Tyrosine phosphorylation of the N-Methyl-D-Aspartate receptor 2B subunit in spinal cord contributes to remifentanil-induced postoperative hyperalgesia: the preventive effect of ketamine

Neuropathic pain is a debilitating pain condition that occurs after nerve damage. Such pain is considered to be a reflection of the aberrant excitability of dorsal horn neurons. Emerging lines of evidence indicate that spinal microglia play a crucial role in neuronal excitability and the pathogenesis of neuropathic pain, but the mechanisms underlying neuron-microglia communications in the dorsal horn remain to be fully elucidated. A recent study has demonstrated that platelet-derived growth factor (PDGF) expressed in dorsal horn neurons contributes to neuropathic pain after nerve injury, yet how PDGF produces pain hypersensitivity remains unknown. Here we report an involvement of spinal microglia in PDGF-induced tactile allodynia. A single intrathecal delivery of PDGF B-chain homodimer (PDGF-BB) to naive rats produced a robust and long-lasting decrease in paw withdrawal threshold in a dose-dependent manner. Following PDGF administration, the immunofluorescence for phosphorylated PDGF β-receptor (p-PDGFRβ), an activated form, was markedly increased in the spinal dorsal horn. Interestingly, almost all p-PDGFRβ-positive cells were double-labeled with an antibody for the microglia marker OX-42, but not with antibodies for other markers of neurons, astrocytes and oligodendrocytes. PDGF-stimulated microglia in vivo transformed into a modest activated state in terms of their cell number and morphology. Furthermore, PDGF-BB-induced tactile allodynia was prevented by a daily intrathecal administration of minocycline, which is known to inhibit microglia activation. Moreover, in rats with an injury to the fifth lumbar spinal nerve (an animal model of neuropathic pain), the immunofluorescence for p-PDGFRβ was markedly enhanced exclusively in microglia in the ipsilateral dorsal horn. Together, our findings suggest that spinal microglia critically contribute to PDGF-induced tactile allodynia, and it is also assumed that microglial PDGF signaling may have a role in the pathogenesis of neuropathic pain.     

Persistent At-Level Thermal Hyperalgesia and Tactile Allodynia Accompany Chronic Neuronal and Astrocyte Activation in Superficial Dorsal Horn following Mouse Cervical Contusion Spinal Cord Injury

In humans, sensory abnormalities, including neuropathic pain, often result from traumatic spinal cord injury (SCI). SCI can induce cellular changes in the CNS, termed central sensitization, that alter excitability of spinal cord neurons, including those in the dorsal horn involved in pain transmission. Persistently elevated levels of neuronal activity, glial activation, and glutamatergic transmission are thought to contribute to the hyperexcitability of these dorsal horn neurons, which can lead to maladaptive circuitry, aberrant pain processing and, ultimately, chronic neuropathic pain. Here we present a mouse model of SCI-induced neuropathic pain that exhibits a persistent pain phenotype accompanied by chronic neuronal hyperexcitability and glial activation in the spinal cord dorsal horn. We generated a unilateral cervical contusion injury at the C5 or C6 level of the adult mouse spinal cord. Following injury, an increase in the number of neurons expressing ΔFosB (a marker of chronic neuronal activation), persistent astrocyte activation and proliferation (as measured by GFAP and Ki67 expression), and a decrease in the expression of the astrocyte glutamate transporter GLT1 are observed in the ipsilateral superficial dorsal horn of cervical spinal cord. These changes have previously been associated with neuronal hyperexcitability and may contribute to altered pain transmission and chronic neuropathic pain. In our model, they are accompanied by robust at-level hyperaglesia in the ipsilateral forepaw and allodynia in both forepaws that are evident within two weeks following injury and persist for at least six weeks. Furthermore, the pain phenotype occurs in the absence of alterations in forelimb grip strength, suggesting that it represents sensory and not motor abnormalities. Given the importance of transgenic mouse technology, this clinically-relevant model provides a resource that can be used to study the molecular mechanisms contributing to neuropathic pain following SCI and to identify potential therapeutic targets for the treatment of chronic pathological pain.     

Selective activation of microglia in spinal cord but not higher cortical regions following nerve injury in adult mouse

Neuronal plasticity along the pathway for sensory transmission including the spinal cord and cortex plays an important role in chronic pain, including inflammatory and neuropathic pain. While recent studies indicate that microglia in the spinal cord are involved in neuropathic pain, a systematic study has not been performed in other regions of the central nervous system (CNS). In the present study, we used heterozygous Cx3cr1 ^GFP/+mice to characterize the morphological phenotypes of microglia following common peroneal nerve (CPN) ligation. We found that microglia showed a uniform distribution throughout the CNS, and peripheral nerve injury selectively activated microglia in the spinal cord dorsal horn and related ventral horn. In contrast, microglia was not activated in supraspinal regions of the CNS, including the anterior cingulate cortex (ACC), prefrontal cortex (PFC), primary and secondary somatosensory cortex (S1 and S2), insular cortex (IC), amygdala, hippocampus, periaqueductal gray (PAG) and rostral ventromedial medulla (RVM). Our results provide strong evidence that nerve injury primarily activates microglia in the spinal cord of adult mice, and pain-related cortical plasticity is likely mediated by neurons.     

Direct evidence for spinal cord microglia in the development of a neuropathic pain-like state in mice

The present study was undertaken to further investigate the role of glial cells in the development of the neuropathic pain-like state induced by sciatic nerve ligation in mice. At 7 days after sciatic nerve ligation, the immunoreactivities (IRs) of the specific astrocyte marker glial fibrillary acidic protein (GFAP) and the specific microglial marker OX-42, but not the specific oligodendrocyte marker O4, were increased on the ipsilateral side of the spinal cord dorsal horn in nerve-ligated mice compared with that on the contralateral side. Furthermore, a single intrathecal injection of activated spinal cord microglia, but not astrocytes, caused thermal hyperalgesia in naive mice. Furthermore, 5-bromo-2'-deoxyuridine (BrdU)-positive cells on the ipsilateral dorsal horn of the spinal cord were significantly increased at 7 days after nerve ligation and were highly co-localized with another microglia marker, ionized calcium-binding adaptor molecule 1 (Iba1), but neither with GFAP nor a specific neural nuclei marker, NeuN, in the spinal dorsal horn of nerve-ligated mice. The present data strongly support the idea that spinal cord astrocytes and microglia are activated under the neuropathic pain-like state, and that the proliferated and activated microglia directly contribute to the development of a neuropathic pain-like state in mice.     

A comparative study of the effect of 17β-estradiol and estriol on peripheral pain behavior in rats

Although estradiol has been reported to influence pain sensitivity, the role of estriol (an estradiol metabolite and another widely used female sex hormone) remains unclear. In this study, pain behavior tests, whole-cell patch clamp recording and Western blotting were used to determine whether estriol plays a role in pain signal transduction and transmission. Either systemic or local administration of 17β-estradiol produced a significant rise of mechanical pain threshold, while estriol lacked this effect in normal and ovariectomized (OVX) rats following estriol replacement. Local administration of 17β-estradiol or estriol significantly decreased ATP-induced spontaneous hind-paw withdrawal duration (PWD), which was blocked by an estrogen receptor antagonist, ICI 182, 780. However, systemic application of estriol in normal or OVX rats lacked this similar effect. In cultured dorsal root ganglion neurons, estriol attenuated α,β-methylene ATP-induced transient currents which were blocked by ICI 182, 780. In complete Freund's adjuvant treated (CFA) rats, systemic application of 17β-estradiol or estriol decreased the mechanical pain threshold significantly, but did not change the inflammatory process. Similar effects were observed after estriol replacement in OVX rats. The expression of c-fos in lumbosacral spinal cord dorsal horn (SCDH) was increased significantly by administration of 17β-estradiol but not estriol, and not by estriol replacement in OVX rats. These results suggest that 17β-estradiol but not estriol plays an anti-hyperalgesic role in physiological pain. However, both peripheral 17β-estradiol and estriol play anti-hyperalgesic roles in ATP-induced inflammatory pain. Systemic application of estriol as well as 17β-estradiol plays hyperalgesic roles in CFA-induced chronic pain.     

CCL-1 in the spinal cord contributes to neuropathic pain induced by nerve injury

Cytokines such as interleukins are known to be involved in the development of neuropathic pain through activation of neuroglia. However, the role of chemokine (C-C motif) ligand 1 (CCL-1), a well-characterized chemokine secreted by activated T cells, in the nociceptive transmission remains unclear. We found that CCL-1 was upregulated in the spinal dorsal horn after partial sciatic nerve ligation. Therefore, we examined actions of recombinant CCL-1 on behavioural pain score, synaptic transmission, glial cell function and cytokine production in the spinal dorsal horn. Here we show that CCL-1 is one of the key mediators involved in the development of neuropathic pain. Expression of CCL-1 mRNA was mainly detected in the ipsilateral dorsal root ganglion, and the expression of specific CCL-1 receptor CCR-8 was upregulated in the superficial dorsal horn. Increased expression of CCR-8 was observed not only in neurons but also in microglia and astrocytes in the ipsilateral side. Recombinant CCL-1 injected intrathecally (i.t.) to naive mice induced allodynia, which was prevented by the supplemental addition of N-methyl-𝒟-aspartate (NMDA) receptor antagonist, MK-801. Patch-clamp recordings from spinal cord slices revealed that application of CCL-1 transiently enhanced excitatory synaptic transmission in the substantia gelatinosa (lamina II). In the long term, i.t. injection of CCL-1 induced phosphorylation of NMDA receptor subunit, NR1 and NR2B, in the spinal cord. Injection of CCL-1 also upregulated mRNA level of glial cell markers and proinflammatory cytokines (IL-1β, TNF-α and IL-6). The tactile allodynia induced by nerve ligation was attenuated by prophylactic and chronic administration of neutralizing antibody against CCL-1 and by knocking down of CCR-8. Our results indicate that CCL-1 is one of the key molecules in pathogenesis, and CCL-1/CCR-8 signaling system can be a potential target for drug development in the treatment for neuropathic pain.     

Galectin-3 Inhibition Is Associated with Neuropathic Pain Attenuation after Peripheral Nerve Injury

Neuropathic pain remains a prevalent and persistent clinical problem because it is often poorly responsive to the currently used analgesics. It is very urgent to develop novel drugs to alleviate neuropathic pain. Galectin-3 (gal3) is a multifunctional protein belonging to the carbohydrate-ligand lectin family, which is expressed by different cells. Emerging studies showed that gal3 elicits a pro-inflammatory response by recruiting and activating lymphocytes, macrophages and microglia. In the study we investigated whether gal3 inhibition could suppress neuroinflammation and alleviate neuropathic pain following peripheral nerve injury. We found that L5 spinal nerve ligation (SNL) increases the expression of gal3 in dorsal root ganglions at the mRNA and protein level. Intrathecal administration of modified citrus pectin (MCP), a gal3 inhibitor, reduces gal3 expression in dorsal root ganglions. MCP treatment also inhibits SNL-induced gal3 expression in primary rat microglia. SNL results in an increased activation of autophagy that contributes to microglial activation and subsequent inflammatory response. Intrathecal administration of MCP significantly suppresses SNL-induced autophagy activation. MCP also inhibits lipopolysaccharide (LPS)-induced autophagy in cultured microglia in vitro. MCP further decreases LPS-induced expression of proinflammatory mediators including IL-1β, TNF-α and IL-6 by regulating autophagy. Intrathecal administration of MCP results in adecreased mechanical and cold hypersensitivity following SNL. These results demonstrated that gal3 inhibition is associated with the suppression of SNL-induced inflammatory process andneurophathic pain attenuation.     

Analgesic Activity of Catalpol in Rodent Models of Neuropathic Pain,and Its Spinal Mechanism

Neuropathic pain is a major health issue that represents considerable social and economic burden worldwidely. In this study, we investigated the potential of catalpol, an iridoid glucoside of Rehmannia glutinosa Steud, to alleviate neuropathic pain. The potential analgesic effects of catalpol were evaluated by chronic constriction injury (CCI) and lumbar 5 spinal nerve ligation (L5 SNL) model. In addition, we explored whether catalpol altered the degree of microglia activation and neuroinflammation in rat spinal cord after CCI induction. Repeated administration of catalpol (1, 5, 25, and 125 mg/kg) reversed mechanical allodynia induced by CCI and L5 SNL in a dose-dependent manner in rats. Levels of activated microglia, activated NF-κB, and proinflammatory cytokines (IL-1β, IL-6, TNF-α) in lumber spinal cord were elevated in rats following CCI induction, and catalpol significantly inhibited these effects. Our results demonstrated that catalpol produces significant antinociceptive action in rodent behavioral models of neuropathic pain and that this effect is associated with modulation of neuroinflammation in spinal cord.     

Spatial and temporal relationship between monocyte chemoattractant protein-1 expression and spinal glial activation following peripheral nerve injury

Peripheral nerve injury can induce spinal microglial/astrocyte activation. Substances released by activated glial cells excite spinal nociceptive neurons. Pharmacological disruption of glial activation or antagonism of substances released by activated glia prevent or reverse pain hypersensitivity. It is not known, however, what causes spinal cord glia to shift from a resting to an activated state. In an attempt to understand the potential role of monocyte chemoattractant protein-1 (MCP-1) in triggering spinal glial activation and its contribution to the development of neuropathic pain, we investigated the effect of peripheral nerve injury on MCP-1 expression in dorsal root ganglia (DRG) and the spinal cord, and established its temporal relationship with activation of spinal microglia and astrocytes. We observed that MCP-1 was induced by chronic constriction of the sciatic nerve in DRG sensory neurons, spinal cord motor neurons and in the superficial dorsal horn, ipsilateral to the injury. Neuronal MCP-1 induction was followed by surrounding microglial activation. After peaking at day 7 after injury, MCP-1 levels began to decline rapidly and had returned to baseline by day 150. In contrast, microglial activation peaked by day 14 and declined afterwards to reach a lower, yet significantly raised level beyond day 22 and remained increased until the end of the test period. Astrocyte activation became detectable later, progressed more slowly and also remained increased until the end of the test period, in parallel with a decreased nociceptive threshold. Our results suggest that neuronal MCP-1 may serve as a trigger for spinal microglial activation, which participates in the initiation of neuropathic pain. Delayed, sustained astrocyte activation may participate with microglia in the persistent phase of pain hypersensitivity.     

Interleukin-1β Plays Key Roles in LPA-Induced Amplification of LPA Production in Neuropathic Pain Model

Lysophosphatidic acid (LPA) is a bioactive lipid mediator that exerts a wide range of biological actions. In recent decades, LPA has been demonstrated as an important initiator of neuropathic pain based on the mechanisms of LPA-induced feed-forward LPA amplification. In this study, we examined the possible involvement of interleukin (IL)-1β in such LPA production. Intrathecal (i.t.) LPA injection rapidly increased the expression of IL-1β mRNA in the spinal dorsal horn as early as 0.5 h after injection, and the level reached peak at 2 h. Through a developed quantitative mass spectrometry for detecting LPA species, the elevated levels of 18:1, 16:0, and 18:0 LPA in the spinal dorsal horn were observed at 3 h after 18:1 LPA injection and this elevation was completely blocked by the pretreatment of IL-1β-neutralizing antibody. Moreover, enzyme assay experiments showed that LPA (i.t.) significantly activated calcium-independent phospholipase A₂ (iPLA₂) and cytosolic phospholipase A₂ (cPLA₂) in the spinal dorsal horn at 1 and 2 h, respectively, and these biochemical changes were also significantly inhibited by IL-1β-neutralizing antibody. Similarly, IL-1β-neutralizing antibody reversed LPA-induced neuropathic pain-like behavior. These findings suggest that the early release of IL-1β is involved in LPA-induced amplification of LPA production, which underlies the initial mechanisms of LPA-induced neuropathic pain.     

Analyzing the role of cannabinoids as modulators of Wnt/β-catenin signaling pathway for their use in the management of neuropathic pain

Neuropathic pain is a debilitating form of treatment-resistant chronic pain caused by damage to the nervous system. Cannabinoids have been known for suppressing neuropathic pain by modulating the endo cannabinoid system. Since the canonical Wnt/β-catenin signaling has recently been implicated in pain sensation, we investigated the impact of major cannabinoids (1–6) from the leaves of Cannabis sativa and an epoxy derivative of compound 2, here upon referred to as 2a, on modulating Wnt/β-catenin signaling pathway. The results presented in this study show that compound 1, 2 and 2a exhibited potent inhibitory activity against Wnt/β-catenin pathway in a dose-dependent manner. Compound 2a was seen to inhibit this pathway at slightly lower concentrations than its parent molecule 2, under similar conditions. Taken together, compound 1, 2 and 2a, by virtue of their inhibition of Wnt/β-catenin signaling pathway, could be developed as effective neuroprotective agents for the management of neuropathic pain.     

Neuroprotective effects of genistein in VSC4.1 motoneurons exposed to activated microglial cytokines

Pro-inflammatory cytokines released from activated microglia may be responsible for neuronal damage and resulting motor deficits associated with CNS disorders such as spinal cord injury, Parkinson’s disease, and multiple sclerosis. Estrogen (17β-estradiol) is capable of ameliorating motoneuron death following spinal cord injury, but has a number of deleterious side effects. Genistein (GEN), an estrogen receptor beta agonist and potent antioxidant, may represent an alternative to estrogen in treating neurodegenerative disorders. However, little is known about the neuroprotective effects of GEN. We therefore tested whether GEN would prevent apoptosis in cultured motoneurons following exposure to pro-inflammatory cytokines released from IFN-γ activated microglia. Exposure of ventral spinal cord 4.1 motoneurons to microglial cytokine supernatant in vitro caused significant apoptosis and reduced mitochondrial membrane potential. An increase in reactive oxygen species, intracellular Ca²⁺, calpain, caspases, cytochrome c, and the bax:bcl-2 ratio were also noted. GEN treatment reversed apoptotic death and cellular changes following cytokine exposure and was associated with increased expression of estrogen receptor β suggesting that GEN may promote neuroprotection via receptor-mediated pathways. The addition of ICI 182, 780, an estrogen receptor antagonist following GEN treatment attenuated neuroprotection, suggesting that GEN may act mainly via estrogen receptor β to protect VSC4.1 motoneurons. We conclude that GEN protects cultured ventral spinal cord 4.1 cells from inflammatory insult and thus may represent a potential beneficial therapy in the treatment of neurodegenerative disorders.     

Roles of extracellular signal-regulated protein kinases 5 in spinal microglia and primary sensory neurons for neuropathic pain

Neuropathic pain that occurs after peripheral nerve injury is poorly controlled by current therapies. Increasing evidence shows that mitogen-activated protein kinase (MAPK) play an important role in the induction and maintenance of neuropathic pain. Here we show that activation of extracellular signal-regulated protein kinases 5 (ERK5), also known as big MAPK1, participates in pain hypersensitivity caused by nerve injury. Nerve injury increased ERK5 phosphorylation in spinal microglia and in both damaged and undamaged dorsal root ganglion (DRG) neurons. Antisense knockdown of ERK5 suppressed nerve injury-induced neuropathic pain and decreased microglial activation. Furthermore, inhibition of ERK5 blocked the induction of transient receptor potential channels and brain-derived neurotrophic factor expression in DRG neurons. Our results show that ERK5 activated in spinal microglia and DRG neurons contributes to the development of neuropathic pain. Thus, blocking ERK5 signaling in the spinal cord and primary afferents has potential for preventing pain after nerve damage.     

Fascin-1 Contributes to Neuropathic Pain by Promoting Inflammation in Rat Spinal Cord

Neuropathic pain is a complicated clinical syndrome caused by heterogeneous etiology. Despite the fact that the underlying mechanisms remain elusive, it is well accepted that neuroinflammation plays a critical role in the development of neuropathic pain. Fascin-1, an actin-bundling protein, has been proved to be involved in the processing of diverse biological events including cellular development, immunity, and tumor invasion etc. Recent studies have shown that Fascin-1 participates in antigen presentation and the regulation of pro-inflammatory agents. However, whether Fascin-1 is involved in neuropathic pain has not been reported. In the present study we examined the potential role of Fascin-1 by using a rodent model of chronic constriction injury (CCI). Our results showed that Fascin-1 increased rapidly in dorsal root ganglions (DRG) and spinal cord (SC) after CCI. The increased Fascin-1 widely expressed in DRG, however, it localized predominantly in microglia, seldom in neuron, and hardly in astrocyte in the SC. Intrathecal injection of Fascin-1 siRNA not only suppressed the activation of microglia and the release of pro-inflammatory mediators, but also attenuated the mechanical allodynia and thermal hyperalgesia induced by CCI.     

Pretreatment with intrathecal amitriptyline potentiates anti-hyperalgesic effects of post-injury intra-peritoneal amitriptyline following spinal nerve ligation

Background

Amitriptyline, a tricyclic antidepressant and potent use-dependent blocker of sodium channels, has been shown to attenuate acute and chronic pain in several preclinical modes. The purpose of this study was to investigate whether intrathecal pretreatment with amitriptyline combined with post-injury intra-peritoneal amitriptyline is more effective than post-injury treatment alone on L5 spinal nerve ligation (SNL)-induced neuropathic pain.

Methods

96 adult male Sprague–Dawley rats were allocated into 4 groups: group S, Sham; group L, L5 spinal nerve Ligation with vehicle treatment; group A, SNL and post-injury intra-peritoneal (Abdominal) amitriptyline twice daily?×?3?days; group P, intrathecal Pretreatment with amitriptyline, SNL and intra-peritoneal amitriptyline twice daily?×?3?days. Responses to thermal and mechanical stimuli, as well as sodium channel expression in injured dorsal root ganglion (DRG) and activated glial cells in spinal dorsal horn (SDH) were measured pre-operatively and on post-operative day (POD) 4, 7, 14, 21 and 28.

Results

SNL-evoked hyper-sensitivity responses to thermal and mechanical stimuli, up-regulated Nav1.3 and down-regulated Nav1.8 expression in DRG, and activated microglia and astrocytes in SDH. In group A, intra-peritoneal amitriptyline alone alleviated thermal hypersensitivity on POD7, reversed Nav1.8 and reduced activated microglia on POD14. In group P, intrathecal pretreatment with amitriptyline not only potentiated the effect of intra-peritoneal amitriptyline on thermal hypersensitivity and Nav1.8, but attenuated mechanical hypersensitivity on POD7 and reduced up-regulated Nav1.3 on POD14. Furthermore, this treatment regimen reduced astrocyte activation on POD14.

Conclusions

Concomitant intrathecal pretreatment and post-injury intra-peritoneal amitriptyline was more effective than post-injury treatment alone on attenuation of behavioral hypersensitivity, decrease of activated microglia and astrocytes and dysregulated Nav1.3 and 1.8.     

Microvesicles shed from microglia activated by the P2X7-p38 pathway are involved in neuropathic pain induced by spinal nerve ligation in rats

Microglia are critical in the pathogenesis of neuropathic pain. In this study, we investigated the role of microvesicles (MVs) in neuropathic pain induced by spinal nerve ligation (SNL) in rats. First, we found that MVs shed from microglia were increased in the cerebrospinal fluid and dorsal horn of the spinal cord after SNL. Next, MVs significantly reduced paw withdrawal threshold (PWT) and paw withdrawal latency (PWL). In addition, the P2X7-p38 pathway was related to the bleb of MVs after SNL. Interleukin (IL)-1β was found to be significantly upregulated in the package of MVs, and PWT and PWL increased following inhibition with shRNA-IL-1β. Finally, the amplitude and frequency of spontaneous excitatory postsynaptic currents increased following stimulation with MVs. Our results indicate that the P2X7-p38 pathway is closely correlated with the shedding of MVs from microglia in neuropathic pain, and MVs had a significant effect on neuropathic pain by participating in the interaction between microglia and neurons.     

Aquaporin 1 - a novel player in spinal cord injury

The role of water channel aquaporin 1 (AQP-1) in uninjured or injured spinal cords is unknown. AQP-1 is weakly expressed in neurons and gray matter astrocytes, and more so in white matter astrocytes in uninjured spinal cords, a novel finding. As reported before, AQP-1 is also present in ependymal cells, but most abundantly in small diameter sensory fibers of the dorsal horn. Rat contusion spinal cord injury (SCI) induced persistent and significant four- to eightfold increases in AQP-1 levels at the site of injury (T10) persisting up to 11 months post-contusion, a novel finding. Delayed AQP-1 increases were also found in cervical and lumbar segments, suggesting the spreading of AQP-1 changes over time after SCI. Given that the antioxidant melatonin significantly decreased SCI-induced AQP-1 increases and that hypoxia inducible factor-1α was increased in acutely and chronically injured spinal cords, we propose that chronic hypoxia contributes to persistent AQP-1 increases after SCI. Interestingly; AQP-1 levels were not affected by long-lasting hypertonicity that significantly increased astrocytic AQP-4, suggesting that the primary role of AQP-1 is not regulating isotonicity in spinal cords. Based on our results we propose possible novel roles for AQP-1 in the injured spinal cords: (i) in neuronal and astrocytic swelling, as AQP-1 was increased in all surviving neurons and reactive astrocytes after SCI and (ii) in the development of the neuropathic pain after SCI. We have shown that decreased AQP-1 in melatonin-treated SCI rats correlated with decreased AQP-1 immunolabeling in the dorsal horns sensory afferents, and with significantly decreased mechanical allodynia, suggesting a possible link between AQP-1 and chronic neuropathic pain after SCI.     

Modification of neuropathic pain sensation through microglial ATP receptors

Neuropathic pain that typically develops when peripheral nerves are damaged through surgery, bone compression in cancer, diabetes, or infection is a major factor causing impaired quality of life in millions of people worldwide. Recently, there has been a rapidly growing body of evidence indicating that spinal glia play a critical role in the pathogenesis of neuropathic pain. Accumulating findings also indicate that nucleotides play an important role in neuron-glia communication through P2 purinoceptors. Damaged neurons release or leak nucleotides including ATP and UTP to stimulate microglia through P2 purinoceptors expressing on microglia. It was shown in an animal model of neuropathic pain that microglial P2X₄ and P2X₇ receptors are crucial in pain signaling after peripheral nerve lesion. In this review, we describe the modification of neuropathic pain sensation through microglial P2X₄ and P2X₇, with the possibility of P2Y₆ and P2Y₁₂ involvement.     

P2 receptors and chronic pain

There is abundant evidence that extracellular ATP and other nucleotides have an important role in pain signaling at both the periphery and in the CNS. The focus of attention now is on the possibility that endogenous ATP and its receptor system might be activated in chronic pathological pain states, particularly in neuropathic and inflammatory pain. Neuropathic pain is often a consequence of nerve injury through surgery, bone compression, diabetes or infection. This type of pain can be so severe that even light touching can be intensely painful; unfortunately, this state is generally resistant to currently available treatments. In this review, we summarize the role of ATP receptors, particularly the P2X₄, P2X₃ and P2X₇ receptors, in neuropathic and inflammatory pain. The expression of P2X₄ receptors in the spinal cord is enhanced in spinal microglia after peripheral nerve injury, and blocking pharmacologically and suppressing molecularly P2X₄ receptors produce a reduction of the neuropathic pain behaviour. Understanding the key roles of these ATP receptors may lead to new strategies for the management of intractable chronic pain.