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Kazuhide Inoue - One of the best experts on this subject based on the ideXlab platform.
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p2y12 receptors in spinal microglia are required for neuropathic pain after peripheral Nerve Injury
The Journal of Neuroscience, 2008Co-Authors: Hidetoshi Tozakisaitoh, Makoto Tsuda, Shinichi Kohsaka, Hiroyuki Miyata, Kazuaki Ueda, Kazuhide InoueAbstract:Extracellular nucleotides have been implicated as signaling molecules used by microglia to sense adverse physiological conditions, such as neuronal damage. They act through purinoceptors, especially the G-protein-coupled P2Y receptor P2Y12R. Emerging evidence has indicated that activated spinal microglia responding to Nerve Injury are key cellular intermediaries in the resulting highly debilitating chronic pain state, namely neuropathic pain. However, the role of microglial P2Y12Rs in neuropathic pain remains unknown. Here, we show that the level of P2Y12R mRNA expression was markedly increased in the spinal cord ipsilateral to the Nerve Injury and that this expression was highly restricted to ionized binding calcium adapter molecule 1-positive microglia. An increase in the immunofluorescence of P2Y12R protein in the ipsilateral spinal cord was also observed after Nerve Injury, and P2Y12R-positive cells were double labeled with the microglial marker OX-42. Blocking spinal P2Y12R by the intrathecal administration of its antagonist AR-C69931MX prevented the development of tactile allodynia (pain hypersensitivity to innocuous stimuli), a hallmark of neuropathic pain syndrome. Furthermore, mice lacking P2ry12 (P2ry12−/−) displayed impaired tactile allodynia after Nerve Injury without any change in basal mechanical sensitivity. Moreover, a single intrathecal administration of AR-C69931MX or oral administration of clopidogrel (a P2Y12R blocker clinically in use) to Nerve-injured rats produced a striking alleviation of existing tactile allodynia. Together, our findings indicate that activation of P2Y12Rs in spinal microglia may be a critical event in the pathogenesis of neuropathic pain and suggest that blocking microglial P2Y12R might be a viable therapeutic strategy for treating neuropathic pain.
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activation of p38 mitogen activated protein kinase in spinal hyperactive microglia contributes to pain hypersensitivity following peripheral Nerve Injury
Glia, 2004Co-Authors: Makoto Tsuda, Yukari Shigemotomogami, Schuichi Koizumi, Akito Mizokoshi, Kazuhide InoueAbstract:Neuropathic pain is an expression of pathological operation of the nervous system, which commonly results from Nerve Injury and is characterized by pain hypersen- sitivity to innocuous stimuli, a phenomenon known as tactile allodynia. The mechanisms by which Nerve Injury creates tactile allodynia have remained largely unknown. We report that the development of tactile allodynia following Nerve Injury requires activation of p38 mito- gen-activated protein kinase (p38MAPK), a member of the MAPK family, in spinal micro- glia. We found that immunofluorescence and protein levels of the dually phosphorylated active form of p38MAPK (phospho-p38MAPK) were increased in the dorsal horn ipsilateral to spinal Nerve Injury. Interestingly, the phospho-p38MAPK immunofluorescence in the dorsal horn was found exclusively in microglia, but not in neurons or astrocytes. The level of phospho-p38MAPK immunofluorescence in individual microglial cells was much higher in the hyperactive phenotype in the ipsilateral dorsal horn than the resting one in the contralateral side. Intrathecal administration of the p38MAPK inhibitor, 4-(4-fluorophenyl)- 2-(4-methylsulfonylphenyl)-5-(4-pyridyl)-1H-imidazole (SB203580), suppresses develop- ment of the Nerve Injury-induced tactile allodynia. Taken together, our results demonstrate that Nerve Injury-induced pain hypersensitivity depends on activation of the p38MAPK signaling pathway in hyperactive microglia in the dorsal horn following peripheral Nerve Injury. © 2003 Wiley-Liss, Inc.
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p2x4 receptors induced in spinal microglia gate tactile allodynia after Nerve Injury
Nature, 2003Co-Authors: Makoto Tsuda, Yukari Shigemotomogami, Schuichi Koizumi, Akito Mizokoshi, Shinichi Kohsaka, Michael W Salter, Kazuhide InoueAbstract:Pain after Nerve damage is an expression of pathological operation of the nervous system1,2, one hallmark of which is tactile allodynia—pain hypersensitivity evoked by innocuous stimuli. Effective therapy for this pain is lacking, and the underlying mechanisms are poorly understood. Here we report that pharmacological blockade of spinal P2X4 receptors (P2X4Rs)3,4,5,6,7, a subtype of ionotropic ATP receptor8, reversed tactile allodynia caused by peripheral Nerve Injury without affecting acute pain behaviours in naive animals. After Nerve Injury, P2X4R expression increased strikingly in the ipsilateral spinal cord, and P2X4Rs were induced in hyperactive microglia but not in neurons or astrocytes. Intraspinal administration of P2X4R antisense oligodeoxynucleotide decreased the induction of P2X4Rs and suppressed tactile allodynia after Nerve Injury. Conversely, intraspinal administration of microglia in which P2X4Rs had been induced and stimulated, produced tactile allodynia in naive rats. Taken together, our results demonstrate that activation of P2X4Rs in hyperactive microglia is necessary for tactile allodynia after Nerve Injury and is sufficient to produce tactile allodynia in normal animals. Thus, blocking P2X4Rs in microglia might be a new therapeutic strategy for pain induced by Nerve Injury.
Clifford J Woolf - One of the best experts on this subject based on the ideXlab platform.
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blocking caspase activity prevents transsynaptic neuronal apoptosis and the loss of inhibition in lamina ii of the dorsal horn after peripheral Nerve Injury
The Journal of Neuroscience, 2005Co-Authors: Joachim Scholz, Isabelle Decosterd, Kimberly A Moore, Tatsuro Kohno, Daniel C Broom, Dong Ho Youn, Charles D Mills, Marc R Suter, Richard E Coggeshall, Clifford J WoolfAbstract:We show that transsynaptic apoptosis is induced in the superficial dorsal horn (laminas I-III) of the spinal cord by three distinct partial peripheral Nerve lesions: spared Nerve Injury, chronic constriction, and spinal Nerve ligation. Ongoing activity in primary afferents of the injured Nerve and glutamatergic transmission cause a caspase-dependent degeneration of dorsal horn neurons that is slow in onset and persists for several weeks. Four weeks after spared Nerve Injury, the cumulative loss of dorsal horn neurons, determined by stereological analysis, is >20%. GABAergic inhibitory interneurons are among the neurons lost, and a marked decrease in inhibitory postsynaptic currents of lamina II neurons coincides with the induction of apoptosis. Blocking apoptosis with the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (zVAD) prevents the loss of GABAergic interneurons and the reduction of inhibitory currents. Partial peripheral Nerve Injury results in pain-like behavioral changes characterized by hypersensitivity to tactile or cold stimuli. Treatment with zVAD, which has no intrinsic analgesic properties, attenuates this neuropathic pain-like syndrome. Preventing Nerve Injury-induced apoptosis of dorsal horn neurons by blocking caspase activity maintains inhibitory transmission in lamina II and reduces pain hypersensitivity.
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partial peripheral Nerve Injury promotes a selective loss of gabaergic inhibition in the superficial dorsal horn of the spinal cord
The Journal of Neuroscience, 2002Co-Authors: Kimberly A Moore, Tatsuro Kohno, Laurie A Karchewski, Joachim Scholz, Hiroshi Baba, Clifford J WoolfAbstract:To clarify whether inhibitory transmission in the superficial dorsal horn of the spinal cord is reduced after peripheral Nerve Injury, we have studied synaptic transmission in lamina II neurons of an isolated adult rat spinal cord slice preparation after complete sciatic Nerve transection (SNT), chronic constriction Injury (CCI), or spared Nerve Injury (SNI). Fast excitatory transmission remains intact after all three types of Nerve Injury. In contrast, primary afferent-evoked IPSCs are substantially reduced in incidence, magnitude, and duration after the two partial Nerve injuries, CCI and SNI, but not SNT. Pharmacologically isolated GABAAreceptor-mediated IPSCs are decreased in the two partial Nerve Injury models compared with naive animals. An analysis of unitary IPSCs suggests that presynaptic GABA release is reduced after CCI and SNI. Partial Nerve Injury also decreases dorsal horn levels of the GABA synthesizing enzyme glutamic acid decarboxylase (GAD) 65 kDa ipsilateral to the Injury and induces neuronal apoptosis, detected by terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling staining in identified neurons. Both of these mechanisms could reduce presynaptic GABA levels and promote a functional loss of GABAergic transmission in the superficial dorsal horn.
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spared Nerve Injury an animal model of persistent peripheral neuropathic pain
Pain, 2000Co-Authors: Isabelle Decosterd, Clifford J WoolfAbstract:Abstract Peripheral neuropathic pain is produced by multiple etiological factors that initiate a number of diverse mechanisms operating at different sites and at different times and expressed both within, and across different disease states. Unraveling the mechanisms involved requires laboratory animal models that replicate as far as possible, the different pathophysiological changes present in patients. It is unlikely that a single animal model will include the full range of neuropathic pain mechanisms. A feature of several animal models of peripheral neuropathic pain is partial denervation. In the most frequently used models a mixture of intact and injured fibers is created by loose ligation of either the whole (Bennett GJ, Xie YK. A peripheral mononeuropathy in rat that produces disorders of pain sensation like those seen in man. Pain 1988;33:87–107) or a tight ligation of a part (Seltzer Z, Dubner R, Shir Y. A novel behavioral model of neuropathic pain disorders produced in rats by partial sciatic Nerve Injury. Pain 1990;43:205–218) of a large peripheral Nerve, or a tight ligation of an entire spinal segmental Nerve (Kim SH, Chung JM. An experimental model for peripheral neuropathy produced by segmental spinal Nerve ligation in the rat. Pain 1992;50:355–363). We have developed a variant of partial denervation, the spared Nerve Injury model. This involves a lesion of two of the three terminal branches of the sciatic Nerve (tibial and common peroneal Nerves) leaving the remaining sural Nerve intact. The spared Nerve Injury model differs from the Chung spinal segmental Nerve, the Bennett chronic constriction Injury and the Seltzer partial sciatic Nerve Injury models in that the co-mingling of distal intact axons with degenerating axons is restricted, and it permits behavioral testing of the non-injured skin territories adjacent to the denervated areas. The spared Nerve Injury model results in early ( 6 months), robust (all animals are responders) behavioral modifications. The mechanical (von Frey and pinprick) sensitivity and thermal (hot and cold) responsiveness is increased in the ipsilateral sural and to a lesser extent saphenous territories, without any change in heat thermal thresholds. Crush Injury of the tibial and common peroneal Nerves produce similar early changes, which return, however to baseline at 7–9 weeks. The spared Nerve Injury model may provide, therefore, an additional resource for unraveling the mechanisms responsible for the production of neuropathic pain.
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peripheral Nerve Injury triggers central sprouting of myelinated afferents
Nature, 1992Co-Authors: Clifford J Woolf, Peter Shortland, Richard E CoggeshallAbstract:THE central terminals of primary afferent neurons are topographically highly ordered in the spinal cord1. Peripheral receptor sensitivity is reflected by dorsal horn laminar location: low-threshold mechanoreceptors terminate in laminae III and IV (refs 2, 3) and high-threshold nociceptors in laminae I, II and V (refs 4, 5). Unmyelinated C fibres, most of which are nociceptors6, terminate predominantly in lamina II (refs 5,7). There is therefore an anatomical framework for the transfer of specific inputs to localized subsets of dorsal horn neurons. This specificity must contribute to the relationship between a low-intensity stimulus and an innocuous sensation and a noxious stimulus and pain. We now show that after peripheral Nerve Injury the central terminals of axotomized myelinated afferents, including the large Aβ fibres, sprout into lamina II. This structural reorganization in the adult central nervous system may contribute to the development of the pain mediated by A-fibres that can follow Nerve lesions in humans8,9.
Makoto Tsuda - One of the best experts on this subject based on the ideXlab platform.
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p2y12 receptors in spinal microglia are required for neuropathic pain after peripheral Nerve Injury
The Journal of Neuroscience, 2008Co-Authors: Hidetoshi Tozakisaitoh, Makoto Tsuda, Shinichi Kohsaka, Hiroyuki Miyata, Kazuaki Ueda, Kazuhide InoueAbstract:Extracellular nucleotides have been implicated as signaling molecules used by microglia to sense adverse physiological conditions, such as neuronal damage. They act through purinoceptors, especially the G-protein-coupled P2Y receptor P2Y12R. Emerging evidence has indicated that activated spinal microglia responding to Nerve Injury are key cellular intermediaries in the resulting highly debilitating chronic pain state, namely neuropathic pain. However, the role of microglial P2Y12Rs in neuropathic pain remains unknown. Here, we show that the level of P2Y12R mRNA expression was markedly increased in the spinal cord ipsilateral to the Nerve Injury and that this expression was highly restricted to ionized binding calcium adapter molecule 1-positive microglia. An increase in the immunofluorescence of P2Y12R protein in the ipsilateral spinal cord was also observed after Nerve Injury, and P2Y12R-positive cells were double labeled with the microglial marker OX-42. Blocking spinal P2Y12R by the intrathecal administration of its antagonist AR-C69931MX prevented the development of tactile allodynia (pain hypersensitivity to innocuous stimuli), a hallmark of neuropathic pain syndrome. Furthermore, mice lacking P2ry12 (P2ry12−/−) displayed impaired tactile allodynia after Nerve Injury without any change in basal mechanical sensitivity. Moreover, a single intrathecal administration of AR-C69931MX or oral administration of clopidogrel (a P2Y12R blocker clinically in use) to Nerve-injured rats produced a striking alleviation of existing tactile allodynia. Together, our findings indicate that activation of P2Y12Rs in spinal microglia may be a critical event in the pathogenesis of neuropathic pain and suggest that blocking microglial P2Y12R might be a viable therapeutic strategy for treating neuropathic pain.
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activation of p38 mitogen activated protein kinase in spinal hyperactive microglia contributes to pain hypersensitivity following peripheral Nerve Injury
Glia, 2004Co-Authors: Makoto Tsuda, Yukari Shigemotomogami, Schuichi Koizumi, Akito Mizokoshi, Kazuhide InoueAbstract:Neuropathic pain is an expression of pathological operation of the nervous system, which commonly results from Nerve Injury and is characterized by pain hypersen- sitivity to innocuous stimuli, a phenomenon known as tactile allodynia. The mechanisms by which Nerve Injury creates tactile allodynia have remained largely unknown. We report that the development of tactile allodynia following Nerve Injury requires activation of p38 mito- gen-activated protein kinase (p38MAPK), a member of the MAPK family, in spinal micro- glia. We found that immunofluorescence and protein levels of the dually phosphorylated active form of p38MAPK (phospho-p38MAPK) were increased in the dorsal horn ipsilateral to spinal Nerve Injury. Interestingly, the phospho-p38MAPK immunofluorescence in the dorsal horn was found exclusively in microglia, but not in neurons or astrocytes. The level of phospho-p38MAPK immunofluorescence in individual microglial cells was much higher in the hyperactive phenotype in the ipsilateral dorsal horn than the resting one in the contralateral side. Intrathecal administration of the p38MAPK inhibitor, 4-(4-fluorophenyl)- 2-(4-methylsulfonylphenyl)-5-(4-pyridyl)-1H-imidazole (SB203580), suppresses develop- ment of the Nerve Injury-induced tactile allodynia. Taken together, our results demonstrate that Nerve Injury-induced pain hypersensitivity depends on activation of the p38MAPK signaling pathway in hyperactive microglia in the dorsal horn following peripheral Nerve Injury. © 2003 Wiley-Liss, Inc.
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p2x4 receptors induced in spinal microglia gate tactile allodynia after Nerve Injury
Nature, 2003Co-Authors: Makoto Tsuda, Yukari Shigemotomogami, Schuichi Koizumi, Akito Mizokoshi, Shinichi Kohsaka, Michael W Salter, Kazuhide InoueAbstract:Pain after Nerve damage is an expression of pathological operation of the nervous system1,2, one hallmark of which is tactile allodynia—pain hypersensitivity evoked by innocuous stimuli. Effective therapy for this pain is lacking, and the underlying mechanisms are poorly understood. Here we report that pharmacological blockade of spinal P2X4 receptors (P2X4Rs)3,4,5,6,7, a subtype of ionotropic ATP receptor8, reversed tactile allodynia caused by peripheral Nerve Injury without affecting acute pain behaviours in naive animals. After Nerve Injury, P2X4R expression increased strikingly in the ipsilateral spinal cord, and P2X4Rs were induced in hyperactive microglia but not in neurons or astrocytes. Intraspinal administration of P2X4R antisense oligodeoxynucleotide decreased the induction of P2X4Rs and suppressed tactile allodynia after Nerve Injury. Conversely, intraspinal administration of microglia in which P2X4Rs had been induced and stimulated, produced tactile allodynia in naive rats. Taken together, our results demonstrate that activation of P2X4Rs in hyperactive microglia is necessary for tactile allodynia after Nerve Injury and is sufficient to produce tactile allodynia in normal animals. Thus, blocking P2X4Rs in microglia might be a new therapeutic strategy for pain induced by Nerve Injury.
Eunkyeong Jo - One of the best experts on this subject based on the ideXlab platform.
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nadph oxidase 2 derived reactive oxygen species in spinal cord microglia contribute to peripheral Nerve Injury induced neuropathic pain
Proceedings of the National Academy of Sciences of the United States of America, 2010Co-Authors: Eunkyeong Jo, Melvin I SimonAbstract:Increasing evidence supports the notion that spinal cord microglia activation plays a causal role in the development of neuropathic pain after peripheral Nerve Injury; yet the mechanisms for microglia activation remain elusive. Here, we provide evidence that NADPH oxidase 2 (Nox2)-derived ROS production plays a critical role in Nerve Injury-induced spinal cord microglia activation and subsequent pain hypersensitivity. Nox2 expression was induced in dorsal horn microglia immediately after L5 spinal Nerve transection (SNT). Studies using Nox2-deficient mice show that Nox2 is required for SNT-induced ROS generation, microglia activation, and proinflammatory cytokine expression in the spinal cord. SNT-induced mechanical allodynia and thermal hyperalgesia were similarly attenuated in Nox2-deficient mice. In addition, reducing microglial ROS level via intrathecal sulforaphane administration attenuated mechanical allodynia and thermal hyperalgesia in SNT-injured mice. Sulforaphane also inhibited SNT-induced proinflammatory gene expression in microglia, and studies using primary microglia indicate that ROS generation is required for proinflammatory gene expression in microglia. These studies delineate a pathway involving Nerve damage leading to microglial Nox2-generated ROS, resulting in the expression of proinflammatory cytokines that are involved in the initiation of neuropathic pain.
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a critical role of toll like receptor 2 in Nerve Injury induced spinal cord glial cell activation and pain hypersensitivity
Journal of Biological Chemistry, 2007Co-Authors: Eunkyeong Jo, Seyoung Choi, Kyungpyo Park, Shizuo Akira, Heung Sik Na, Seog Bae OhAbstract:Abstract The activation of spinal cord glial cells has been implicated in the development of neuropathic pain upon peripheral Nerve Injury. The molecular mechanisms underlying glial cell activation, however, have not been clearly elucidated. In this study, we found that damaged sensory neurons induce the expression of tumor necrosis factor-α, interleukin-1β, interleukin-6, and inducible nitric-oxide synthase genes in spinal cord glial cells, which is implicated in the development of neuropathic pain. Studies using primary glial cells isolated from toll-like receptor 2 knock-out mice indicate that damaged sensory neurons activate glial cells via toll-like receptor 2. In addition, behavioral studies using toll-like receptor 2 knock-out mice demonstrate that the expression of toll-like receptor 2 is required for the induction of mechanical allodynia and thermal hyperalgesia due to spinal Nerve axotomy. The Nerve Injury-induced spinal cord microglia and astrocyte activation is reduced in the toll-like receptor 2 knock-out mice. Similarly, the Nerve Injury-induced pro-inflammatory gene expression in the spinal cord is also reduced in the toll-like receptor 2 knock-out mice. These data demonstrate that toll-like receptor 2 contributes to the Nerve Injury-induced spinal cord glial cell activation and subsequent pain hypersensitivity.
Marzia Malcangio - One of the best experts on this subject based on the ideXlab platform.
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role of spinal microglia in rat models of peripheral Nerve Injury and inflammation
European Journal of Pain, 2007Co-Authors: Anna K Clark, Clive Gentry, Elizabeth J Bradbury, Stephen B Mcmahon, Marzia MalcangioAbstract:Mounting evidence supports the hypothesis that spinal microglia modulate the development and maintenance of some chronic pain states. Here we examined the role of spinal microglia following both peripheral inflammatory insult and peripheral Nerve Injury. We observed significant ipsilateral dorsal horn microglia activation 2 weeks after Injury and bilateral activation 50 days following Nerve Injury as well as 24 h following intraplantar zymosan but not intraplantar complete Freund's adjuvant (CFA). Ipsilateral but not contralateral microglia activation was associated with hind paw mechanical hyperalgesia. Spinal injection of the glial metabolic inactivator fluorocitrate attenuated ipsilateral hyperalgesia and bilateral spinal microglia activation after peripheral Nerve Injury. Intrathecal fluorocitrate reversed hyperalgesia after intraplantar zymosan and produced no reversal of CFA-induced hyperalgesia. These data suggest a role for spinal glia in the persistence of mechanical hyperalgesia following peripheral Nerve Injury. However, activation of spinal microglia contralaterally did not correlate to nociception. Furthermore, it would appear that the time course of microglia activation and their contribution to inflammatory pain is dependent on the inflammatory stimulus administered.
