HCN2-mediated increase in C-nociceptor I h contributes to mechanical hyperalgesia in the CFA model of inflammatory pain. Changes in HCN2 post-translational SUMOylation and protein expression have not been systematically documented for a given dorsal root ganglia (DRG) throughout the time course of inflammation., Methods: This study examined HCN2 protein expression and post-translational SUMOylation in a rat model of CFA-induced hindpaw inflammation. L5 DRG cryosections were used in immunohistochemistry experiments and proximity ligation assays to investigate HCN2 expression and SUMOylation, respectively, on days 1 and 3 post-CFA., Results: Unilateral CFA injection elicited a significant bilateral increase in HCN2 staining intensity in small diameter DRG neurons on day 1 post-CFA, and a significant bilateral increase in the number of small neurons expressing HCN2 but not staining intensity on day 3 post-CFA. HCN2 channels were hyper-SUMOylated in small diameter neurons of ipsilateral relative to contralateral DRG on days 1 and 3 post-CFA., Conclusions: Unilateral CFA injection elicits unilateral mechanical hyperalgesia, a bilateral increase in HCN2 expression and a unilateral increase in post-translational SUMOylation. This suggests that enhanced HCN2 expression in L5 DRG is not sufficient for mechanical hyperalgesia in the early stages of inflammation and that hyper-SUMOylation of HCN2 channels may also be necessary., Significance: Nociceptor HCN2 channels mediate an increase in I h that is necessary for mechanical hyperalgesia in a CFA model of chronic pain, but the mechanisms producing the increase in nociceptor I h have not been resolved. The data presented here suggest that the increase in I h during the early stages of inflammation may be mediated by an increase in HCN2 protein expression and post-translational SUMOylation., (© 2020 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC®.)

Background: Anodal transcranial direct current stimulation over the primary cortex has been shown to activate regions of the brain involved in the descending modulation of pain sensitivity. However, more research is required to dissect the spinal cord analgesic mechanisms associated with the development of central sensitization., Methods: In this randomized, double blind, crossover study 12 healthy participants had baseline mechanical stimulus response (S/R) functions measured before and after the development of capsaicin-induced ongoing pain sensitivity. The effects of 20 min of either real or sham transcranial direct current stimulation (tDCS, 2 mA) over the primary motor cortex on dynamic mechanical allodynia (DMA) and mechanical pain sensitivity (MPS) were then investigated., Results: Topical application of capsaicin resulted in an increase in area under the pain ratings curve for both DMA (p < .01) and MPS (p < .01). The effects of tDCS on the area under the curve ratio (i.e. post-/pre-treatment) revealed significant analgesic effects over DMA (p < .05) and MPS (p < .05) when compared with sham., Conclusions: This study demonstrates that anodal tDCS over the primary motor cortex can reduce both dynamic and static forms of mechanical pain sensitivity associated with the development of DMA and MPS, respectively. The use of tDCS may provide a novel mechanism-driven therapy in chronic pain patients with altered mechanical S/R functions., Significance: This research shows new evidence that anodal tDCS over the primary motor cortex can reduce dynamic and static forms of mechanical pain sensitivity in the capsaicin model of ongoing pain. By using this approach, it may be possible to provide mechanism-driven analgesia in chronic pain patients who have dynamic mechanical allodynia and/or secondary mechanical hyperalgesia., (© 2020 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC ®.)

The antihyperalgesic and sedative effects of the α2-subunit preferring GABA A positive allosteric modulator (GAM), N-desmethyl-clobazam (NDMC), 20 and 60 mg, were assessed in a randomized, placebo and active-controlled (clonazepam 1,5 mg), 4-way crossover study, in healthy volunteers, using the ultraviolet B-induced experimental pain model. Single (20, 40, 60 mg) and repeated doses (20 mg over 15 days) of NDMC pharmacokinetics were evaluated. Thirty-two subjects participated in the study. Primary outcome parameter was maximal change in the area of cutaneous UVB irradiation-induced secondary hyperalgesia (ASH). ASH decreased under all treatments. Mean (SD) relative change was 79 (22)%, 83 (24)%, 77 (30)% and 92 (16)% for placebo, NDMC20, NDMC60 and clonazepam, respectively. Neither absolute change nor relative change in ASH was significantly different between NDMC60 and placebo (mean difference = 2.3 cm 2 [95% CI 4.0-8.5], p = .462 and 0.4% [-11.9 to 12.6], p = .952, respectively). An overall treatment effect was found on level of sedation. Compared to placebo, sedation was higher under clonazepam (mean difference = 39 mm [30-49] on a visual analogue scale, p < .001) while NDMC was free of sedative effect. NDMC pharmacokinetics after single doses showed poor absorption, but was linear. Steady-state plasma concentrations of NDMC20 were attained within 14 days, with low between-subjects variability. Mean steady-state concentration (C S-S , SD) reached 209 (22) ng/ml. NDMC absence of sedative effect and its overall well-characterized safety coming from years of utilization as a metabolite from clobazam, raise the prospect of dose escalating trials in patients to quantify its clinical utility. SIGNIFICANCE: This article, presenting the Phase I data of the new antihyperalgesic compound, α2-subunit GABA A positive allosteric modulator, N-desmethyl-clobazam (NDMC) is exploring the modulation of a new target in the treatment of neuropathic pain. Based on these results and on its preclinical properties NDMC would qualify as a good tool compound to seek confirmation of the clinical utility of selective GABA allosteric modulators in neuropathic pain patients., (© 2020 European Pain Federation - EFIC®.)

Background: Hyperalgesia is a heightened pain response to a noxious stimulus and is a hallmark of many common neuropathic and chronic pain conditions. In a double-blind placebo-controlled drug-crossover trial, the effects of concomitant and delayed minocycline treatment on the initiation and resolution of muscle hyperalgesia were tested., Methods: An initial cohort (n = 10) received repeated injections (5 µg: days 0, 2 and 4) of nerve growth factor (NGF) in the flexor carpi ulnaris muscle of the forearm and pressure pain thresholds were collected at day 0 (control), day 7 (peak) and day 14 (recovery). A second cohort (n = 18) underwent an identical procedure, however, half received a placebo between days 0 and 7 before switching to minocycline from days 7 to 14 (P1/M2), while the remaining subjects received minocycline (day 0: 200mg then 100mg b.i.d. for 7 days) before switching to placebo (M1/P2)., Results: The initial cohort exhibited a diffuse muscular pain hypersensitivity with a decrease in pressure pain thresholds at day 7 before a partial return to normalcy at day 14. The P1/M2 treatment group exhibited an identical peak in hypersensitivity at day 7, however, after switching to minocycline in week 2 showed a significant reduction in muscle hyperalgesia compared with the initial cohort at day 14. The M1/P2 treatment group had significantly less (~43%) hyperalgesia at day 7 compared with the other groups., Conclusions: The study indicates that the administration of minocycline can reduce experimentally induced muscle pain regardless of the time of administration., Significance: In a double-blind placebo-controlled drug-crossover study, the common antibiotic minocycline was found to reduce the muscle hyperalgesia induced by intramuscular injection of nerve growth factor. The results of the study showed that both concomitant (pre-emptive) and delayed administration of minocycline can ameliorate the onset and facilitate the resolution of experimentally induced muscle hyperalgesia., (© 2020 European Pain Federation - EFIC®.)

Background: Intramuscular injection of Nerve Growth Factor (NGF) may influence the responsiveness of active chemo-sensitive channels affecting muscle pain sensitivity. This double-blinded crossover study in healthy humans assessed contraction-evoked pain responses and pain sensitivity during acute ischaemia in the tibialis anterior (TA) muscle before and 24 hr after five distributed NGF injections (1 µg, 4 cm interval) compared with control injections (isotonic-saline)., Methods: Twenty-one subjects participated in two experimental phases, each including five sessions over 7 days, with a gap of 4 weeks in-between. Muscle pain intensity evoked with daily functional tasks (Likert scale score) was collected using a paper diary. Pain intensity evoked by ischaemic and non-ischaemic contractions numerical rating scale (NRS) was collected at Day0 and Day1. Pressure pain thresholds (PPTs) on the TA were recorded before (Day0), 3 hr, 1, 3, and 7 days post-injection, and after the ischaemic-contractions and post-cuff deflation at Day0 and Day1., Results: Increased Likert scores of pain were present for 7 days after NGF compared to control injections (p < .05). Higher NRS pain scores of ischaemic-contractions were seen when contracting the muscle injected with NGF compared to baseline (p = .003) and control (p = .012). Pain during non-ischaemic contractions was not significantly affected by NGF injections. Decreased PPTs were found at 3 hr, Day1 and Day3 post-injection (p < .05) in both conditions. Compared with pre-contractions, PPTs were increased following ischaemic contractions at Day0 (p < .05) and Day1 (p < .05) in both conditions., Conclusion: This study showed that ischaemic contraction-evoked pain was facilitated in an NGF-sensitized muscle., Significance: Acidification of the muscle environment may affect muscle nociceptors and pain by different mechanisms, including activation of ASIC 3 and TRPV1. In this study, pain evoked following ischaemic contractions was increased in the Nerve Growth Factor (NGF)-sensitized muscle compared with non-ischaemic contractions and in the non-sensitized muscle. These findings illustrate that responses of peripheral afferents under ischaemic conditions are altered by a pre-sensitized muscle. This highlights the role of growth factors, including NGF, in peripheral muscle sensitization with clinical implications for ischaemic myalgia., (© 2019 European Pain Federation - EFIC®.)

Background and Purpose: The P2X3 receptor is an ATP-gated ion channel expressed by sensory afferent neurons and is used as a target to treat chronic sensitisation conditions. The first-in-class, selective P2X3 and P2X2/3 receptor antagonist, the diaminopyrimidine MK-7264 (gefapixant), has progressed to Phase III trials for refractory or unexplained chronic cough. We used patch clamp to elucidate the pharmacology and kinetics of MK-7264 and rat models of hypersensitivity and hyperalgesia to test its efficacy on these conditions., Experimental Approach: Whole-cell patch clamp of 1321N1 cells expressing human P2X3 and P2X2/3 receptors was used to determine mode of MK-7264 action, potency, and kinetics. The analgesic efficacy was assessed using paw withdrawal threshold and limb weight distribution in rat models of inflammatory, osteoarthritic, and neuropathic sensitisation., Key Results: MK-7264 is a reversible allosteric antagonist at human P2X3 and P2X2/3 receptors. Experiments with the slowly desensitising P2X2/3 heteromer revealed concentration- and state-dependency to wash-on, with faster rates and greater inhibition when applied before agonist compared to during agonist application. The wash-on rate (τ value) for MK-7264 at maximal concentrations was much lower when applied before compared to during agonist application. In vivo, MK-7264 displayed efficacy comparable to naproxen in inflammatory and osteoarthritic sensitisation models and gabapentin in neuropathic sensitisation models, increasing paw withdrawal threshold and decreasing weight-bearing discomfort., Conclusions and Implications: MK-7264 is a reversible and selective P2X3 and P2X2/3 antagonist, exerting allosteric antagonism via preferential activity at closed channels. Its efficacy in rat models supports its clinical investigation for chronic sensitisation conditions., (© 2019 The British Pharmacological Society.)

Chemotherapy-induced peripheral neuropathy is a disabling condition induced by several frequently used chemotherapeutic drugs including the front-line agent oxaliplatin (OXA). Symptoms are predominantly sensory with the development of neuropathic pain. Alternative dosing protocols and treatment discontinuation are the only available therapeutic strategies. The aim of our work was to evaluate the potential of a synthetic derivative of progesterone, 17α-hydroxyprogesterone caproate (HPGC), in the prevention and treatment of OXA-evoked painful neuropathy. We also evaluated glial activation at the dorsal root ganglia (DRG) and spinal cord levels as a possible target mechanism underlying HPGC actions. Male rats were injected with OXA and HPGC following a prophylactic (HPGCp) or therapeutic (HPGCt) scheme (starting either before or after chemotherapy). The development of hypersensitivity and allodynic pain and the expression of neuronal and glial activation markers were evaluated. When compared to control animals, those receiving OXA showed a significant decrease in paw mechanical and thermal thresholds, with the development of allodynia. Animals treated with HPGCp showed patterns of response similar to those detected in control animals, while those treated with HPGCt showed a suppression of both hypersensitivities after HPGC administration. We also observed a significant increase in the mRNA levels of activating transcription factor 3, the transcription factor (c-fos), glial fibrillary acidic protein, ionized calcium binding adaptor protein 1, interleukin 1 beta (IL-1β) and tumor necrosis factor alpha (TNFα) in DRG and spinal cord of OXA-injected animals, and significantly lower levels in rats receiving OXA and HPGC. These results show that HPGC administration reduces neuronal and glial activation markers and is able to both prevent and suppress OXA-induced allodynia, suggesting a promising therapeutic strategy., (© 2019 Peripheral Nerve Society.)

Background: The antineoplastic drugs cisplatin and vincristine induce peripheral neuropathies. The sigma-1 receptor (σ1R) is expressed in areas of pain control, and its blockade with the novel selective antagonist MR-309 has shown efficacy in nociceptive and neuropathic pain models. Our goal was to test whether this compound reduces neuropathic signs provoked by these antitumoural drugs., Methods: Rats were treated with cisplatin or vincristine to induce neuropathies. The effects of acute or repeated administration of MR-309 were tested on mechanical and thermal sensitivity, electrophysiological activity of Aδ-primary afferents in the rat skin-saphenous nerve preparation, and gastrointestinal or cardiovascular functions., Results: Rats treated with antitumourals developed tactile allodynia, while those treated with vincristine also developed mechanical hyperalgesia. These in vivo modifications correlated with electrophysiological hyperactivity (increased spontaneous activity and hyperresponsiveness to innocuous and noxious mechanical stimulation). Animals treated with cisplatin showed gastrointestinal impairment and those receiving vincristine showed cardiovascular toxicity. A single dose of MR-309 strongly reduced both nociceptive behaviour and electrophysiological changes. Moreover, its concomitant administration with the antitumourals blocked the development of neuropathic symptoms, thus restoring mechanical sensitivity, improving the impairment of feeding behaviour and gastrointestinal transit in the cisplatin-treated group along with ameliorating the altered vascular reactivity recorded in rats treated with vincristine., Conclusion: σ1R antagonist, MR-309, reduces sensorial and electrophysiological neuropathic signs in rats treated with cisplatin or vincristine and, in addition, reduces gastrointestinal and cardiovascular side effects., Significance: σ1R antagonism could be an interesting and new option to palliate antitumoural neuropathies., (© 2018 European Pain Federation - EFIC®.)

The 25-methoxy hispidol A (25-MHA) is a triterpenoid, isolated from the immature fruit of Poncirus trifoliata (Rutaceae). The pretreatment with 25-MHA markedly (p < 0.001) attenuated the formalin-induced biphasic responses as well as acetic acid-induced writhing responses. The intraperitoneal administration of 25-MHA significantly attenuated the mechanical hyperalgesia (p < 0.001) and allodynia (p < 0.05). Similarly, 25-MHA also significantly attenuated (p < 0.001) complete Freund's adjuvant (CFA)-induced paw edema in mice. The 25-MHA treatment significantly attenuated the production of nuclear kappa B (NF-κB) (p65 nuclear subunit). The cytokines are the important mediators of inflammation and pain; however, treatment with 25-MHA exhibited significant inhibition (p < 0.001) on the mRNA expression levels of various inflammatory mediators. The 25-MHA administration also significantly enhanced antioxidant enzymes (p < 0.001) and inhibited the oxidative stress markers. The current study indicates that 25-MHA significantly (p < 0.001) inhibited the nitric oxide (NO) in mice plasma. Similarly, the haematoxylin and eosin (H&E) staining shows that 25-MHA administration significantly inhibited the inflammatory process in the mice paw tissue compared with the CFA-treated group. The 25-MHA treatment did not exhibited any toxicity on the liver, kidney, muscles strength, and motor co-ordination in mice. The 25-MHA was coadministered with the various drugs such as tramadol, piroxicam, and gabapentin to observe the synergistic effect., (© 2018 John Wiley & Sons, Ltd.)

Background: The study aimed to investigate the physiology, psychophysics, pathology and their relationship in reversible nociceptive nerve degeneration, and the physiology of acute hyperalgesia., Methods: We enrolled 15 normal subjects to investigate intraepidermal nerve fibre (IENF) density, contact heat-evoked potential (CHEP) and thermal thresholds during the capsaicin-induced skin nerve degeneration-regeneration; and CHEP and thermal thresholds at capsaicin-induced acute hyperalgesia., Results: After 2-week capsaicin treatment, IENF density of skin was markedly reduced with reduced amplitude and prolonged latency of CHEP, and increased warm and heat pain thresholds. The time courses of skin nerve regeneration and reversal of physiology and psychophysics were different: IENF density was still lower at 10 weeks after capsaicin treatment than that at baseline, whereas CHEP amplitude and warm threshold became normalized within 3 weeks after capsaicin treatment. Although CHEP amplitude and IENF density were best correlated in a multiple linear regression model, a one-phase exponential association model showed better fit than a simple linear one, that is in the regeneration phase, the slope of the regression line between CHEP amplitude and IENF density was steeper in the subgroup with lower IENF densities than in the one with higher IENF densities. During capsaicin-induced hyperalgesia, recordable rate of CHEP to 43 °C heat stimulation was higher with enhanced CHEP amplitude and pain perception compared to baseline., Conclusions: There were differential restoration of IENF density, CHEP and thermal thresholds, and changed CHEP-IENF relationships during skin reinnervation. CHEP can be a physiological signature of acute hyperalgesia., Significance: These observations suggested the relationship between nociceptive nerve terminals and brain responses to thermal stimuli changed during different degree of skin denervation, and CHEP to low-intensity heat stimulus can reflect the physiology of hyperalgesia., (© 2018 European Pain Federation - EFIC®.)

Background: Current medical treatments for chemotherapy-induced pain (CIP) are either ineffective or have adverse side effects. Acupuncture may alleviate CIP, but its effectiveness against this condition has not been studied. Paclitaxel causes neuropathic pain in cancer patients., Methods: We evaluated the effects of electroacupuncture (EA) on paclitaxel-induced CIP in a rat model. Paclitaxel (2 mg/kg) or vehicle was injected (i.p.) on alternate days of 0-6. The resulting pain was treated with 10 Hz/2 mA/0.4 ms pulse EA for 30 min at the equivalent of human acupoint GB30 (Huantiao) once every other day between days 14 and 26. For sham control, EA needles were inserted into GB30 without stimulation. Von Frey filaments with bending forces of 2-8 g and 15 g were used to assess mechanical allodynia and hyperalgesia, respectively, on day 13 and once every other day between 14-26 days and then for 2-3 weeks after EA treatment., Results: Compared to sham control, EA significantly alleviated paclitaxel-induced mechanical allodynia and hyperalgesia, as shown by less frequent withdrawal responses to the filaments. The alleviation of allodynia/hyperalgesia lasted up to 3 weeks after the EA treatment. EA significantly inhibited phosphorylation of Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) in the spinal cord. KN-93, a selective inhibitor of p-CaMKII, inhibited mechanical allodynia/hyperalgesia and p-CaMKII. 5-HT1A receptor antagonist blocked EA inhibition of allodynia/hyperalgesia and p-CaMKII., Conclusions: Electroacupuncture activates 5-HT 1A receptors in the spinal cord and inhibits p-CaMKII to alleviate both allodynia and hyperalgesia. The data support acupuncture/EA as a complementary therapy for CIP., Significance: Electroacupuncture (EA) activates spinal 5-HT1A receptors to inhibit p-CaMKII to alleviate paclitaxel-induced pain. Acupuncture/EA may be used as a complementary therapy for CIP., (© 2017 European Pain Federation - EFIC®.)

Background: Human experimental pain models provide an important translational link between pre-clinical models and clinical pain. Using topical capsaicin and continuous heat application, the novel capsaicin/heat ongoing pain (CHOP) model induces long-lasting experimental pain of which the perceived intensity can be individually adjusted., Methods: In the CHOP model, capsaicin or control cream is applied to a 10 × 10 cm skin area and a heating pad is applied over the area after cream removal. Two experiments in healthy participants were performed for model characterization. In Experiment 1, a constant temperature was applied for 60 min; in Experiment 2, temperature was adjusted to maintain a constant perceived intensity for 60 min., Results: Experiment 1: across participants, constant temperature induced initial habituation followed by an increase in sensation back to baseline. Cluster analysis revealed that half the participants sensitized to the constant temperature, while the other half did not. The degree of sensitization was related to the baseline pain unpleasantness, relative to pain intensity. Experiment 2: constant perceived intensity was achieved in the painful and a non-painful control condition. The two conditions did not differ regarding possibly confounding variables, including blood pressure, heart rate, inflammation or physiological stress as measured by surrogate markers. Secondary allodynia and hyperalgesia were reported more following painful compared to control stimulation. Sensitizers as determined in Experiment 1 were also more pain sensitive in Experiment 2., Conclusion: The CHOP model reproduces some aspects of clinical pain, such as longer duration, sensitization, secondary allodynia and hyperalgesia., Significance: Here we demonstrate a novel pain model that can be applied for up to an hour without tissue damage. The CHOP model allows for investigation of primary and secondary hyperalgesia as well as top-down influences on sensitization, thereby providing an experimental model that can be used to assess clinically-oriented questions., (© 2017 European Pain Federation - EFIC®.)

Background: Histamine H3 receptors are mainly expressed on CNS neurons, particularly along the nociceptive pathways. The potential involvement of these receptors in pain processing has been suggested using H3 receptor inverse agonists., Methods: The antinociceptive effect of S 38093, a novel inverse agonist of H3 receptors, has been evaluated in several neuropathic pain models in rat and compared with those of gabapentin and pregabalin., Results: While S 38093 did not change vocalization thresholds to paw pressure in healthy rats, it exhibited a significant antihyperalgesic effect in the Streptozocin-induced diabetic (STZ) neuropathy model after acute and chronic administration and, in the chronic constriction injury (CCI) model only after chronic administration, submitted to the paw-pressure test. Acute S 38093 administration at all doses tested displayed a significant cold antiallodynic effect in a model of acute or repeated administration of oxaliplatin-induced neuropathy submitted to cold tail immersion, cold allodynia being the main side effect of oxaliplatin in patients. The effect of S 38093 increased following chronic administration (i.e. twice a day during 5 days) in the CCI and STZ models except in the oxaliplatin models where its effect was already maximal from the first administration The kinetics and size of effect of S 38093 were similar to gabapentin and/or pregabalin. Finally, the antinociceptive effect of S 38093 could be partially mediated by α2 adrenoreceptors desensitization in the locus coeruleus., Conclusions: These results highlight the interest of S 38093 to relieve neuropathic pain and warrant clinical trials especially in chemotherapeutic agent-induced neuropathic pain., Significance: S 38093, a new H3 antagonist/inverse agonist, displays antiallodynic and antihyperalgesic effect in neuropathic pain, especially in oxaliplatin-induced neuropathy after chronic administration. This effect of S 38093 in neuropathic pain could be partly mediated by α2 receptors desensitization in the locus coeruleus., (© 2017 European Pain Federation - EFIC®.)

Background and Purpose: The non-psychoactive phytocannabinoid cannabidiol (CBD) can affect the pharmacological effects of Δ 9 -tetrahydrocannabinol (THC). We tested the possible synergy between CBD and THC in decreasing mechanical sensitivity in a mouse model of paclitaxel-induced neuropathic pain. We also tested the effects of CBD on oxaliplatin- and vincristine-induced mechanical sensitivity., Experimental Approach: Paclitaxel-treated mice (8.0 mg·kg -1 i.p., days 1, 3, 5 and 7) were pretreated with CBD (0.625-20.0 mg·kg -1 i.p.), THC (0.625-20.0 mg·kg -1 i.p.) or CBD + THC (0.04 + 0.04-20.0 + 20.0 mg·kg -1 i.p.), and mechanical sensitivity was assessed on days 9, 14 and 21. Oxaliplatin-treated (6.0 mg·kg -1 i.p., day 1) or vincristine-treated mice (0.1 mg·kg -1 i.p. days 1-7) were pretreated with CBD (1.25-10.0 mg·kg -1 i.p.), THC (10.0 mg·kg -1 i.p.) or THC + CBD (0.16 mg·kg -1 THC + 0.16 mg·kg -1 CBD i.p.)., Key Results: Both CBD and THC alone attenuated mechanical allodynia in mice treated with paclitaxel. Very low ineffective doses of CBD and THC were synergistic when given in combination. CBD also attenuated oxaliplatin- but not vincristine-induced mechanical sensitivity, while THC significantly attenuated vincristine- but not oxaliplatin-induced mechanical sensitivity. The low dose combination significantly attenuated oxaliplatin- but not vincristine-induced mechanical sensitivity., Conclusions and Implications: CBD may be potent and effective at preventing the development of chemotherapy-induced peripheral neuropathy, and its clinical use may be enhanced by co-administration of low doses of THC. These treatment strategies would increase the therapeutic window of cannabis-based pharmacotherapies., (© 2017 The British Pharmacological Society.)

Objective: Exercise is commonly recommended for patients with osteoarthritis (OA) pain. However, whether exercise is beneficial in ameliorating ongoing pain that is persistent, resistant to nonsteroidal antiinflammatory drugs (NSAIDs), and associated with advanced OA is unknown., Methods: Rats treated with intraarticular (IA) monosodium iodoacetate (MIA) or saline underwent treadmill exercise or remained sedentary starting 10 days postinjection. Tactile sensory thresholds and weight bearing were assessed, followed by radiography at weekly intervals. After 4 weeks of exercise, ongoing pain was assessed using conditioned place preference (CPP) to IA or rostral ventromedial medulla (RVM)-administered lidocaine. The possible role of endogenous opioids in exercise-induced pain relief was examined by systemic administration of naloxone. Knee joints were collected for micro-computed tomography (micro-CT) analysis to examine pathologic changes to subchondral bone and metaphysis of the tibia., Results: Treadmill exercise for 4 weeks reversed MIA-induced tactile hypersensitivity and weight asymmetry. Both IA and RVM lidocaine D35, administered post-MIA, induced CPP in sedentary but not exercised MIA-treated rats, indicating that exercise blocks MIA-induced ongoing pain. Naloxone reestablished weight asymmetry in MIA-treated rats undergoing exercise and induced conditioned place aversion, indicating that exercise-induced pain relief is dependent on endogenous opioids. Exercise did not alter radiographic evidence of OA. However, micro-CT analysis indicated that exercise did not block lateral subchondral bone loss or trabecular bone loss in the metaphysis, but did block MIA-induced medial bone loss., Conclusion: These findings support the conclusion that exercise induces pain relief in advanced, NSAID-resistant OA, likely through increased endogenous opioid signaling. In addition, treadmill exercise blocked MIA-induced bone loss in this model, indicating a potential bone-stabilizing effect of exercise on the OA joint., (© 2017, American College of Rheumatology.)

Background: Chronic pain patients with sensitization may exhibit decreased sensitivity to normally pruritogenic sensory stimuli and moreover occasionally perceive these as painful. This study explored the relationship between itch and pain, by evaluating histaminergic and non-histaminergic itch evoked in capsaicin-induced allodynic and hyperalgesic areas., Methods: In 28 healthy volunteers, capsaicin (100 μg/0.1 mL) was injected intradermally in the volar forearm to establish secondary dysesthesias. After the capsaicin-induced pain subsided, the areas of allodynia and hyperalgesia were mapped and itch was provoked inside these areas by histamine (10 mg/mL) and cowhage (25-40 spicules). The evoked itch and pain were recorded on a visual analogue scale (VAS 0-10 cm). Contralateral injection of 0.1 mL isotonic saline served as a control., Results: Histaminergic and non-histaminergic evoked itch were significantly decreased when provoked in allodynic skin (p < 0.05). The area-under-the-curve of the evoked itch was reduced -43% from 18.0 ± 2.6 cm 10 min in normal skin to 10.3 ± 1.8 cm 10 min in allodynic skin (p < 0.01) for cowhage and -56% from 20.0 ± 3.5 cm 10 min in normal skin to 8.8 ± 2.3 cm 10 min allodynic skin (p < 0.001) for histamine. The pain responses to the pruritogens were not significantly altered between the areas of allodynia and normal skin (p > 0.1). An additional experiment showed that pinprick hyperalgesia in the absence of allodynia was sufficient to evoke the observed reduced sensitivity to itch stimuli., Conclusions: Cutaneous sensitization (secondary allodynia and hyperalgesia) reduced itch responses regardless of the type of itch model applied and without attenuation of the associated pruritogen-induced pain responses. This could explain the decreased sensitivity to itch provocations previously observed in patients with chronic pain., Significance: This study shows that the neuronal sensitization processes underlying the development secondary hyperalgesia involve significant gating of histaminergic as well as non-histaminergic pruriceptive transmission. Because these itch provocations normally target specific subpopulations of C-nociceptors they could be of relevance for exploratory purposes in pain patients., (© 2017 European Pain Federation - EFIC®.)

The aim of this study was to demonstrate the differences of metabolomics changes in a hyperalgesia model and find potent biomarkers of hyperalgesia. Seven rats were placed in metabolic cages. An emulsion containing 500 μg of Complete Freund's adjuvant (CFA) was used to induce hyperalgesia. Urine samples were collected prior to the injection of CFA and on post-injection days 1, 3 and 7. Ultraperformance liquid chromatography, coupled with quadrupole-time-of-flight mass spectrometry (UHPLC-Q-TOF/MS), was used for a quantitative analysis of urinary metabolic changes in the CFA-induced hyperalgesia model. Differences between the metabolic profiles of the rats in the four groups were analyzed using partial least squares discriminant analysis. Thirty-four potential urine metabolite biomarkers were identified, which changed in a trend similar to the pain threshold. These potential biomarkers were involved in 11 metabolic pathways, as follows: alanine, aspartate, and glutamate metabolism; ascorbate and aldarate metabolism; glycerolipid metabolism; glycerophospholipid metabolism; histidine metabolism; phenylalanine metabolism; sphingolipid metabolism; tryptophan metabolism; tyrosine metabolism; valine, leucine and isoleucine biosynthesis; and vitamin B6 metabolism. These results may improve our understanding of hyperalgesia and provide a basis for the clinical diagnosis of hyperalgesia., (Copyright © 2016 John Wiley & Sons, Ltd.)

Background: Tissue pH is lowered in inflamed tissues, and the increased proton concentration activates acid-sensing ion channels (ASICs), contributing to pain and hyperalgesia. ASICs can be upregulated by nerve growth factor (NGF). The aim of this study was to investigate two new human experimental pain models combining NGF- and acid-induced pain in a randomized, controlled, double-blind study., Methods: In experiment 1, volunteers (N = 16) received an injection of either NGF or isotonic saline in each infrapatellar fat pad (IFP). One day after 5 mL of phosphate-buffered acidic saline was infused into each IFP at a rate of 20 mL/h. In experiment 2, the tibialis anterior (TA) muscle of additional volunteers (N = 16) was examined, following the same procedure except that the volume and infusion rate of acid were different (10 mL, 30 mL/h). Continuous pain ratings were recorded during and after acid infusions. In addition, soreness scores on a Likert scale and pressure pain thresholds (PPTs) were assessed., Results: The PPT of the IFP was significantly decreased at the NGF injection site on day 1, but acid-provoked pain ratings and the change in PPT from pre- to postinfusion between the knees were similar. In the muscle pain model, local mechanical hyperalgesia developed 3 h after the NGF injection and a significant additional decrease in PPT was found after acid infusion compared to preinfusion., Conclusions: NGF sensitization in the IFP was not facilitated by acid, whereas an acid-provoked enhancement of muscle hyperalgesia was found. NGF sensitization of adipose tissue responds differently to acid provocation compared to muscle tissue., Significance: Quantification of two novel pain models combining NGF and acid. Hyperalgesia developed after NGF injection in the infrapatellar fat pad, but it was not facilitated by acid provocation. Contrary, NGF-induced hyperalgesia in muscle tissue was enhanced by acid., (© 2016 European Pain Federation - EFIC®.)

Background: Multiple mechanisms contribute to the stimulus-evoked pain hypersensitivity that may be experienced after peripheral inflammation. Persistent pathological stimuli in many pain conditions affect the expression of certain genes through epigenetic alternations. The main purpose of our study was to investigate the role of epigenetic modification on potassium-chloride co-transporter 2 (KCC2) gene expression in the persistence of inflammatory pain., Methods: Persistent inflammatory pain was induced through the injection of complete Freund's adjuvant (CFA) in the left hind paw of rats. Acetyl-histone H3 and H4 level was determined by chromatin immunoprecipitation in the spinal dorsal horn. Pain behaviour and inhibitory synaptic function of spinal cord were determined before and after CFA injection. KCC2 expression was determined by real time RT-PCR and Western blot. Intrathecal KCC2 siRNA (2 μg per 10 μL per rat) or HDAC inhibitor (10 μg per 10 μL per rat) was injected once daily for 3 days before CFA injection., Results: Persistent inflammatory pain epigenetically suppressed KCC2 expression through histone deacetylase (HDAC)-mediated histone hypoacetylation, resulting in decreased inhibitory signalling efficacy. KCC2 knock-down caused by intrathecal administration of KCC2 siRNA in naïve rats reduced KCC2 expression in the spinal cord, leading to sensitized pain behaviours and impaired inhibitory synaptic transmission in their spinal cords. Moreover, intrathecal HDAC inhibitor injection in CFA rats increased KCC2 expression, partially restoring the spinal inhibitory synaptic transmission and relieving the sensitized pain behaviour., Conclusion: These findings suggest that the transcription of spinal KCC2 is regulated by histone acetylation epigenetically following CFA., Significance: Persistent pain suppresses KCC2 expression through HDAC-mediated histone hypoacetylation and consequently impairs the inhibitory function of inhibitory interneurons. Drugs such as HDAC inhibitors that suppress the influences of persistent pain on the expression of KCC2 may serve as a novel analgesic., (© 2016 European Pain Federation - EFIC®.)

Background: Opioids are known to relieve pain, and also aggravate pre-existing hyperalgesia. In animal studies, the N-methyl-d-aspartate-receptor antagonist nitrous oxide (N2 O) was able to prevent hyperalgesia. The present study evaluated the effect of N2 O on hyperalgesia after remifentanil infusion in healthy volunteers., Methods: Twenty-one healthy volunteers were enrolled in this placebo-controlled cross-over study. Transcutaneous electrical stimulation at high current densities induced spontaneous acute pain and stable areas of hyperalgesia. Each volunteer underwent the following four sessions: (1) 50-50% N2 -O2 and i.v. saline; (2) 50-50% N2 -O2 and i.v. remifentanil 0.1 μg/kg/min; (3) 50-50% N2 O-O2 and i.v. saline; (4) 50-50% N2 O-O2 and i.v. remifentanil 0.1 μg/kg/min. Inhaled gas mixtures lasted for 60 min, i.v. drug administration for 30 min. Visual analogue scale pain intensity, areas of pinprick hyperalgesia and touch-evoked allodynia were assessed repeatedly for 160 min., Results: Data of 19 volunteers were analysed. There were significant time and treatment effects regarding areas of hyperalgesia and allodynia (p < 0.02). The area of hyperalgesia was significantly reduced in the N2 O + remifentanil session compared to the remifentanil session (35.88 ± 22.37 vs. 43.55 ± 18.48 cm(2) , p = 0.004). The area of allodynia was significantly reduced in the N2 O + remifentanil session compared to the remifentanil session (29.95 ± 16.15 vs. 34.80 ± 15.35 cm(2) , p = 0.008). The pain intensity was significantly reduced in the N2 O + remifentanil session compared to the remifentanil session (37.96 ± 12.78 vs. 42.15 ± 13.34 mm, p < 0.0001)., Conclusions: Nitrous oxide significantly reduced hyperalgesia, allodynia and pain intensity aggravated after remifentanil administration in a human volunteer model. WHAT DOES THIS STUDY ADD?: This study brings the evidence that N2 O reduces the remifentanil aggravated secondary hyperalgesia in human volunteers exposed to a well-known model of electrical pain. N2 O was able to oppose the hyperalgesia, the allodynia and the pain intensity consecutive to remifentanil use in this specific pain model., (© 2016 European Pain Federation - EFIC®)

Background: Chronic opioid treatment is complicated by the development of tolerance and hyperalgesia. Social environment alters both opioid-induced behaviours and nociceptive mechanisms. Our previous studies demonstrated that, in adolescent rodents, the susceptibility to acquire opioid dependence and reward is dependent on the nature of social housing conditions. Specifically, our previous studies demonstrate that housing morphine-treated mice with drug-naïve animals mitigates the abuse liability of opioids. Thus, this study tested the effect of social housing conditions on the development of adaptive processes to morphine antinociception., Method: Adolescent males were group-housed in different conditions. In the mixed treatment condition, mice treated with 20 mg/kg morphine (i.e. 'morphine cage-mates') and saline (i.e. 'saline cage-mates') were housed together. In the separated treatment conditions, all mice in the cage received morphine (i.e. 'morphine only') or saline (i.e. 'saline only'). All animals were tested for baseline pain sensitivity and for the response to morphine in the tail withdrawal, hot plate, acetone and von Frey filament tests, during and after discontinuation of opioid treatment., Results: Both morphine cage-mate and morphine only animals developed antinociceptive tolerance. However, this effect was more robust and persistent in the morphine only group. Notably, morphine only animals, but not morphine cage-mates, developed opioid-induced hyperalgesia., Conclusion: This study demonstrates that housing morphine-treated mice with drug-naïve animals mitigates the development of opioid-induced hyperalgesia and antinociceptive tolerance. Thus, this study indicates that social environment influences the effectiveness of opioid pain management., (© 2016 European Pain Federation - EFIC®)

Background: The assessment of facial expressions associated with pain has been used to evaluate pain in humans and has recently found application in non-human mammals. These so called 'grimace scales' have the potential to be developed into a widely accepted non-invasive method of measuring pain in laboratory rodents. Currently, common methodologies to assess pain rely on nociceptive tests that assess stimulus evoked withdrawal responses. These tests, however, are limited to the assessment of a reflexive response without an affective component. This study aimed to use the recently developed Rat Grimace Scale (RGS) and assess its relationship with a conventional nociceptive test (the application of von Frey filaments)., Methods: Fifty-two adult, male Wistar rats were randomized to one of five treatment groups: intraplantar carrageenan, intraplantar complete Freund's adjuvant (CFA), plantar incision, anaesthetic control and saline injection control. The RGS and response to mechanical hypersensitivity testing was evaluated at predetermined time points before and after treatment until withdrawal responses returned to baseline levels., Results: The RGS score significantly increased in all pain models. The peak RGS score also coincided with the development of paw hypersensitivity. However, mechanical hypersensitivity persisted after RGS scores returned to baseline., Conclusion: This study confirms that the three pain models induce pain in rodents and showed that peak pain coincided with peak mechanical hypersensitivity. However, mechanical hypersensitivity remained once pain subsided, mimicking the human experience of CFA injection. These findings further our understanding of the roles of, and relationship between, these assays in the assessment of nociception and pain., (© 2015 European Pain Federation - EFIC®)

Background and Purpose: The Rho GTPase, Rac1, is involved in the pathogenesis of neuropathic pain induced by malformation of dendritic spines in the spinal dorsal horn (sDH) neurons. In the present study, the contribution of spinal Rac1 to peripheral inflammatory pain was studied., Experimental Approach: Effects of s.c. bee venom (BV) injection on cellular localization of Rac1 in the rat sDH was determined with double labelling immunofluorescence. Activation of Rac1 and its downstream effector p21-activated kinase (PAK), ERKs and p38 MAPK in inflammatory pain states was evaluated with a pull-down assay and Western blotting. The preventive and therapeutic analgesic effects of intrathecal administration of NSC23766, a selective inhibitor of Rac1, on BV-induced spontaneous nociception and pain hypersensitivity were investigated., Key Results: Rac1 labelling was mainly localized within neurons in both the superficial and deep layers of the sDH in rats of naïve, vehicle-treated and inflamed (BV injected) groups. GTP-Rac1-PAK and ERKs/p38 were activated following s.c. BV injection. Post-treatment with intrathecal NSC23766 significantly inhibited GTP-Rac1 activity and phosphorylation of Rac1-PAK, ERKs and p38 MAPK in the sDH. Both pre-treatment and post-treatment with intrathecal NSC23766 dose-dependently attenuated the paw flinches, primary thermal and mechanical hyperalgesia and the mirror-image thermal hyperalgesia induced by BV injection, but without affecting the baseline pain sensitivity and motor coordination., Conclusions and Implications: The spinal GTP-Rac1-PAK-ERK/p38MAPK signalling pathway is involved in both the development and maintenance of peripheral inflammatory pain and can be used as a potential molecular target for developing a novel therapeutic strategy for clinical pain., (© 2015 The British Pharmacological Society.)

Background: Chemotherapeutic agents, such as cisplatin, are known to induce a persistent polyneuropathy. The mechanisms underlying the development of this pain are complex, and have only been investigated rodent models using male animals, despite an equivalent presentation of neuropathy between the sexes, clinically., Methods: Male and female C57Bl/6, Tlr3(-/-) Tlr4(-/-) , Myd88(-/-) , Trif(lps2) and Myd88(-/-) /Trif(lps2) mice received 6 i.p. injections of cisplatin (2.3 mg/kg/day) every other day over the course of 2 weeks. Changes in tactile threshold were monitored during this time, continuing through day 23, using von Frey filaments., Results: Male WT mice develop a persistent tactile allodynia resulting from cisplatin administration. Female mice develop an initial allodynia, but thresholds return to baseline by day 23. Deletion of TLR3, TLR4, MyD88 and Trif/MyD88 protects animals from the development of cisplatin-induced polyneuropathy, and there are no sex differences. Trif(lps2) male mice show a persistent tactile allodynia following cisplatin administration, while female mice show a reduced allodynia, and remain higher in threshold than their male counterparts. On day 18, animals were given the analgesic gabapentin, and thresholds were tested 45 min after. Gabapentin was effective in transiently reversing mechanical allodynia in those mice with lowered thresholds., Conclusions: It is important to continue examining both sexes in various pain models, as a mononeuropathy and polyneuropathy show sex differences in pain development and the role of TLR signalling., (© 2015 European Pain Federation - EFIC®)

Background: EphB receptors and their ephrinB ligands are implicated in modulating spinal nociceptive information processing. Here, we investigated whether cyclooxygenase-2 (COX-2), acts as a downstream effector, participates in the modulation of spinal nociceptive information related to ephrinB/EphB signalling., Methods: Thermal hyperalgesia and mechanical allodynia were measured by using radiant heat and von Frey filaments test, respectively. Real-time PCR (RT-PCR) was used to detect the expression of spinal COX-2 mRNA. Spinal COX-2 and extracellular signal-regulated kinase (ERK) protein were determined by Western blot analysis., Results: Intrathecal injection of ephrinB2-Fc caused thermal hyperalgesia and mechanical allodynia, which were accompanied by increased expression of spinal COX-2 mRNA and protein. Inhibition of spinal COX-2 prevented and reversed pain behaviours induced by the intrathecal injection of ephrinB2-Fc. Blockade of EphB receptors by intrathecal injection of EphB2-Fc reduced complete Freund's adjuvant (CFA)-induced inflammatory pain behaviours, which were accompanied by decreased expression of spinal COX-2 mRNA and protein. Furthermore, treatment with U0126, a mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) inhibitor, suppressed spinal ERK activation and COX-2 mRNA and protein expression induced by intrathecal injection of ephrinB1-Fc., Conclusions: These results confirmed the important involvement of COX-2 in the modulation of spinal nociceptive information related to ephrinBs-EphBs signalling., (© 2015 European Pain Federation - EFIC®)

Background: Nerve growth factor (NGF) causes early heat and delayed mechanical hyperalgesia. Axonal transport might contribute to lasting responses. Temporal hyperalgesia development was investigated by administering NGF in paraspinal skin. Transient receptor potential ankyrin 1 (TRPA1) is up-regulated by NGF and chemical responsiveness to cinnamon aldehyde (TRPA1 agonist) was quantified., Methods: Eight healthy volunteers received 1 μg human recombinant NGF (i.d. 50 μL) to L4/L5 processi spinosi skin. Mechanical, thermal and electrical sensitization was assessed at 3-6 h and at days 1, 2, 3, 5, 7, 10, 14 and 21, and pain upon cinnamon aldehyde (20%, 60 μL) recorded at days 3 and 21., Results: Heat hyperalgesia developed with an initial maximum at 3 h [heat pain threshold -3.9°; peak pain ratings +22 visual analogue scale (VAS)] that decreased by day 1, subsequently increased to a maximum around day 5 (-5 ± 0.2 °C, +41 ± 4 VAS), and thereafter declined to ∼20% at day 21. Mechanical and electrical hyperexcitability developed within 3 days and gradually increased to peak between days 14 and 21. Pain intensity upon cinnamon aldehyde stimulation was doubled at the NGF site at day 3 and was still increased by about 50% at day 21., Conclusions: NGF causes immediate heat hyperalgesia probably linked to an up-regulation and sensitization of transient receptor potential vanilloid 1 and possibly other proteins involved in heat transduction. The delayed mechanical hyperalgesia is apparently independent of the time required for axonal transport of NGF receptor complexes. Local mRNA translation at axonal terminals and protein accumulation is hypothesized being involved in sustained NGF-evoked hyperalgesia., (© 2014 European Pain Federation - EFIC®)

Background: Synovial fluid in inflamed joint shows a drop in pH, which activates proton-gated ion channels in nociceptors. No studies have ever tried to develop and characterize acid-induced joint pain., Methods: Rats were injected intra-articularly with pH 4.0 acidic saline twice, 5 days apart. Pain-related behaviour tests including weight-bearing asymmetry, paw withdrawal threshold and knee compression threshold were conducted. To clarify the roles of proton-gated ion channels, rats were injected intra-articularly with selective antagonists for ASIC1a, ASIC3 and TRPV1 on day 5 (before the second injection) or on day 14. Underlying peripheral and central pain mechanisms were evaluated using joint histology, interleukin-1β concentrations in the synovium, single-fibre recording of the knee afferent and expression of phosphorylated cyclic adenosine monophosphate-responsive element-binding protein (p-CREB) in the spinal dorsal horn., Results: Repeated injections of acidic saline induced weight-bearing asymmetry, decrease in paw withdrawal threshold and knee compression threshold bilaterally, which lasted until day 28. Early administration of ASIC3 antagonist reduced the bilateral and long-lasting hyperalgesia. Neither articular degeneration nor synovial inflammation was observed. C-fibre of the knee afferent was activated by acidic saline, which was attenuated by pre-injection of ASIC3 antagonist. p-CREB expression was transiently up-regulated bilaterally on day 6, but not on day 14., Conclusion: We developed and characterized a model of acid-induced long-lasting bilateral joint pain. Peripheral ASIC3 and spinal p-CREB played important roles for the development of hyperalgesia. This animal model gives insights into the mechanisms of joint pain, which is helpful in developing better pain treatments., (© 2014 European Pain Federation - EFIC®)

Despite being the mainstay of pain management, opioids are limited in their clinical utility by adverse effects, such as tolerance and paradoxical hyperalgesia. Research of the past 15 years has extended beyond neurons, to implicate central nervous system immune signaling in these adverse effects. This article will provide an overview of these central immune mechanisms in opioid tolerance and paradoxical hyperalgesia, including those mediated by Toll-like receptor 4, purinergic, ceramide, and chemokine signaling. Challenges for the future, as well as new lines of investigation will be highlighted., (© 2015 American Headache Society.)

Background: Previous studies suggest that persistent post-surgical pain (PPSP) is correlated with preoperative pain status and amplification of central sensitization. Protein kinase Mζ (PKMζ) is an essential substrate of the late long-term potentiation underlying central sensitization, which is one mechanism of pain memory formation. However, the potential contributions of spinal PKMζ to PPSP, a condition in which preoperative pain is prevalent, are not known., Methods: Here, a modified 'hyperalgesia priming' model was established to simulate the clinical situation. This model used intraplantar injections of carrageenan (Car) as priming stimuli to elicit persistent nociceptive sensitization after plantar incision in rats. Upon treatment with PKMζ inhibitor ZIP, Scr-ZIP or protein kinase Cs (PKCs) inhibitor NPC-15437, altered behaviour and spinal PKMζ/PKCs expression were observed., Results: A long-lasting hypersensitivity induced by Car-priming was identified and precipitated by subsequent plantar incision in this 'two-hit' paradigm. Post-treatment with ZIP, but not Scr-ZIP and NPC-15437, after the resolution of Car-priming selectively relieved hypersensitivity. In contrast, pre-priming NPC-15437 treatment only prevented Car-induced initial transient hyperalgesia. Immunoassays showed a significant decrease in spinal PKMζ expression after plantar incision with post-priming ZIP treatment as compared with Scr-ZIP and NPC-15437, but no notable differences in PKCs expression were observed., Conclusions: Spinal PKCs solely contribute to the initial induction of persistent pain, whereas PKMζ plays an essential role in spinal plasticity storage. PKMζ is responsible for the maintenance of peripheral inflammation-primed PPSP. Therefore, spinal PKMζ may be a therapeutic target to prevent surgery-induced chronic pain in patients with preoperative pain., (© 2014 European Pain Federation - EFIC®)

Background: We investigated both the efficacy and the sub-chronic toxicity of Tephrosia toxicaria Pers. in the zymosan-induced temporomandibular joint (TMJ) inflammatory hypernociception in rats evaluating the possible role of heme oxygenase-1 (HO-1)., Methods: Rats were pretreated with T. toxicaria (0.2, 2.0 or 20 mg/kg) 60 min before the intra-articular injection of zymosan (2 mg, 40 μL) in the left TMJ. In another series of experiments, rats were treated with ZnPP-IX (3 mg/kg), a specific HO-1 inhibitor, before T. toxicaria (20 mg/kg). Von Frey test was used to evaluate inflammatory hypernociception (g) 4 h after zymosan injection. Six hours after zymosan injection, the synovial lavage was collected for total cell count and myeloperoxidase (MPO) activity, and joint tissue for histopathological analysis and immunohistochemistry for HO-1. To evaluate the sub-chronic toxicity, mice received T. toxicaria (20 mg/kg) or saline once a day for 14 days to analyse body mass, organ weight and biochemical parameters., Results: T. toxicaria partially reversed the zymosan-induced head withdrawal threshold, the number of cells and the MPO activity. T. toxicaria reduced the inflammatory cell influx in the synovial membrane. TMJ immunohistochemical analyses treated with T. toxicaria showed increased HO-1 expression. These effects of T. toxicaria were not observed in the presence of ZnPP-IX. T. toxicaria treatment for 14 days did not show significant signs of toxicity when administrated to mice., Conclusions: T. toxicaria did not produce any signs of toxicity and effectively decreased zymosan-induced TMJ inflammatory hypernociception dependent, at least in part, upon the HO-1 pathway integrity., (© 2014 European Pain Federation - EFIC®)

Background: One of the major unresolved issues in treating pain is the paradoxical hyperalgesia produced by opiates, and accumulating evidence implicate that EphBs receptors and ephrinBs ligands are involved in mediation of spinal nociceptive information and central sensitization, but the manner in which ephrinB/EphB signalling acts on spinal nociceptive information networks to produce hyperalgesia remains enigmatic. The objective of this research was to investigate the role of ephrinB/EphB signalling in remifentanil-induced hyperalgesia (RIH) and its downstream effector., Methods: We characterized the remifentanil-induced pain behaviours by evaluating thermal hyperalgesia and mechanical allodynia in a rat hind paw incisional model. Protein expression of EphB1 receptor and ephrinB1 ligand in spinal dorsal horn cord was determined by Western blotting, and Fos was determined by immunohistochemistry assay, respectively. To figure out the manner in which ephrinB/EphB signalling acts with N-methyl-d-aspartic acid (NMDA) receptor, we used MK-801, an antagonist of NMDA receptor, trying to suppressed the hyperalgesia induced by ephrinB1-Fc, an agonist of ephrinB/EphB., Results: Continuing infusion of remifentanil produced a thermal hyperalgesia and mechanical allodynia, which was accompanied with increased protein expression of spinal-level EphB1 receptor, ephrinB1 ligand and Fos; what appeared above was suppressed by pretreatment with EphB1-Fc, an antagonist of ephrinB/EphB or MK-801, and increased pain behaviours induced by intrathecal injection of ephrinB1-Fc, an agonist of ephrinB/EphB, were suppressed by MK-801., Conclusions: Our findings indicated that ephrinB/EphB signalling is involved in RIH. EphrinB/EphB signalling might be the upstream of NMDA receptor., (© 2014 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC®.)

Objective: To investigate age-related differences in mechanical sensitivity to inflammatory pain and determine the contribution of transient receptor potential ankyrin 1 (TRPA1) to mechanical hypersensitivity during chronic inflammation in young and aged mice with complete Freund's adjuvant (CFA)-induced arthritis., Methods: Mechanical sensitivity in young (3-month-old) and aged (24-month-old) wild-type (TRPA1(+/+) ) mice and TRPA1-deficient (TRPA1(-/-) ) mice was measured behaviorally for 8 weeks following injection of CFA into the plantar hind paw. The severity of inflammation was evaluated by histologic analyses and hind-paw measurements. Ex vivo preparations of the skin saphenous nerve from mice were assessed for C-fiber sensitivity., Results: Among naive (uninjured) wild-type mice, aged animals were less sensitive than young animals to mechanical stimuli. Afferent recordings of C-fibers from TRPA1(-/-) mice indicated that TRPA1 contributes to the normal mechanical sensitivity in both age groups. Following injection of CFA, both young and aged TRPA1(+/+) mice exhibited mechanical hypersensitivity. In young TRPA1(-/-) mice injected with CFA, peak development of mechanical hypersensitivity was delayed until week 4, when they exhibited a sharp decrease (9-fold) in the mechanical paw withdrawal threshold, whereas aged TRPA1(-/-) mice did not exhibit mechanical hypersensitivity at any time during the 8 weeks after CFA injection. Recordings of C-fibers from the saphenous nerve supported these findings, with results indicating that both young and aged TRPA1(+/+) mice exhibited increased action potential firing at 8 weeks after CFA injection (increases of 25% and 60%, respectively). Interestingly, among TRPA1(-/-) mice injected with CFA, mechanical firing was increased markedly in the C-fibers of young mice (increase of 80%) but not in the C-fibers of aged mice., Conclusion: These findings reveal marked differences in the long-term mechanical behavioral sensitivity of aged and young mice, and suggest that TRPA1 may be a key contributor to the transition from acute to chronic inflammatory pain in response to mechanical stimuli as well as to the development of nociceptor sensitization selectively in aged mice., (Copyright © 2014 by the American College of Rheumatology.)

Background: Both nerve growth factor (NGF) and ultraviolet-B (UV-B) irradiation sensitize nociceptive nerve endings and increase axonal excitability of nociceptors. Combining NGF and UV-B treatment is supra-additive for sensory sensitization and even caused spontaneous pain in about 70% of the subjects., Methods: UV-B irradiation was performed at day 21 after intradermal NGF injection in 13 volunteers. Pain thresholds, electrically induced axon reflex erythema and pain (1.5-fold pain threshold, 5-100 Hz) was analysed at days 22, 24, 28, 35, 49 and 70 and correlated to hyperalgesia and spontaneous pain., Results: Electrical pain threshold after combined NGF/UVB was reduced below single treatment at 24 h but not at 72 h post-UV-B irradiation. At the NGF/UV-B site, electrical pain was enhanced at all frequencies compared with single NGF and UV-B sites at 24 and 72 h with pain ratings exceeding control values about twofold to threefold [65 ± 7 vs. 25 ± 8 visual analogue scale (VAS) (24 h) and 55 ± 9 vs. 22 ± 5 VAS (72 h)]. Hyperalgesia to electrical stimulation correlated with hyperalgesia to pinprick (Spearman r = 0.44; p < 0.001, Bonferroni corr.) and supra-threshold heat (Spearman r = 0.55; p < 0.001) stimulation at 24 h only. Electrical pain thresholds at the NGF/UV-B site weakly correlated to spontaneous pain levels (Spearman r = 0.3; p = 0.025, without Bonferroni correction). In contrast, electrically induced pain or axon reflex erythema did not correlate to spontaneous pain levels., Conclusions: The combination of NGF and UV-B increases axonal excitability that contributes to hyperalgesia and might also facilitate ongoing spontaneous pain.

Background: The role of glial cell line-derived neurotrophic factor (GDNF) in pain and muscular nociceptor activities is not well understood. We examined pain-related behaviour and mechanical response of muscular thin-fibre afferents after intramuscular injection of GDNF in rats., Methods: GDNF and antagonist to transient receptor potential V1 or acid-sensing ion channels were injected into rat gastrocnemius muscle and muscular mechanical hyperalgesia was assessed with a Randall-Selitto analgesiometer. Activities of single C- (conduction velocity < 2.0 m/s) and Aδ-fibres (conduction velocity 2.0-12.0 m/s) were recorded from extensor digitorum longus muscle-nerve preparations in vitro. The changes in the responses to mechanical stimuli before and after GDNF injection were recorded., Results: Mechanical hyperalgesia was observed from 1 h to 1 day after GDNF (0.03 μM, 20 μL) injection. The decreased withdrawal threshold was temporarily reversed after intramuscular injection of amiloride (50 mM, 20 μL), but not capsazepine (50 μM, 20 μL). In single-fibre recordings, both phosphate buffered saline (PBS) and GDNF failed to induce any significant discharges. GDNF significantly enhanced the mechanical response when compared with the PBS group, but only in Aδ-fibre afferents. C-fibres were not affected. Significantly lowered threshold and increased response magnitude to mechanical stimuli were observed 30 or 60-120 min after injection. These times are compatible with the timing of the onset of the hyperalgesic effect of GDNF., Conclusions: These results suggest that GDNF increased the response of muscular Aδ-fibre afferents to mechanical stimuli, resulting in muscular mechanical hyperalgesia., (© 2013 European Pain Federation - EFIC®)

Background: Nitric oxide is a key signalling molecule in the pathogenesis of inflammation, but its role in acute pancreatitis and related abdominal pain induced by secretory phospholipase A2 (sPLA2 ) from Crotalus durissus terrificus (Cdt) venom has not been investigated., Methods: Male Wistar rats were i.v. injected with L-NAME (20 mg/kg), aminoguanidine (AG, 50 mg/kg), 7-nitroindazole (7-NI, 10 mg/kg) or vehicle 10 min before or 60 min after the injection of sPLA2 (300 μg/kg) into the common bile duct. After 4 h of sPLA2 injection, abdominal hyperalgesia and inflammation were assessed in addition to serum amylase, nitrite/nitrate (NOx), pancreas lipoperoxidation and 3-nitrotyrosine (3-NT) contents., Results: sPLA2 -induced acute pancreatitis, related abdominal hyperalgesia, hyperamylasemia and increased concentration of NOx were not correlated with lipoperoxidation or increased 3-NT in the pancreas. Pretreatment with all the nitric oxide synthase (NOS) inhibitors significantly reduced abdominal mechanical hyperalgesia, but only iNOS blockade by AG suppressed pancreas oedema and serum NOx increase. The therapeutic approach with all the NOS inhibitors produced a similar reduction pattern of the abdominal hyperalgesia, but AG treatment also inhibited serum hyperamylasemia and NOx concentrations and pancreatic myeloperoxidase. The nNOS blockade by 7-NI treatment also inhibited myeloperoxidase activity in both pancreas and lung., Conclusions: Therapeutic blockade of iNOS or nNOS provides benefits in terms of inhibition of the acute pancreatitis-related abdominal hyperalgesia, while iNOS inhibition also ameliorates the inflammatory cell influx to the pancreas and reduces the resultant hyperamylasemia and NOx levels, thus representing alternative pharmacological strategies for treatment of clinical pancreatitis associated with increased PLA2 ., (© 2013 European Pain Federation - EFIC®)