Long-lasting Mechanical Allodynia Research Articles

A Cross-Sectional and Longitudinal Integrated Study on Brain Functional Changes in a Neuropathic Pain Rat Model.

Human and animal imaging studies demonstrated that chronic pain profoundly alters the structure and the functionality of several brain regions and even causes mental dysfunctions such as depression and anxiety disorders. In this article, we conducted a multimodal study cross-sectionally and longitudinally, to evaluate how neuropathic pain affects the brain. Using the spared nerve injury (SNI) model which promotes long-lasting mechanical allodynia, results showed that neuropathic pain deeply modified the intrinsic organization of the brain functional network 2 weeks after injury. There are significant changes in the activity of the left thalamus (Th_L) and left olfactory bulb (OB_L) brain regions after SNI, as evidenced by both the blood oxygen level-dependent (BOLD) signal and c-Fos expression. Importantly, these changes were closely related to mechanical pain behavior of rats. However, it is worth noting that after morphine administration for analgesia, only the increased activity in the TH region is reversed, while the decreased activity in the OB region becomes more prominent. Functional connectivity (FC) and c-Fos correlation analysis further showed these two regions of interest (ROIs) exhibit different FC patterns with other brain regions. Our study comprehensively revealed the adaptive changes of brain neural networks induced by nerve injury in both cross-sectional and longitudinal dimensions and emphasized the abnormal activity and FC of Th_L and OB_L in the pathological condition. It provides reliable assistance in exploring the intricate mechanisms of diseases.

Scorpion Venom Component; BmK AGAP Potentiates the Analgesic Effects of Lidocaine During Sciatic Nerve Block

Background: Various analgesic adjuvants, including opioids, have been tested and demon-strated to be clinically beneficial when added to local anesthetics to increase the duration of analgesia with the risk of various adverse effects. This study was designed to test the hypothesis that a scorpion venom component, BmK AGAP, may have a synergistic effect with lidocaine. Methods: We performed partial sciatic nerve ligation on 84 rats to induce a rapid onset and long-lasting mechanical allodynia. An equal volume (600μl) of lidocaine and BmK AGAP were prepared with saline. The rats were randomly assigned to one of seven groups. Group A (n = 12) received saline as the control; Group B (n = 12) received lidocaine alone; Group C (n = 12) received BmK AGAP alone; Group D, E, F and G (n = 12 each) received lidocaine and different concentrations of BmK AGAP combined. The von Frey filaments were used to assess mechanical allodynia in the rats. Results: We observed a decrease in pain intensity and a prolonged duration of analgesia in rats that received BmK AGAP with lidocaine. Conclusion: BmK AGAP with lidocaine decreased pain intensity and increased the duration of analgesia.

CXCL13 contributes to chronic pain of a mouse model of CRPS-I via CXCR5-mediated NF-κB activation and pro-inflammatory cytokine production in spinal cord dorsal horn

BackgroundComplex regional pain syndrome type-I (CRPS-I) causes excruciating pain that affect patients’ life quality. However, the mechanisms underlying CRPS-I are incompletely understood, which hampers the development of target specific therapeutics.MethodsThe mouse chronic post-ischemic pain (CPIP) model was established to mimic CRPS-I. qPCR, Western blot, immunostaining, behavioral assay and pharmacological methods were used to study mechanisms underlying neuroinflammation and chronic pain in spinal cord dorsal horn (SCDH) of CPIP mice.ResultsCPIP mice developed robust and long-lasting mechanical allodynia in bilateral hindpaws. The expression of inflammatory chemokine CXCL13 and its receptor CXCR5 was significantly upregulated in ipsilateral SCDH of CPIP mice. Immunostaining revealed CXCL13 and CXCR5 was predominantly expressed in spinal neurons. Neutralization of spinal CXCL13 or genetic deletion of Cxcr5 (Cxcr5−/−) significantly reduced mechanical allodynia, as well as spinal glial cell overactivation and c-Fos activation in SCDH of CPIP mice. Mechanical pain causes affective disorder in CPIP mice, which was attenuated in Cxcr5−/− mice. Phosphorylated STAT3 co-expressed with CXCL13 in SCDH neurons and contributed to CXCL13 upregulation and mechanical allodynia in CPIP mice. CXCR5 coupled with NF-κB signaling in SCDH neurons to trigger pro-inflammatory cytokine gene Il6 upregulation, contributing to mechanical allodynia. Intrathecal CXCL13 injection produced mechanical allodynia via CXCR5-dependent NF-κB activation. Specific overexpression of CXCL13 in SCDH neurons is sufficient to induce persistent mechanical allodynia in naïve mice.ConclusionsThese results demonstrated a previously unidentified role of CXCL13/CXCR5 signaling in mediating spinal neuroinflammation and mechanical pain in an animal model of CRPS-I. Our work suggests that targeting CXCL13/CXCR5 pathway may lead to novel therapeutic approaches for CRPS-I.

Spinal Astrocyte-Neuron Lactate Shuttle Contributes to the Pituitary Adenylate Cyclase-Activating Polypeptide/PAC1 Receptor-Induced Nociceptive Behaviors in Mice.

We have previously shown that spinal pituitary adenylate cyclase-activating polypeptide (PACAP)/PACAP type 1 (PAC1) receptor signaling triggered long-lasting nociceptive behaviors through astroglial activation in mice. Since astrocyte-neuron lactate shuttle (ANLS) could be essential for long-term synaptic facilitation, we aimed to elucidate a possible involvement of spinal ANLS in the development of the PACAP/PAC1 receptor-induced nociceptive behaviors. A single intrathecal administration of PACAP induced short-term spontaneous aversive behaviors, followed by long-lasting mechanical allodynia in mice. These nociceptive behaviors were inhibited by 1,4-dideoxy-1,4-imino-d-arabinitol (DAB), an inhibitor of glycogenolysis, and this inhibition was reversed by simultaneous L-lactate application. In the cultured spinal astrocytes, the PACAP-evoked glycogenolysis and L-lactate secretion were inhibited by DAB. In addition, a protein kinase C (PKC) inhibitor attenuated the PACAP-induced nociceptive behaviors as well as the PACAP-evoked glycogenolysis and L-lactate secretion. Finally, an inhibitor for the monocarboxylate transporters blocked the L-lactate secretion from the spinal astrocytes and inhibited the PACAP- and spinal nerve ligation-induced nociceptive behaviors. These results suggested that spinal PAC1 receptor-PKC-ANLS signaling contributed to the PACAP-induced nociceptive behaviors. This signaling system could be involved in the peripheral nerve injury-induced pain-like behaviors.

TREK-1 potassium channels participate in acute and long-lasting nociceptive hypersensitivity induced by formalin in rats

TREK-1 channels are expressed in small nociceptive dorsal root ganglion (DRG) neurons where they participate in acute inflammatory and neuropathic pain. However, the role of TREK-1 in persistent pain is not well understood. The aim of this study was to investigate the local peripheral and spinal participation of TREK-1 in formalin-induced acute and long-lasting nociceptive hypersensitivity. Local peripheral or intrathecal pre-treatment with spadin, selective blocker of TREK-1, increased acute flinching behavior and secondary mechanical allodynia and hyperalgesia behavior observed 6 days after formalin injection. Local peripheral or intrathecal pre-treatment with BL-1249, selective opener of TREK-1, decreased long-lasting secondary mechanical allodynia and hyperalgesia induced by formalin. Pre-treatment with BL-1249 prevented the pro-nociceptive effect of spadin on acute nociception and long-lasting mechanical allodynia and hyperalgesia in rats. Pre-treatment with two recombinant channels that produce a high TREK-1 current, S300A and S333A (non-phosphorylated state of TREK-1), reduced formalin-induced acute pain and long-lasting mechanical allodynia and hyperalgesia. Besides, post-treatment with S300A, S333A or BL-1249 reversed long-lasting mechanical allodynia and hyperalgesia induced by formalin. Formalin increased TREK-1 expression at 1 and 6 days in DRG and dorsal spinal cord in rats, whereas that it increased c-fos expression at the DRG. Intrathecal repeated transfection of rats with S300A and S333A or injection with BL-1249 reduced formalin-induced enhanced c-fos expression. Data suggest that TREK-1 activity at peripheral and spinal sites reduces neuronal excitability in the process of acute and long-lasting nociception induced by formalin in rats.

Endothelin receptor type A is involved in the development of oxaliplatin-induced mechanical allodynia and cold allodynia acting through spinal and peripheral mechanisms in rats.

Oxaliplatin, a platinum-based chemotherapeutic agent, frequently causes severe neuropathic pain typically encompassing cold allodynia and long-lasting mechanical allodynia. Endothelin has been shown to modulate nociceptive transmission in a variety of pain disorders. However, the action of endothelin varies greatly depending on many variables, including pain causes, receptor types (endothelin type A (ETA) and B (ETB) receptors) and organs (periphery and spinal cord). Therefore, in this study, we investigated the role of endothelin in a Sprague–Dawley rat model of oxaliplatin-induced neuropathic pain. Intraperitoneal administration of bosentan, a dual ETA/ETB receptor antagonist, effectively blocked the development or prevented the onset of both cold allodynia and mechanical allodynia. The preventive effects were exclusively mediated by ETA receptor antagonism. Intrathecal administration of an ETA receptor antagonist prevented development of long-lasting mechanical allodynia but not cold allodynia. In marked contrast, an intraplantar ETA receptor antagonist had a suppressive effect on cold allodynia but only had a partial and transient effect on mechanical allodynia. In conclusion, ETA receptor antagonism effectively prevented long-lasting mechanical allodynia through spinal and peripheral actions, while cold allodynia was prevented through peripheral actions.

Allodynia by Splenocytes From Mice With Acid-Induced Fibromyalgia-Like Generalized Pain and Its Sexual Dimorphic Regulation by Brain Microglia.

Fibromyalgia (FM), a disease of unknown etiology characterized by chronic generalized pain, is partly recapitulated in an animal model induced by repeated acid saline injections into the gastrocnemius muscle. Here, we attempted to investigate the sex difference in pain hypersensitivity (mechanical allodynia and hypersensitivity to electrical stimulation) in the repeated acid saline-induced FM-like generalized pain (AcGP) model. The first unilateral acid injection into gastrocnemius muscle at day 0/D0 and second injection at D5 (post day 0, P0) induced transient and long-lasting mechanical allodynia, respectively, on both sides of male and female mice. The pretreatment with gonadectomy did not affect the first injection-induced allodynia in both sexes, but gradually reversed the second injection-induced allodynia in male but not female mice. Moreover, the AcGP in male mice was abolished by intracerebroventricular minocycline treatments during D4–P4 or P5–P11, but not by early treatments during D0–D5 in male but not female mice, suggesting that brain microglia are required for AcGP in late-onset and sex-dependent manners. We also found that the intravenous treatments of splenocytes derived from male but not female mice treated with AcGP caused allodynia in naive mice. In addition, the purified CD4+ T cells derived from splenocytes of acid-treated male mice retained the ability to cause allodynia in naive mice. These findings suggest that FM-like AcGP has multiple sexual dimorphic mechanisms.

Deletion of Acid-Sensing Ion Channel 3 Relieves the Late Phase of Neuropathic Pain by Preventing Neuron Degeneration and Promoting Neuron Repair.

Neuropathic pain is one type of chronic pain that occurs as a result of a lesion or disease to the somatosensory nervous system. Chronic excessive inflammatory response after nerve injury may contribute to the maintenance of persistent pain. Although the role of inflammatory mediators and cytokines in mediating allodynia and hyperalgesia has been extensively studied, the detailed mechanisms of persistent pain or whether the interactions between neurons, glia and immune cells are essential for maintenance of the chronic state have not been completely elucidated. ASIC3, a voltage-insensitive, proton-gated cation channel, is the most essential pH sensor for pain perception. ASIC3 gene expression is increased in dorsal root ganglion neurons after inflammation and nerve injury and ASIC3 is involved in macrophage maturation. ASIC currents are increased after nerve injury. However, whether prolonged hyperalgesia induced by the nerve injury requires ASIC3 and whether ASIC3 regulates neurons, immune cells or glial cells to modulate neuropathic pain remains unknown. We established a model of chronic constriction injury of the sciatic nerve (CCI) in mice. CCI mice showed long-lasting mechanical allodynia and thermal hyperalgesia. CCI also caused long-term inflammation at the sciatic nerve and primary sensory neuron degeneration as well as increased satellite glial expression and ATF3 expression. ASIC3 deficiency shortened mechanical allodynia and attenuated thermal hyperalgesia. ASIC3 gene deletion shifted ATF3 expression from large to small neurons and altered the M1/M2 macrophage ratio, thereby preventing small neuron degeneration and relieved pain.

Spinal caspase-3 contributes to tibial fracture-associated postoperative allodynia via up-regulation of LRRTM1 expression in mice

BackgroundBone fracture may subsequently cause chronic postoperative pain after orthopedic surgery, but mechanisms remain elusive. The necessity of caspase-3 in neuroinflammation and synaptic plasticity has been summarized in pathological pain. Leucine-rich repeat transmembrane protein 1 (LRRTM1) mediates synaptic delivery of AMPA receptor and synaptogenesis. This study evaluated whether caspase-3 and LRRTM1 are required for fracture-associated postoperative allodynia. MethodsA model of tibial fracture with intramedullary pinning in mice was established for the induction of postoperative pain, verified by measurement of mechanical paw withdrawal threshold and cold scores response to acetone. The caspase-3 specific inhibitor, recombinant caspase-3 and LRRTM1 knockdown by shRNA were utilized for the investigation of pathogenesis as well as the prevention of allodynia. Also, the activity of caspase-3 and the expression of LRRTM1 in the spinal dorsal horn were examined by Western blot and RT-qPCR. ResultsThis study reported that tibial fracture and orthopedic surgery produced long-lasting mechanical allodynia and cold allodynia, along with the up-modulation of spinal caspase-3 activity (but not caspase-3 expression) and LRRTM1 expression. Spinal caspase-3 inhibition prevented fracture-associated behavioral allodynia in a dose-dependent manner. Caspase-3 inhibitor also reduced the spinal increased LRRTM1 level after tibial fracture with pinning. Spinal LRRTM1 deficiency impaired fracture-caused postoperative pain. Intrathecal recombinant caspase-3 facilitated acute pain hypersensitivity and spinal LRRTM1 expression in naïve mice, reversing by LRRTM1 knockdown. ConclusionOur current results demonstrate the spinal up-regulation of LRRTM1 by caspase-3 activation in the development of tibial fracture-associated postoperative pain in mice.

Dysregulation of EAAT2 and VGLUT2 Spinal Glutamate Transports via Histone Deacetylase 2 (HDAC2) Contributes to Paclitaxel-induced Painful Neuropathy.

Effective treatments for chemotherapy-induced peripheral neuropathy (CIPN) remain unavailable. Given the significance of spinal cord glutamate transporters in neuronal plasticity and central sensitization, this study investigated the role of excitatory amino acid transporter 2 (EAAT2) and vesicular-glutamate transporter 2 (VGLUT2) in the development of paclitaxel-induced painful neuropathy. Paclitaxel (2 mg/kg, i.p., cumulative dose 8 mg/kg) induced long-lasting mechanical allodynia (>28 days) with increased glutamate concentration and decreased EAAT2 expression with no changes in GABA/glycine or VGAT (vesicular GABA transporter) in rat spinal dorsal horn. VGLUT2 expression was upregulated and coexpressed with enhanced synaptophysin, characterizing nociceptive afferent sprouting and new synapse formation of glutamatergic neurons in the spinal cord dorsal horn. HDAC2 and transcription factor YY1 were also upregulated, and their interaction and colocalization were confirmed following paclitaxel treatment using co-immunoprecipitation. Inhibition or knockdown of HDAC2 expression by valproic acid, BRD6688, or HDAC2 siRNA not only attenuated paclitaxel-induced mechanical allodynia but also suppressed HDAC2 upregulation, glutamate accumulation, and the corresponding changes in EAAT2/VGLUT/synaptophysin expression and HDAC2/YY1 interaction. These findings indicate that loss of the balance between glutamate release and reuptake due to dysregulation EAAT2/VGLUT2/synaptophysin cascade in the spinal dorsal horn plays an important role in the development of paclitaxel-induced neuropathic pain. HDAC2/YY1 interaction as a complex appears essential in regulating this pathway, which can potentially be a therapeutic target to relieve CIPN by reversing central sensitization of spinal nociceptive neurons.

Effects of Aging on Formalin-Induced Pain Behavior and Analgesic Activity of Gabapentin in C57BL/6 Mice.

Improved living conditions have induced an increase of lifespan often accompanied by comorbidities, responsible for pain, and by cognitive impairment and dementia, impairing communication capabilities. In most cases, the elderly do not receive pain relief because of underdiagnosis and of aging-induced changes of systems affecting nociceptive response. Unrelieved pain is involved in the development of behavioral symptoms, as agitation, representing a difficult challenge in this fragile population. Aged C57BL/6 mice and amyloid precursor protein (APP) mice display behavioral disturbances that mimic behavioral and psychological symptoms of dementia (BPSD). Therefore, this original study focuses on the influence of aging on nociception to provide insight into the occurrence of BPSD. We have investigated how aging can affect nociception after formalin administration and gabapentin effect in C57BL/6 mice, since it represents one of the treatments of choice for chronic neuropathic pain. Based on our results, changes of nociceptive behavior in response to an algogen stimulus occur during aging. Formalin-induced behavioral pattern in older C57BL/6 mice presents a temporal shift and an increase in the peak amplitudes. Our data show that the effectiveness of gabapentin is influenced by the age of the animal; though preliminary, the latter provide evidence upon which formalin test induced long-lasting mechanical allodynia might be a reliable as rapid and viable persistent pain model. The disclosed differences in effectiveness of gabapentin according to age can form the rational basis to deepen the study of pain treatment in the elderly.

Thiamine, riboflavin, and nicotinamide inhibit paclitaxel-induced allodynia by reducing TNF-α and CXCL-1 in dorsal root ganglia and thalamus and activating ATP-sensitive potassium channels.

Some B vitamins exhibit activities in models of nociceptive pain, inflammatory pain, and neuropathic pain induced by nerve lesions and also in certain painful conditions in humans. In the present study, we investigated the effects of thiamine, riboflavin, and nicotinamide in a neuropathic pain model induced by the chemotherapeutic paclitaxel in mice. Four intraperitoneal (i.p.) administrations of paclitaxel (2mg/kg day, cumulative dose 8mg/kg) induced a long-lasting mechanical allodynia. Per os (p.o.) administration of two doses of thiamine (150, 300 and 600mg/kg), nicotinamide (250, 500 and 1000mg/kg) or riboflavin (125, 250 and 500mg/kg), on the seventh day after the first administration of paclitaxel, the mechanical allodynia was attenuated. The antinociceptive activity of all B vitamins was attenuated by glibenclamide (20 and 10mg/kg, p.o.). Naltrexone (5 and 10mg/kg, i.p.) attenuated the antinociceptive activity of thiamine.Thiamine, riboflavin, and nicotinamide also reduced the concentrations of tumor necrosis factor-α (TNF-α) and CXCL-1 in dorsal root ganglia (DRG) and thalamus.In conclusion, thiamine, riboflavin, and nicotinamide exhibit antinociceptive activity in the neuropathic pain model induced by paclitaxel. Inhibition of TNF-αand CXCL-1 production in DRG and thalamus, as well as activation of ATP-sensitive potassium channels,underly theirantinociceptive activity.

In Silico Screening Identified Novel Small-molecule Antagonists of PAC1 Receptor.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptors are present in the spinal dorsal horn and dorsal root ganglia, suggesting an important role of PACAP signaling systems in the modulation of spinal nociceptive transmission. Previously, we found that intrathecal injection of PACAP or maxadilan, a selective PACAP type I (PAC1) receptor agonist, induced transient aversive responses followed by a long-lasting mechanical allodynia in mice, suggesting that PACAP-PAC1 receptor systems are involved in chronic pain and that selective PAC1 antagonists may become a new class of analgesics. Although several PAC1 antagonists, such as PACAP 6-38, have been reported, all of them are peptide compounds. In the present study, we identified new small-molecule antagonists of the PAC1 receptor using in silico screening and in vitro/vivo pharmacological assays. The identified small-molecule compounds, named PA-8 and PA-9, dose dependently inhibited the phosphorylation of CREB induced by PACAP in PAC1-, but not VPAC1- or VPAC2-receptor-expressing CHO cells. PA-8 and PA-9 also dose dependently inhibited PACAP-induced cAMP elevation with an IC50 of 2.0 and 5.6 nM, respectively. In vivo pharmacological assays showed that intrathecal injection of these compounds blocked the induction of PACAP-induced aversive responses and mechanical allodynia in mice. In contrast, the compounds when administered alone exerted neither agonistic nor algesic actions in the in vitro/vivo assays. The compounds identified in the present study are new and the first small-molecule antagonists of the PAC1 receptor; they may become seed compounds for developing novel analgesics.

Anion exchanger 3 in dorsal root ganglion contributes to nerve injury-induced chronic mechanical allodynia and thermal hyperalgesia.

To determine the role of anion exchanger 3 (AE3) in dorsal root ganglion (DRG) in nerve injury-induced chronic nociception in the rat. Spared nerve injury (SNI) was used to induce neuropathic pain. Von Frey filaments and Hargreaves test were used to assess tactile allodynia and thermal hyperalgesia, respectively. Drugs were given by intrathecal administration. Western blotting was used to determine AE3 expression in DRG. SNI produced long-lasting mechanical allodynia and thermal hyperalgesia. AE3 was found in DRG of sham-operated rats. SNI enhanced baseline AE3 expression in L4 and L5 DRGs at days 7 and 14, respectively. In contrast, SNI did not affect AE3 expression in L6 DRG. AE3 expression returned to baseline levels 21 days after SNI. Intrathecal 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) (5-50 μg) pretreatment prevented SNI-induced allodynia and, at a lesser extent, hyperalgesia. Moreover, DIDS (50 μg) reduced SNI-induced AE3 upregulation in L4, but not L5, DRGs. Intrathecal DIDS (5-50 μg) or anti-AE3 antibody (1 μg), but not vehicle, post-treatment (6 days) partially reversed SNI-induced allodynia and hyperalgesia. DIDS or anti-AE3 antibody post-treatment diminished SNI-induced AE3 upregulation in L4 and L5 DRGs. Data suggest that AE3 is present in DRG and contributes to mechanical allodynia and thermal hyperalgesia in neuropathic rats.

Contralateral Electroacupuncture Relieves Chronic Neuropathic Pain in Rats with Spared Nerve Injury.

BackgroundAcupuncture and electroacupuncture (EA) are widely applied in the treatment of various conditions, including pain. Acupuncture stimulation is applied not only in areas close to pain sites, but also in distal regions or on the contralateral side of the body. Identifying which acupuncture paradigms produce best therapeutic effects is of clinical significance.Material/MethodsSpared nerve injury (SNI) was applied to establish a rat model of neuropathic pain. We applied 14 sessions of EA (BL 60 and BL 40, 1–2 mA, and 2 Hz, 30 min per session) every other day from days 3 to 29 after surgery on the contralateral or ipsilateral side of pain. von Frey hair was applied to examine mechanical allodynia in the SNI model and analgesic effects of EA. All experimental procedures were approved by the Animal Care and Use Committee of our university, according to the guidelines of the International Association for the Study of Pain.ResultsSNI produced significant and long-lasting mechanical allodynia (p<0.001) in injured paws. Repeated EA on the contralateral side of the pain significantly attenuated mechanical allodynia from 14 days after surgery (p<0.05). By contrast, ipsilateral EA did not show analgesic effects (p>0.05).ConclusionsThese findings indicate that contralateral EA is superior to local EA in some types of pain disorders. Further investigations are needed for a more comprehensive understanding of the central mechanisms of acupuncture.

Role of Thrombin in Soluble Thrombomodulin-Induced Suppression of Peripheral HMGB1-Mediated Allodynia in Mice.

High mobility group box1 (HMGB1), a nuclear protein, once released into the extracellular space under pathological conditions, plays a pronociceptive role in redox-dependent distinct active forms, all-thiol HMGB1 (at-HMGB1) and disulfide HMGB1 (ds-HMGB1), that accelerate nociception through the receptor for advanced glycation endproducts (RAGE) and Toll-like receptor 4 (TLR4), respectively. Thrombomodulin (TM), an endothelial membrane protein, and soluble TM, known as TMα, promote thrombin-mediated activation of protein C and also sequester HMGB1, which might facilitate thrombin degradation of HMGB1. The present study aimed at clarifying the role of thrombin in TMα-induced suppression of peripheral HMGB1-dependent allodynia in mice. Thrombin-induced degradation of at-HMGB1 and ds-HMGB1 was accelerated by TMα in vitro. Intraplantar (i.pl.) injection of bovine thymus-derived HMGB1 in an unknown redox state, at-HMGB1, ds-HMGB1 or lipopolysaccharide (LPS), known to cause HMGB1 secretion, produced long-lasting mechanical allodynia in mice, as assessed by von Frey test. TMα, when preadministered i.pl., prevented the allodynia caused by bovine thymus-derived HMGB1, at-HMGB1, ds-HMGB1 or LPS, in a dose-dependent manner. The TMα-induced suppression of the allodynia following i.pl. at-HMGB1, ds-HMGB1 or LPS was abolished by systemic preadministration of argatroban, a thrombin-inhibiting agent, and accelerated by i.pl. co-administered thrombin. Our data clearly indicate that TMα is capable of promoting the thrombin-induced degradation of both at-HMGB1 and ds-HMGB1, and suppresses the allodynia caused by either HMGB1 in a thrombin-dependent manner. Considering the emerging role of HMGB1 in distinct pathological pain models, the present study suggests the therapeutic usefulness of TMα for treatment of intractable and/or persistent pain.

Formalin injection produces long-lasting hypersensitivity with characteristics of neuropathic pain

The purpose of this study was to investigate whether 1%, 2% or 5% formalin injection produce hypersensitivity with characteristics of the neuropathic pain induced by spinal nerve injury. Formalin injection (1%, 2% and 5%) produced concentration-dependent long-lasting (at least 14 days) mechanical allodynia and hyperalgesia in both paws. Likewise, L5/L6 spinal nerve ligation induced allodynia and hyperalgesia in both paws. The intensity of hypersensitivity was greater in the ipsilateral than in the contralateral paw in all models. Systemic gabapentin or morphine completely reduced 1% formalin-induced hypersensitivity. In contrast, both drugs were not able to fully diminish 2–5% formalin- and nerve injury-induced hypersensitivity. Indomethacin produced a significant effect in the chronic 1% formalin test. Conversely, this drug did not modify 2 or 5% formalin- and nerve injury-induced hypersensitivity. Spinal nerve injury and 2–5%, but not 1%, formalin injection enhanced ATF3 protein expression and immunofluorescence in dorsal root ganglia (DRG) in a time-dependent manner. Furthermore, 2–5%, but not 1%, formalin injection or spinal nerve injury also enhanced α2δ−1 subunit protein levels in DRG. Our results suggest that 5% and, at lesser extent, 2% formalin injection produces long-lasting hypersensitivity with a pharmacological and molecular pattern that resembles neuropathic pain induced by spinal nerve ligation.

Spinal CXCL5 contributes to nerve injury-induced neuropathic pain via modulating GSK-3β phosphorylation and activity in rats

BackgroundNeuroinflammation is identified to be crucial in the development of neuropathic pain, whereas definite molecular mechanisms remain obscure. Recently, chemokine CXCL5 is manifested to participate in the inflammatory process of central nervous system, however, little is known about the potential effect of spinal CXCL5 on pathogenesis of pain. This study investigated whether and how CXCL5 and its receptor CXCR2 regulated neuropathic pain in a rat model of chronic constriction injury (CCI) of the sciatic nerves. MethodsRecombinant CXCL5, a neutralizing antibody against CXCL5, selective CXCR2 antagonist SB225002 and GSK-3β inhibitor TDZD-8 were injected intrathecally. PWT and PWL were documented to assess mechanical allodynia and thermal hyperalgesia. Simultaneously, levels of CXCL5 and CXCR2 in spinal dorsal horn were measured by RT-qPCR after nociceptive testing. Western blot was utilized to evaluate spinal GSK-3β expression and phosphorylation. ResultsWe found that CCI engendered rapid and long-lasting mechanical allodynia and thermal hyperalgesia, which was accompanied by dramatical rise of spinal CXCL5 and CXCR2 expression. CCI also caused an increase of pGSK-3β (Tyr216) and a decrease of pGSK-3β (Ser9) without affecting total protein level of GSK-3β. Moreover, spinal blockage of CXCL5/CXCR2 pathway attenuated neuropathic pain and inhibited the enhancement of GSK-3β activity. Also, intrathecal delivery of exogenous CXCL5 dose-dependently induced nociceptive hypersensitivity in naïve rats, which was prevented by the supplemental addition of TDZD-8. ConclusionThese present findings indicate that up-regulation of spinal CXCL5 and CXCR2 is involved in neuropathic pain after nerve injury, through regulating GSK-3β activity in rats.

Spinal activity of interleukin 6 mediates myelin basic protein-induced allodynia

Mechanosensory fibers are enveloped by myelin, a unique multilamellar membrane permitting saltatory neuronal conduction. Damage to myelin is thought to contribute to severe pain evoked by innocuous tactile stimulation (i.e., mechanical allodynia). Our earlier (Liu et al., 2012) and present data demonstrate that a single injection of a myelin basic protein-derived peptide (MBP84-104) into an intact sciatic nerve produces a robust and long-lasting (>30days) mechanical allodynia in female rats. The MBP84-104 peptide represents the immunodominant epitope and requires T cells to maintain allodynia. Surprisingly, only systemic gabapentin (a ligand of voltage-gated calcium channel α2δ1), but not ketorolac (COX inhibitor), lidocaine (sodium channel blocker) or MK801 (NMDA antagonist) reverse allodynia induced by the intrasciatic MBP84-104. The genome-wide transcriptional profiling of the sciatic nerve followed by the bioinformatics analyses of the expression changes identified interleukin (IL)-6 as the major cytokine induced by MBP84-104 in both the control and athymic T cell-deficient nude rats. The intrasciatic MBP84-104 injection resulted in both unilateral allodynia and unilateral IL-6 increase the segmental spinal cord (neurons and astrocytes). An intrathecal delivery of a function-blocking IL-6 antibody reduced the allodynia in part by the transcriptional effects in large-diameter primary afferents in DRG. Our data suggest that MBP regulates IL-6 expression in the nervous system and that the spinal IL-6 activity mediates nociceptive processing stimulated by the MBP epitopes released after damage or disease of the somatosensory nervous system.

Role of myelin auto-antigens in pain: a female connection.

Following injury to the peripheral nerve, pain secondary to low threshold innocuous mechanical stimulation (mechanical allodynia) is commonly observed. Its origins are poorly understood and the pain is often refractory to common analgesic therapeutics. The past two decades has brought major advances in our understanding of the complexity of systems that underlie processing of neuropathic pain (pain arising from damage or disease of the somatosensory system). Beginning with the argument by Robert Galambos in the 1960s that glia played a major regulatory role in neural function, current thinking has evolved to emphasize the major role of glia-mediated neuroinflammation in the development of a chronic pain state. Recent use of unbiased high-throughput genetic and proteomics screening technology in preclinical models of nerve injury associated with pain states, attention has become focused on the role of T cell related genes and adaptive immune pathways (Costigan et al., 2009; Liu et al., 2012). In this perspective, we discuss the organizing principle that following local nerve injury, the ability to evoke a pain state with low-threshold, otherwise innocuous, mechanical stimulation (light touch), represents a local, sex dependent, manifestation of an adaptive immune response mediated by myelin basic protein (MBP).