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Spinal cord injury (SCI) often causes sensitization of spinal dorsal horn excitatory neurons via disruption of inhibitory output that results in exaggerated nociceptive transmission. Gamma-aminobutyric acid (GABA) is a major inhibitory neurotransmitter and thought to be critical for spinal inhibitory synaptic transmission. However, SCI causes hypofunctional GABAergic inhibitory output via multiple mechanisms, including loss of GABAergic neurons, downregulation of GABA synthesis enzymes, decrease of primary afferent innervation into GABAergic neurons, and shifts of Cl− gradient in the spinal dorsal horn. These disruptions of GABAergic inhibitory output critically contribute to neuronal hyperexcitability in the spinal dorsal horn and chronic neuropathic pain states following SCI. In this book chapter, we focus on spinal GABAergic mechanisms of chronic neuropathic pain development following SCI in rodent animals.

Spinal cord injury (SCI) causes maladaptive changes to nociceptive synaptic circuits within the injured spinal cord. Changes also occur at remote regions including the brain stem, limbic system, cortex, and dorsal root ganglia. These maladaptive nociceptive synaptic circuits frequently cause neuronal hyperexcitability in the entire nervous system and enhance nociceptive transmission, resulting in chronic central neuropathic pain following SCI. The underlying mechanism of chronic neuropathic pain depends on the neuroanatomical structures and electrochemical communication between pre- and postsynaptic neuronal membranes, and propagation of synaptic transmission in the ascending pain pathways. In the nervous system, neurons are the only cell type that transmits nociceptive signals from peripheral receptors to supraspinal systems due to their neuroanatomical and electrophysiological properties. However, the entire range of nociceptive signaling is not mediated by any single neuron. Current literature describes regional studies of electrophysiological or neurochemical mechanisms for enhanced nociceptive transmission post-SCI, but few studies report the electrophysiological, neurochemical, and neuroanatomical changes across the entire nervous system following a regional SCI. We, along with others, have continuously described the enhanced nociceptive transmission in the spinal dorsal horn, brain stem, thalamus, and cortex in SCI-induced chronic central neuropathic pain condition, respectively. Thus, this review summarizes the current understanding of SCI-induced neuronal hyperexcitability and maladaptive nociceptive transmission in the entire nervous system that contributes to chronic central neuropathic pain.

In the present study, we examined superoxide-mediated excitatory nociceptive transmission on at-level neuropathic pain following spinal thoracic 10 contusion injury (SCI) in male Sprague Dawley rats. Methods: Mechanical sensitivity at body trunk, neuronal firing activity, and expression of superoxide marker/ionotropic glutamate receptors (iGluRs)/CamKII were measured in the T7/8 dorsal horn, respectively. Results: Topical treatment of superoxide donor t-BOOH (0.4 mg/kg) increased neuronal firing rates and pCamKII expression in the naïve group, whereas superoxide scavenger Tempol (1 mg/kg) and non-specific ROS scavenger PBN (3 mg/kg) decreased firing rates in the SCI group (*p &lt, 0.05). SCI showed increases of iGluRs-mediated neuronal firing rates and pCamKII expression (*p &lt, 0.05), however, t-BOOH treatment did not show significant changes in the naïve group. The mechanical sensitivity at the body trunk in the SCI group (6.2 ± 0.5) was attenuated by CamKII inhibitor KN-93 (50 mg, 3.9 ± 0.4) or Tempol (1 mg, 4 ± 0.4) treatment (*p &lt, 0.05). In addition, the level of superoxide marker Dhet showed significant increase in SCI rats compared to the sham group (11.7 ± 1.7 vs. 6.6 ± 1.5, *p &lt, 0.05). Conclusions: Superoxide and the pCamKII pathway contribute to chronic at-level neuropathic pain without involvement of iGluRs following SCI.

Invasive bioelectrodes are widely used as an effective treatment for several acute and chronic diseases. In earlier work using high surface area invasive porous bioelectrodes evaluated in an animal model of alcoholism withdrawal, we demonstrated significantly improved electrophysiological and behavioral responses. In this study, we further modify the surface of these invasive porous bioelectrodes with noble metal (Ag, Au, Pt) nanoparticles. Compared to both conventional and porous bioelectrodes, noble metal sensitized invasive porous bioelectrodes show markedly increased low threshold (LT) and wide dynamic range (WDR) neuronal activity. In particular, Pt-sensitized invasive porous bioelectrodes show the highest WDR neuronal activity only upon insertion. In addition, Ag-sensitized invasive porous bioelectrodes, whose surface area is about 37 times greater than that of conventional bioelectrodes, show improved electrochemical properties with higher LT and WDR neuronal activity when stimulated. In an animal model of chronic alcoholism, using normal and alcohol-treated Sprague-Dawley (SD) rats evaluated with the elevated plus maze (EPM) test, the Ag-sensitized invasive porous bioelectrodes show about 20% higher open arms time. These results suggest that these noble metal-sensitized invasive bioelectrodes may offer improved therapeutic outcomes for the treatment of chronic alcoholism, and given these enhanced electrophysiological properties, for other conditions as well.

Drug addiction is a chronic relapsing disease, which causes serious social and economic problems. The most important trial for the successful treatment of drug addiction is to prevent the high rate of relapse to drug-seeking behaviors. Opponent process as a motivational theory with excessive drug seeking in the negative reinforcement of drug dependence reflects both loss of brain reward system and recruitment of brain stress system. The negative emotional state produced by brain stress system during drug withdrawal might contribute to the intense drug craving and drive drug-seeking behaviors via negative reinforcement mechanisms. Decrease in dopamine neurotransmission in the nucleus accumbens and recruitment of corticotropin-releasing factor in the extended amygdala are hypothesized to be implicated in mediating this motivated behavior. Also, a brain stress response system is hypothesized to increase drug craving and contribute to relapse to drug-seeking behavior during the preoccupation and anticipation stage of dependence caused by the exposure to stress characterized as the nonspecific responses to any demands on the body. Acupuncture has proven to be effective for reducing drug addiction and stress-related psychiatric disorders, such as anxiety and depression. Furthermore, acupuncture has been shown to correct reversible brain malfunctions by regulating drug addiction and stress-related neurotransmitters. Accordingly, it seems reasonable to propose that acupuncture attenuates relapse to drug-seeking behavior through inhibition of stress response. In this review, a brief description of stress in relapse to drug-seeking behavior and the effects of acupuncture were presented.

Methamphetamine (METH) increases metabolic neuronal activity in the mesolimbic dopamine (DA) system and mediates the reinforcing effect. To explore the underlying mechanism of acupuncture intervention in reducing METH-induced behaviors, we investigated the effect of acupuncture on locomotor activity, ultrasonic vocalizations, extracellular DA release in the nucleus accumbens (NAcs) using fast-scan cyclic voltammetry and alterations of brain temperature (an indicator of local brain metabolic activity) produced by METH administration. When acupuncture was applied to HT7, but not TE4, both locomotor activity and 50-kHz ultrasonic vocalizations were suppressed in METH-treated rats. Acupuncture at HT7 attenuated the enhancement of electrically stimulated DA release in the NAc of METH-treated rats. Systemic injection of METH produced a sustained increase in NAc temperature, which was reversed by the DA D1 receptor antagonist SCH 23390 or acupuncture at HT7. Acupuncture inhibition of METH-induced NAc temperature was prevented by pre-treatment with a group II metabotropic glutamate receptors (mGluR2/3) antagonist EGLU into the NAc or mimicked by injection of an mGluR2/3 agonist DCG-IV into the NAc. These results suggest that acupuncture reduces extracellular DA release and metabolic neuronal activity in the NAc through activation of mGluR2/3 and suppresses METH-induced affective states and locomotor behavior.

There is growing public interest in alternative approaches to addiction treatment and scientific interest in elucidating the neurobiological underpinnings of acupuncture. Our previous studies showed that acupuncture at a specific Shenmen (HT7) points reduced dopamine (DA) release in the nucleus accumbens (NAc) induced by drugs of abuse. The present study was carried out to evaluate the effects of HT7 acupuncture on γ-aminobutyric acid (GABA) neuronal activity in the ventral tegmental area (VTA) and the reinstatement of cocaine-seeking behavior. Using microdialysis and in vivo single-unit electrophysiology, we evaluated the effects of HT7 acupuncture on VTA GABA and NAc DA release and VTA GABA neuronal activity in rats. Using a within-session reinstatement paradigm in rats self-administering cocaine, we evaluated the effects of HT7 stimulation on cocaine-primed reinstatement. Acupuncture at HT7 significantly reduced cocaine suppression of GABA release and GABA neuron firing rates in the VTA. HT7 acupuncture attenuated cocaine-primed reinstatement, which was blocked by VTA infusions of the selective GABAB receptor antagonist 2-hydroxysaclofen. HT7 stimulation significantly decreased acute cocaine-induced DA release in the NAc, which was also blocked by 2-hydroxysaclofen. HT7 acupuncture also attenuated cocaine-induced sensitization of extracellular DA levels in the NAc. Moreover, HT7 acupuncture reduced both locomotor activity and neuronal activation in the NAc induced by acute cocaine in a needle-penetration depth-dependent fashion. These results suggest that acupuncture may suppress cocaine-induced DA release in the NAc and cocaine-seeking behavior through activation of VTA GABA neurons. Acupuncture may be an effective therapy to reduce cocaine relapse by enhancing GABAergic inhibition in the VTA.

We provide evidence for the effectiveness of acupuncture in an animal model of alcohol dependence., A withdrawal-associated impairment in β-endorphin neurotransmission in the arcuate nucleus (ARC) of the hypothalamus is associated with alcohol dependence characterized by a chronic relapsing disorder. Although acupuncture activates β-endorphin neurons in the ARC projecting to the nucleus accumbens (NAc), a role for ARC β-endorphin neurons in alcohol dependence and acupuncture effects has not been examined. Here, we show that acupuncture at Shenmen (HT7) points attenuates behavioral manifestation of alcohol dependence by activating endorphinergic input to the NAc from the ARC. Acupuncture attenuated ethanol withdrawal tremor, anxiety-like behaviors, and ethanol self-administration in ethanol-dependent rats, which are mimicked by local injection of β-endorphin into the NAc. Acupuncture also reversed the decreased β-endorphin levels in the NAc and a reduction of neuronal activity in the ARC during ethanol withdrawal. These results suggest that acupuncture may provide a novel, potential treatment strategy for alcohol use disorder by direct activation of the brain pathway.

Background: The present study was designed to examine the functional recovery following spinal cord injury (SCI) by adjusting the parameters of impact force and dwell-time using the Infinite Horizon (IH) impactor device. Methods: Sprague-Dawley rats (225.240 g) were divided into eight injury groups based on force of injury (Kdyn) and dwell time (seconds), indicated as Force-Dwell time: 150-4, 150-3, 150-2, 150-1, 150-0, 200-0, 90-2 and sham controls, respectively. Results: After T10 SCI, higher injury force produced greater spinal cord displacement (P ＜ 0.05) and showed a significant correlation (r = 0.813) between the displacement and the force (P ＜ 0.05). In neuropathic pain-like behavior, the percent of paw withdrawals scores in the hindpaw for the 150-4, 150-3, 150-2, 150-1 and the 200-0 injury groups were significantly lowered compared with sham controls (P ＜ 0.05). The recovery of locomotion had a significant within-subjects effect of time (P ＜ 0.05) and the 150-0 group had increased recovery compared to other groups (P ＜ 0.05). In addition, the 200-0 and the 90-2 recovered significantly better than all the 150 kdyn impact groups that included a dwell-time (P ＜ 0.05). In recovery of spontaneous bladder function, the 150-4 injury group took significantly longer recovery time whereas the 150-0 and the 90-2 groups had the shortest recovery times. Conclusions: The present study demonstrates SCI parameters optimize development of mechanical allodynia and other pathological outcomes. (Korean J Pain 2016; 29: 86-95)

Previous studies revealed that acupuncture suppressed both morphine self-administration and morphine-seeking behavior after abstinence. Based on these results, this study examined whether acupuncture attenuated morphine-craving under a progressive ratio (PR) schedule and investigated the possible neuronal mechanism. Male Sprague-Dawley rats were trained to self-administer morphine (0.5 mg/kg) at a fixed ratio for 9 days, and rats who achieved stable infusion were switched to a PR schedule. When animals had taken no more morphine for 1 hour, the number of infusions was defined as the break point (BP). After PR training, animals that had established a stable BP received acupuncture the next day. Acupuncture was applied for 1 minute immediately before the test session. Bicuculline (1.0 mg/kg) and SCH 50911 (2.0 mg/kg) were given 30 minutes prior to acupuncture. The c-Fos levels in the ventral tegmental area (VTA) and the nucleus accumbens (NAc) were examined. Acupuncture at SI5 reduced the BP significantly. Moreover, the effects of acupuncture were blocked by either bicuculline or SCH 50911. Immunofluorescence revealed that acupuncture at SI5 decreased c-Fos expressions in the VTA and the NAc. This study demonstrates that acupuncture at SI5 is effective for the treatment of morphine-craving and that this effect is mediated via the GABA pathway.

Spinal cord injury (SCI) frequently results in chronic neuropathic pain (CNP). However, the understanding of brain neural circuits in CNP modulation is unclear. The present study examined the changes of ventral tegmental area (VTA) putative GABAergic and dopaminergic neuronal activity with CNP attenuation in rats. SCI was established by T10 clip compression injury (35 g, 1 min) in rats, and neuropathic pain behaviors, in vivo extracellular single-cell recording of putative VTA gamma-aminobutyric acid (GABA)/dopamine neurons, extracellular GABA level, glutamic acid decarboxylase (GAD), and vesicular GABA transporters (VGATs) were measured in the VTA, respectively. The results revealed that extracellular GABA level was significantly increased in the CNP group (50.5 ± 18.9 nM) compared to the sham control group (10.2 ± 1.7 nM). In addition, expression of GAD

Objectives In the previous studies, it has been demonstrated that acupuncture treatment was effective on the suppression of withdrawal signs as well as self-administration behavior induced by morphine. Based on, the present study has investigated whether acupuncture could attenuate the anxiety-like behavior induced by withdrawal from chronic morphine treatment. Materials & methods Male Sprague-Dawley rats weighing 270–300 g were treated saline or morphine hydrochloride (10 mg/kg, s.c.) for 2 weeks. Following abstinence of 5 days in home cage, rats were subjected to the measurement of anxiety-like behavior in the elevated plus maze. Bicuculline (1 mg/kg, i.p.) and SCH 50911 (2 mg/kg, i.p.) were used to investigate the possible mechanism of acupuncture effects focusing on the GABA receptors system. Results Acupuncture at HT7 increased the time spent in open arms significantly. Also, these effects of acupuncture at HT7 were blocked by GABAA receptor antagonist. Conclusion Results of the present study suggest that acupuncture at HT7 can attenuate anxiety-like behavior induced by withdrawal from chronic morphine treatment through the mediation of GABAA receptor system.

The hyperactive state of sensory neurons in the spinal cord enhances pain transmission. Spinal glial cells have also been implicated in enhanced excitability of spinal dorsal horn neurons, resulting in pain amplification and distortions. Traumatic injuries of the neural system such as spinal cord injury (SCI) induce neuronal hyperactivity and glial activation, causing maladaptive synaptic plasticity in the spinal cord. Recent studies demonstrate that SCI causes persistent glial activation with concomitant neuronal hyperactivity, thus providing the substrate for central neuropathic pain. Hyperactive sensory neurons and activated glial cells increase intracellular and extracellular glutamate, neuropeptides, adenosine triphosphates, proinflammatory cytokines, and reactive oxygen species concentrations, all of which enhance pain transmission. In addition, hyperactive sensory neurons and glial cells overexpress receptors and ion channels that maintain this enhanced pain transmission. Therefore, post-SCI neuronal-glial interactions create maladaptive synaptic circuits and activate intracellular signaling events that permanently contribute to enhanced neuropathic pain. In this review, we describe how hyperactivity of sensory neurons contributes to the maintenance of chronic neuropathic pain via neuronal-glial interactions following SCI.

Acupuncture as a therapeutic intervention has been widely used for treatment of many pathophysiological disorders. For achieving improved therapeutic effects, relatively thick acupuncture needles have been frequently used in clinical practice with, in turn, enhanced stimulation intensity. However due to the discomforting nature of the larger-diameter acupuncture needles there is considerable interest in developing advanced acupuncture therapeutical techniques that provide more comfort with improved efficacy. So motivated, we have developed a new class of acupuncture needles, porous acupuncture needles (PANs) with hierarchical micro/nano-scale conical pores upon the surface, fabricated via a simple and well known electrochemical process, with surface area approximately 20 times greater than conventional acupuncture needles. The performance of these high-surface-area PANs is evaluated by monitoring the electrophysiological and behavioral responses from the in vivo stimulation of Shenmen (HT7) points in Wistar rats, showing PANs to be more effective in controlling electrophysiological and behavioral responses than conventional acupuncture needles. Comparative analysis of cocaine induced locomotor activity using PANs and thick acupuncture needles shows enhanced performance of PANs with significantly less pain sensation. Our work offers a unique pathway for achieving a comfortable and improved acupuncture therapeutic effect.

There is growing public interest in alternative approaches to addiction treatment and scientific interest in elucidating the neurobiological underpinnings of acupuncture. Our previous studies showed that acupuncture at a specific Shenmen (HT7) points reduced dopamine (DA) release in the nucleus accumbens (NAc) induced by drugs of abuse. The present study was carried out to evaluate the effects of HT7 acupuncture on γ-aminobutyric acid (GABA) neuronal activity in the ventral tegmental area (VTA) and the reinstatement of cocaine-seeking behavior. Using microdialysis and in vivo single-unit electrophysiology, we evaluated the effects of HT7 acupuncture on VTA GABA and NAc DA release and VTA GABA neuronal activity in rats. Using a within-session reinstatement paradigm in rats self-administering cocaine, we evaluated the effects of HT7 stimulation on cocaine-primed reinstatement. Acupuncture at HT7 significantly reduced cocaine suppression of GABA release and GABA neuron firing rates in the VTA. HT7 acupuncture attenuated cocaine-primed reinstatement, which was blocked by VTA infusions of the selective GABA

In the present study, we compared the roles of gracile neurons in mechanically-induced neuropathic pain caused by spinal injury and L5 spinal nerve ligation in rats. Behavioral and electrophysiological methods were used to measure mechanical allodynia in the hindpaws, and excitability of the gracile neurons in the medulla, respectively. In the spinal hemisection and spinal contusion models, mechanical allodynia developed in both hindpaws and lasted over a month. Three weeks following the hemisection, gracile neurons identified as wide-dynamic-range (WDR) and low-threshold (LT) neurons, showed increased neuronal activity to non-noxious mechanical stimuli compared to control groups, whereas the spinal contusion groups did not show evoked activity (*p

Chronic central neuropathic pain after central nervous system injuries remains refractory to therapeutic interventions. A novel approach would be to target key intracellular signaling proteins that are known to contribute to continued activation by phosphorylation of kinases, transcription factors, and/or receptors that contribute to changes in membrane excitability. We demonstrate that one signaling kinase, calcium/calmodulin-dependent kinase II (CaMKII), is critical in maintaining aberrant dorsal horn neuron hyperexcitability in the neuropathic pain condition after spinal cord injury (SCI). After contusion SCI at spinal level T10, activated CaMKII (phosphorylated, pCaMKII) expression is significantly upregulated in the T7/8 spinal dorsal horn in neurons, but not glial cells, and in oligodendrocytes in the dorsal column in the same rats that displayed at-level mechanical allodynia. Furthermore, identified spinothalamic neurons demonstrated significant increases of pCaMKII after SCI compared to sham-treated control animals. However, neither astrocytes nor microglia showed pCaMKII expression in either sham-treated or SCI rats. To demonstrate causality, treatment of SCI rats with KN-93, which prevents CaMKII activation, significantly attenuated at-level mechanical allodynia and aberrant wide dynamic range neuronal activity evoked by brush, pressure, and pinch stimuli and a graded series of von Frey stimuli, respectively. Persistent CaMKII activation contributes to chronic central neuropathic pain by mechanisms that involve maintained hyperexcitability of wide dynamic range dorsal horn neurons. Furthermore, targeting key signaling proteins is a novel, useful therapeutic strategy for treating chronic central neuropathic pain.

In this study, we examined whether topical treatment of glutamate receptor antagonists attenuate hyperexcitability of lumbar spinal dorsal horn neurons following low thoracic hemisection spinal cord injury in rats. Four weeks after spinal hemisection, neuronal activity in response to mechanical stimuli applied on the peripheral receptive field was significantly increased in three different phenotypes of lumbar spinal dorsal horn neurons: wide dynamic range (WDR), low threshold (LT) and high threshold (HT). Topical application of MK-801 (NMDA receptor antagonist, 50 microg) significantly attenuated the activity of WDR, but not LT and HT neurons; whereas, NBQX (AMPA receptor antagonist, 0.5 and 1 microg) significantly attenuated neuronal activity in all three phenotypes of neurons (*p0.05). However, MCPG (group I/II metabotropic glutamate receptor antagonist, 100 microg) had no effect. The present study, in the context of previous work, suggests that ionotropic glutamate receptor activation play critical roles in the maintenance of neuronal hyperexcitability and neuropathic "below-level" pain behavior following spinal hemisection injury.

In behavior methods to quantify neuropathic pain, visual observations of limb-withdrawal reflexes to stimuli are not always clear-cut, so this method is partly subjective. Our current data suggest that measurement of electrophysiological EMG magnitudes enables more reliable and objective assessment for quantifying nocifensive behaviors related to neuropathic pain.

Recent work regarding chronic central neuropathic pain (CNP) following spinal cord injury (SCI) suggests that activation of key signaling molecules such as members of the mitogen activated protein kinase (MAPK) family play a role in the expression of at-level mechanical allodynia. Specifically, Crown and colleagues (2005, 2006) have shown that the development of at-level CNP following moderate spinal cord injury is correlated with increased expression of the activated (and thus phosphorylated) forms of the MAPKs extracellular signal related kinase and p38 MAPK. The current study extends this work by directly examining the role of p38 MAPK in the maintenance of at-level CNP following spinal cord injury. Using a combination of behavioral, immunocytochemical, and electrophysiological measures we demonstrate that increased activation of p38 MAPK occurs in the spinal cord just rostral to the site of injury in rats that develop at-level mechanical allodynia after moderate SCI. Immunocytochemical analyses indicate that the increases in p38 MAPK activation occurred in astrocytes, microglia, and dorsal horn neurons in the spinal cord rostral to the site of injury. Inhibiting the enzymatic activity of p38 MAPK dose dependently reverses the behavioral expression of at-level mechanical allodynia and also decreases the hyperexcitability seen in thoracic dorsal horn neurons after moderate SCI. Taken together, these novel data are the first to demonstrate causality that increased activation of p38 MAPK in multiple cell types play an important role in the maintenance of at-level CNP following spinal cord injury.

ObjectivesThe occurrence of a relapse during abstinence is an important issue that must be addressed during treatment for drug addiction. We investigated the influence of drug exposure pattern on morphine-seeking behaviour following withdrawal. We also studied the role of the hippocampus in this process to confirm its involvement in drug relapse.MethodsMale Sprague–Dawley rats that were trained to self-administer morphine (1.0 mg/kg) using 2, 4, 6, 8, or 10 h daily sessions underwent withdrawal in their home cages and were re-exposed to the operant chamber to evaluate morphine-seeking behaviour. During the relapse session, rats were intravenously injected with morphine (0.25 mg/kg) or saline before re-exposure to the chamber. In the second experiment, rats were administered a microinjection of saline or cobalt chloride (CoCl2, 1 mM), a synaptic blocker, into the CA1 of the hippocampus prior to the relapse test.ResultsIn the first experiment, more morphine-seeking behaviour was observed in the 2 h group (animals trained to self-administer morphine during a 2 h daily session spread over 21 days) during the relapse session, despite all groups being exposed to similar amounts of morphine during the training period before withdrawal. In the second experiment, pretreatment with CoCl2 markedly reduced morphine-seeking behaviour in the 2 h group.ConclusionsThe present findings suggest that the exposure pattern influences the degree of relapse and that control of memorisation is important for prevention of relapse.

In this study, we examined whether a competitive AMPA receptor antagonist, NBQX, attenuates mechanical allodynia and hyperexcitability of spinal neurons in remote, caudal regions in persistent central neuropathic pain following spinal cord injury in rats. Spinal cord injury was produced by unilateral T13 transverse spinal hemisection, from dorsal to ventral, in male Sprague Dawley rats (200-250 g). Mechanical thresholds were measured behaviorally, and the excitability of wide-dynamic-range (WDR) dorsal horn neurons in the lumbar cord (L4-L5) was measured to assess central neuropathicpain. On postoperation day (POD) 28 after spinalhemisection, mechanical thresholds were significantly decreased in both injured (ipsilateral) and noninjured (contralateral) hindpaws compared with preinjury and sham control, respectively (P0.05). Intrathecal administration of NBQX (0.25, 0.5, 1 mM) significantly reversed the decreased mechanical thresholds in both hindpaws, dose dependently (P0.05). The excitability of WDR neurons was significantly enhanced on both sides of the lumbar dorsal horn 28 days following spinal hemisection (P0.05). The hyperexcitability of WDR neurons was attenuated by topical administration of NBQX (0.125, 0.25, 0.5, 1 mM), dose dependently (P0.05). Regression analysis indicated that at least three molecules of NBQX bond per receptor complex, and are needed to achieve inhibition of WDR hyperexcitability. In conclusion, our study demonstrates that the AMPA receptor plays an important role in behaviors related to the maintenance of central neuropathic pain below the level of spinal cord injury.

In this study, we investigated the role of the spinal GABAergic system in central neuropathic painlike outcomes following spinal cord injury (SCI) produced by a spinal hemitransection at T13 of the rat. After SCI, mechanical allodynia develops bilaterally in both hind paws of the rat, lasting longer than 40 days, as evidenced by an increase in paw withdrawal frequency in response to a weak von Frey filament. In naive rats, intrathecal (i.t.) administration in the lumbar spinal cord of GABAA and GABAB receptor antagonists, bicuculline (1-5 microg) and phaclofen (0.1-5 microg), respectively, causes a dose-dependent increase in the magnitude of mechanical allodynia. The SCI-induced mechanical allodynia in both hind-paws is attenuated by i.t. administration in the lumbar spinal cord of GABAA or GABAB receptor agonists, muscimol (1 microg) or baclofen (0.5 microg), respectively. In electrophysiological experiments, rats with SCI show a bilateral increase in hyperexcitability in response to natural stimuli in wide dynamic range (WDR) neurons in the lumbar spinal dorsal horn. The topical application of muscimol (1 microg) or baclofen (0.5 microg) onto the lumbar cord surface reduce the SCIinduced increased responsiveness of WDR neurons. Inhibitory effects of muscimol and baclofen on both the behavioral mechanical allodynia and the hyperexcitability in WDR neuron with SCI compared to controls, were antagonized by pre-treatment of bicuculline (10 microg) and phaclofen (5 microg), respectively. This study provides behavioral and electrophysiological evidence for the important role of the loss of spinal inhibitory tone, mediated by activation of both GABAA and GABAB receptors, in the development of central neuropathic pain following SCI.

This study examined the antinociceptive effect of electroacupuncture (EA) to heterotopic acupoints on formalin-induced pain in rats. EA (2 ms, 10 Hz, and 3 mA) was delivered to heterotopic acupoints HE7 and PE7, or non-acupoints at the right fore limb, for 30 min and was immediately followed by subcutaneous formalin injection into the left hind paw, respectively. The quantified pain score, electromyogram (EMG) response of the C-fiber reflex, and cFos immunoreactivity were assessed, respectively. EA to heterotopic acupoints significantly reduced both early- and late-phase pain-like behaviors and significantly decreased the EMG responses of the C-fiber reflex after formalin injection. By contrast, EA to non-acupoints had no significant effects on pain-like behavior or the EMG response. In addition, EA to heterotopic acupoints decreased cFos immunoreactivity in the lumbar spinal dorsal horn. Therefore, EA induced pre-emptive antinociception via the extra-segmental inhibition of the formalin-induced pain, suggesting that EA to heterotopic acupoints is a useful treatment for inflammatory pain.

Of the glutamate receptor types, the metabotropic glutamate receptors (mGluRs) are G proteins coupled and can initiate a number of intracellular pathways leading to hyperexcitability of spinal neurons. In this study, we tested the expression of mGluRs to determine which cell types might contribute to sustained neuronal hyperexcitability in the lumbar enlargement with postoperative day (POD) 7 (early), 14 (late), and 30 (chronic phase) following spinal cord injury (SCI) by unilateral hemisection at T13 in Sprague-Dawley rats. Expression was determined by confocal analyses of immunocytochemical reaction product of neurons (NeuN positive) and astrocytes (GFAP positive) in the dorsal horn on both sides of the L4 segment. Neurons were divided into two sizes: small (20 microm) and large (35 microm), for physiological reasons. We report a significant increase of mGluR(1) expression in large and small neurons of the dorsal horn on both sides of the cord in late and chronic phases when compared to control sham groups. Expression of mGluR(2/3) significantly increased in large neurons on the ipsilateral (hemisected) side in the late phase. Expression of mGluR(5) significantly increased in large neurons in early, late, and chronic phases. In addition, mGluR(1) and mGluR(5) expression after hemisection was significantly increased in astrocytes in early, late, and chronic phases; whereas mGluR(2/3) did not display any significant changes. In conclusion, our data demonstrate long-term changes in expression levels of Group I mGluRs (mGluR(1) and mGluR(5)) in both neurons and astrocytes in segments below a unilateral SCI. Thus, permanent alterations in dorsal horn receptor expression may play important roles in transmission of nociceptive responses in the spinal cord following SCI.

We tested the effect of age at the time of spinal cord injury (SCI) on locomotor recovery, in open field tests, and mechanical hyperalgesia, using paw withdrawal frequency (PWF) in response to noxious mechanical stimuli, in male Sprague-Dawley rats after spinal hemisection at T13 in young (40 days), adult (60 days) and middle-age (1 year) groups. Behavioral outcomes were measured weekly for 4 weeks in both SCI and sham groups. Following SCI, the young and adult groups recovered significantly more locomotor function, at a more rapid rate, than did the middle-age group. The PWF of the young group was significantly increased, the adult group was significantly decreased, and the middle-age group showed no significant change in fore- and hindlimbs when compared to other age groups, pre-injury and sham controls. These results support age-dependent behavioral outcomes after SCI.

Spinal cord injury (SCI) often leads to central pain syndrome including hyperalgesia to mechanical stimulation. Since there is evidence that nerve growth factor (NGF) contributes to pain-related behaviors, we wished to determine if anti-NGF might inhibit abnormal somatosensory behaviors that develop following SCI in rats. SCI was performed in male Sprague-Dawley rats by T13 spinal hemisection. After spinal hemisection, animals were untreated or treated daily with anti-NGF or saline intraperitoneally for 10 days. In groups of both hemisection only and hemisection with saline treatment, mechanical hyperalgesia developed in both hindlimbs, as evidenced by a decrease in paw withdrawal thresholds. Mechanical responsiveness of wide dynamic range (WDR) neurons on both sides of spinal cord also increased. The anti-NGF treated group demonstrated significant suppression of both mechanical hyperalgesia and increased WDR neuronal responsiveness. These results indicate that anti-NGF prevents the development of abnormal somatosensory behavior and suggest a potential pre-emptive analgesic treatment for central pain.

In the previous study, acupuncture at HT7 has shown to attenuate the self-administration of morphine at a low dose (0.1mg/kg). In this study, it was further investigated whether acupuncture at HT7 could attenuate the morphine self-administration at a high dose (0.5mg/kg). Male Sprague-Dawley rats weighing 270-300g were used. After surgery of catheterization, animals were trained to self-administer morphine solution (0.5mg/kg) using daily 1h session under fixed ratio 1 schedule for 3 weeks. Animals that had shown stable morphine-taking (establish baseline: variation less than 20% of the mean of three consecutive days) were subjected to the acupuncture treatment. Bicuculline and SCH 50911 were used to investigate the possible relation between the effect of acupuncture and the GABA receptor system. Acupuncture at HT7, but not at control acupoint, LI5, suppressed spontaneous morphine-taking behavior significantly. In addition, the effect of acupuncture was blocked by both GABA receptor antagonists. The results of this study suggest that acupuncture at HT7 suppresses morphine-taking behavior through the mediation of GABA receptor system.

The present study examined the effects of sympathetic stimulation on the activity of primary afferent neurons that had peripheral axons being injured previously by a spinal nerve ligation. About 22% of afferents with injured fibers that showed spontaneous discharge were excited by sympathetic stimulation or systemic injection of adrenaline. Most sympathetically-excited afferent neurons had axons that conducted in the A-fiber range. This sympathetically-evoked afferent excitation was not affected by cutting the spinal nerve at a place close to the dorsal root ganglion (DRG). Yohimbine, alpha2-antagonist, suppressed sympathetically-evoked afferent excitation which was not affected by alpha1-antagonist prazosin. Clonidine, alpha2-agonist, exerted an excitatory effect, whereas alpha1-agonist phenylephrine had no effect on the activity of afferents with injured fibers. No afferent fibers in control preparations responded to sympathetic stimulation. The results suggest that after a spinal nerve ligation, injured DRG neurons with fast-conducting fibers become sensitive to sympathetic activity via activation of alpha2-adrenoceptors.

In this study, we examined whether blocking spinal cord injury (SCI)-induced increases in reactive oxygen species (ROS) by a ROS scavenger would attenuate below-level central neuropathic pain and promote recovery of locomotion. Rats with T10 SCI developed mechanical allodynia in both hind paws and overproduction of ROS, as assayed by Dhet intensity, in neurons in the lumbar 4/5 dorsal horn ((∗)P

In the spinal cord, neuron and glial cells actively interact and contribute to neurofunction. Surprisingly, both cell types have similar receptors, transporters and ion channels and also produce similar neurotransmitters and cytokines. The neuroanatomical and neurochemical similarities work synergistically to maintain physiological homeostasis in the normal spinal cord. However, in trauma or disease states, spinal glia become activated, dorsal horn neurons become hyperexcitable contributing to sensitized neuronal-glial circuits. The maladaptive spinal circuits directly affect synaptic excitability, including activation of intracellular downstream cascades that result in enhanced evoked and spontaneous activity in dorsal horn neurons with the result that abnormal pain syndromes develop. Recent literature reported that spinal cord injury produces glial activation in the dorsal horn; however, the majority of glial activation studies after SCI have focused on transient and/or acute time points, from a few hours to 1 month, and peri-lesion sites, a few millimeters rostral and caudal to the lesion site. In addition, thoracic spinal cord injury produces activation of astrocytes and microglia that contributes to dorsal horn neuronal hyperexcitability and central neuropathic pain in above-level, at-level and below-level segments remote from the lesion in the spinal cord. The cellular and molecular events of glial activation are not simple events, rather they are the consequence of a combination of several neurochemical and neurophysiological changes following SCI. The ionic imbalances, neuroinflammation and alterations of cell cycle proteins after SCI are predominant components for neuroanatomical and neurochemical changes that result in glial activation. More importantly, SCI induced release of glutamate, proinflammatory cytokines, ATP, reactive oxygen species (ROS) and neurotrophic factors trigger activation of postsynaptic neuron and glial cells via their own receptors and channels that, in turn, contribute to neuronal-neuronal and neuronal-glial interaction as well as microglia-astrocytic interactions. However, a systematic review of temporal and spatial glial activation following SCI has not been done. In this review, we describe time and regional dependence of glial activation and describe activation mechanisms in various SCI models in rats. These data are placed in the broader context of glial activation mechanisms and chronic pain states. Our work in the context of work by others in SCI models demonstrates that dysfunctional glia, a condition called "gliopathy", is a key contributor in the underlying cellular mechanisms contributing to neuropathic pain.

Background Paclitaxel is a widely used chemotherapeutic drug for breast and ovarian cancer. Unfortunately, it induces neuropathic pain, which is a dose-limiting side effect. Free radicals have been implicated in many neurodegenerative diseases. The current study tests the hypothesis that a free radical scavenger plays an important role in reducing chemotherapy-induced neuropathic pain. Methods Neuropathic pain was induced by intraperitoneal injection of paclitaxel (2 mg/kg) on four alternate days (days 0, 2, 4, and 6) in male Sprague-Dawley rats. Phenyl N-tert-butylnitrone (PBN), a free radical scavenger, was administered intraperitoneally as a single dose or multiple doses before or after injury. Mechanical allodynia was measured by using von Frey filaments. Results The administration of paclitaxel induced mechanical allodynia, which began to manifest on days 7-10, peaked within 2 weeks, and plateaued for at least 2 months after the first paclitaxel injection. A single injection or multiple intraperitoneal injections of PBN ameliorated paclitaxel-induced pain behaviors in a dose-dependent manner. Further, multiple administrations of PBN starting on day 7 through day 15 after the first injection of paclitaxel completely prevented the development of mechanical allodynia. However, an intraperitoneal administration of pbn for 8 days starting with the first paclitaxel injection did not prevent the development of pain behavior. Conclusions This study clearly shows that PBN alleviated mechanical allodynia induced by paclitaxel in rats. Furthermore, our data show that PBN given on days 7 through 15 after the first paclitaxel injection prevented the development of chemotherapy-induced neuropathic pain. This clearly has a clinical implication.

In the present study, we have examined whether spinal hemisection injury induces changes in the electrophysiological properties of thalamic ventral posteriorlateral (VPL) neurons in rats. Male Sprague-Dawley rats were subjected to unilateral spinal cord injury by transverse hemisection at the T13 spinal segment. Four weeks after the T13 spinal hemisection, the injured rats displayed robust allodynic behaviors on both sides of hindpaws compared to sham controls (P < 0.05). Extracellular recordings taken 4 weeks after the hemisection revealed that wide dynamic range (WDR) neurons had significantly increased spontaneous and brush-, pressure-, and pinch-evoked activities, respectively, on both sides of the thalamic VPL regions (P < 0.05). In contrast, low threshold (LT) neurons showed only an increase in the brush-evoked activity compared to sham controls (P < 0.05). However, afterdischarge activity in both types of neurons showed no changes. In addition, both sides of the thalamic VPL regions showed higher incidences of WDR neurons. In conclusion, our data demonstrate that spinal unilateral injury induces bilaterally increased evoked activity in thalamic VPL neurons.

In the present study, we examined whether activation of p-38alpha MAPK modulates mechanical allodynia and neuronal hyperexcitability, and if propentofylline (PPF, a glial modulator) modulates specifically localized activated p-38alpha MAPK expression in caudal regions remote from a low thoracic hemisection injury in rats. T13 spinal hemisection produces bilateral mechanical allodynia in hindpaws with evoked (in response to mechanical stimuli) neuronal hyperexcitability in lumbar spinal wide dynamic range (WDR) neurons compared to sham controls. The mechanical allodynia and the evoked activity of WDR neurons is attenuated by intrathecal and topical administration of SB203580, an inhibitor of p-38 MAPK activation, dose dependently (p0.05); however, the spontaneous activity showed no significant differences compared to sham controls. After T13 spinal hemisection, significantly increased phosphorylated (activated form) p-38alpha MAPK expression was present in both superficial and deep dorsal horn neurons as well as in microglia, but not in astrocytes, in the lumbar spinal cord compared to sham controls (p0.05). Intrathecal application of PPF significantly attenuated the expression of phosphorylated p-38alpha MAPK in superficial dorsal horn neurons (10 mM) and in microglia (1 and 10 mM) in the lumbar spinal cord compared to the hemisection group (p0.05). In conclusion, our present data demonstrate that activated neuronal and microglial, but not astrocytic, p-38alpha MAPK contributes to the maintenance of neuronal hyperexcitability in caudal regions following spinal cord injury.

In this study, we evaluated whether propentofylline, a methylxanthine derivative, modulates spinal glial activation and GABAergic inhibitory tone by modulation of glutamic acid decarboxylase (GAD)65, the GABA synthase enzyme, in the spinal dorsal horn following spinal cord injury (SCI). Sprague–Dawley rats (225–250 g) were given a unilateral spinal transverse injury, from dorsal to ventral, at the T13 spinal segment. Unilateral spinal injured rats developed robust bilateral hindlimb mechanical allodynia and hyperexcitability of spinal wide dynamic range (WDR) neurons in the lumbar enlargement (L4–L5) compared to sham controls, which was attenuated by intrathecal (i.t.) administration of GABA, dose-dependently (0.01, 0.1, 0.5 μg). Western blotting and immunohistochemical data demonstrated that the expression level of GAD65 protein significantly decreased on both sides of the lumbar dorsal horn (L4/5) after SCI (p < 0.05). In addition, astrocytes and microglia showed soma hypertrophy as determined by increased soma area and increased GFAP and CD11b on both sides of the lumbar dorsal horn compared to sham controls, respectively (p < 0.05). Intrathecal treatment with propentofylline (PPF 10 mM) significantly attenuated the astrocytic and microglial soma hypertrophy and mechanical allodynia (p < 0.05). Additionally, the Western blotting and immunohistochemistry data demonstrated that i.t. treatment of PPF significantly prevented the decrease of GAD65 expression in both sides of the lumbar dorsal horn following SCI (p < 0.05). In conclusion, our present data demonstrate that propentofylline modulates glia activation and GABAergic inhibitory tone by modulation of GAD65 protein expression following spinal cord injury.

Spinal cord injury (SCI) often leads to chronic central pain (CCP) syndromes such as allodynia and hyperalgesia. Although several experimental animal models for CCP studies exist, little is known about the effect of age on the development of CCP following SCI. In this study, we evaluated behavioral responses to mechanical and thermal stimuli following SCI using three different age groups of adult Sprague-Dawley rats: young (40 days), adult (60 days), and middle-age (12 months). SCI was produced by unilateral hemisection of the spinal cord at T13. Behavioral measures of locomotor function were assayed in open field tests and somatosensory function by paw withdrawal frequency (PWF) to innocuous mechanical stimuli and paw withdrawal latency (PWL) to radiant heat stimuli on both the forelimbs and hindlimbs. Prior to hemisection, the PWF was not different between the three groups; however, the PWL of the young group was significantly greater than the adult and middle-age group. After spinal hemisection, spontaneous locomotor recovery occurred more rapidly in young and adult than in middle-age rats. In both forelimbs and hindlimbs, the young group displayed a significant increase in PWF and a significant decrease in PWL compared to presurgical and sham values or values from the adult and middle-age groups. These results indicate that younger rats developed more robust neuropathic behaviors than middle-age rats, indicating that age selection is an important factor in animal models of CCP syndromes following SCI. Additionally, our data suggest that age at the time of injury may be one risk factor in predicting the development of CCP after SCI in people.

Spontaneous pain, allodynia and hyperalgesia are well known phenomena following peripheral nerve or tissue injury, and it is speculated that secondary hyperalgesia and allodynia, are generally thought to depend on a hyperexcitability (sensitization) of neurons in the dorsal horn. It is supposed that the sensitization may be due to various actions of neurotransmitters (SP, CGRP, excitatory amino acids) released from the primary afferent fibers. In this study, we examined effects of the iontophoretically applied SP and CGRP on the response to EAA receptor agonists (NMDA and non-NMDA) in the WDR dorsal horn neurones and see if the effects of SP or CGRP mimic the characteristic response pattern known in various pain models. The main results are summarized as follows: 1) SP specifically potentiated NMDA response. 2) CGRP non-specifically potentiated both NMDA and AMPA responses. Potentiation of NMDA response, however, was significantly greater than that of AMPA response. 3) 50% of SP applied cells and 15.8% of CGRP applied cells showed reciprocal changes(potentiation of NMDA response and suppression of AMPA response). These results are generally consistent with the sensitization characteristics in diverse pain models and suggests that the modulatory effects of SP and CGRP on NMDA and non-NMDA (AMPA) response are, at least in part, contribute to the development of sensitization in various pain models.

Spinal cord injury often leads to central neuropathic pain syndromes, such as allodynic and hyperalgesic behaviors. Electrophysiologically, spinal dorsal horn neurons show enhanced activity to non-noxious and noxious stimuli as well as increased spontaneous activity following spinal cord injury, which often called hyperexcitability or central sensitization. Under hyperexcitable states, spinal neurons lose their ability of discrimination and encoding somatosensory information followed by abnormal somatosensory recognition to non-noxious and noxious stimuli. In the present review, we summarize a variety of pathophysiological mechanisms of neuronal hyperexcitability for treating or preventing central neuropathic pain syndrome following spinal cord injury.

Background: Allodynia, hyperalgesia, and spontaneous pain are symptoms characterized by chronic central pain which was frequently observed following a spinal cord injury (SCI). However, the underlying mechanism has not been fully understood. This study was conducted to investigate whether the loss of the GABAergic system in the spinal dorsal horn was involved in the development of central pain following a spinal cord injury. Methods: SCI was induced by a hemisection of the spinal cord at T13 in adult male Sprague-Dawley rats. Mechanical allodynia was tested by measuring paw withdrawal frequency in response to repeated applications of a von Frey hair to the plantar surface of the hind-paw. Single neuronal activity of the dorsal horn neurons (L4-L6) was recorded extracellularly using a carbon filament-filled glass microelectrode (2-4 MΩ). The drugs were intrathecally or topically administrated on the spinal surface for behavioral and electrophysiological experiments, respectively. Results: After a left spinal hemisection at T13, behavioral signs of mechanical allodynia developed on both hind-paws and responsiveness of spinal dorsal horn neurons increased on both sides of the spinal dorsal horn. GABA receptor agonists including GABAA and GABAB receptor subtypes suppressed mechanical allodynia on both sides of hind-paws and decreased responsiveness of spinal dorsal horn neurons on both sides of spinal cord. Conclusions: These results indicate that a loss of the GABAergic system within the spinal cord plays a key role on the development of central pain following a spinal cord injury.