Low frequencies, but not high frequencies of bi-polar spinal cord stimulation reduce cutaneous and muscle hyperalgesia induced by nerve injury

Spinal cord stimulation (SCS) is an established treatment for neuropathic pain. However, SCS is not effective for all the patients and the mechanisms underlying the reduction in pain by SCS are not clearly understood. To elucidate the mechanisms of pain relief by SCS, we utilized the spared nerve injury model. Sprague–Dawley rats were anesthetized, the tibial and common peroneal nerves were tightly ligated, and an epidural SCS lead implanted in the upper lumbar spinal cord. SCS was delivered daily at one of 4 different frequencies (4 Hz, 60 Hz, 100 Hz, and 250 Hz) at approximately 85% of motor threshold 2 weeks after nerve injury for 4 days. Mechanical withdrawal threshold of the paw and compression withdrawal threshold of the hamstring muscles were measured before and after SCS on each day. All rats showed a decrease in withdrawal threshold of the paw and the muscle 2 weeks after nerve injury. Treatment with either 4 Hz or 60 Hz SCS significantly reversed the decreased withdrawal threshold of the paw and muscle. The effect was cumulative with a greater reversal by the fourth treatment when compared to the first treatment. Treatment with 100 Hz, 250 Hz or sham SCS had no significant effect on the decreased withdrawal threshold of the paw or muscle that normally occurs after nerve injury. In conclusion, SCS at 4 Hz and 60 Hz was more effective in reducing hyperalgesia than higher frequencies of SCS (100 Hz and 250 Hz); and repeated treatments result in a cumulative reduction in hyperalgesia.     

Reactive oxygen species contribute to neuropathic pain by reducing spinal GABA release

Although both a loss of spinal inhibitory neurotransmission and the involvement of oxidative stress have been regarded as important mechanisms in the pathogenesis of pain, the relationship between these 2 mechanisms has not been studied. To determine whether reactive oxygen species (ROS) involvement in pain mechanisms is related to the diminished inhibitory transmission in the substantia gelatinosa (SG) of the spinal dorsal horn, behavioral studies and whole-cell recordings were performed in FVB/NJ mice. Neuropathic pain was induced by a tight ligation of the L5 spinal nerve (SNL). Pain behaviors in the affected foot were assessed by behavioral testing for mechanical hyperalgesia. Pain behaviors developed by 3 days and lasted more than 8 weeks. Both systemic and intrathecal administration of an ROS scavenger, phenyl-N-tert-butylnitrone (PBN), temporarily reversed mechanical hyperalgesia up to 2 hours, 1 week after SNL. In nonligated mice, an intrathecal injection of an ROS donor, tert-butyl hydroperoxide (t-BOOH), dose-dependently induced mechanical hyperalgesia for 1.5 hours. In whole-cell voltage clamp recordings of SG neurons, perfusion with t-BOOH significantly decreased the frequency of mIPSCs, and this effect was reversed by PBN. Furthermore, t-BOOH decreased the frequency of GABA_A receptor-mediated mIPSCs without altering their amplitudes but did not affect glycine receptor-mediated mIPSCs. In SNL mice, mIPSC frequency in SG neurons was significantly reduced as compared with that of normal mice, which was restored by PBN. The antihyperalgesic effect of PBN on mechanical hyperalgesia was attenuated by intrathecal bicuculline, a GABA_A receptor blocker. Our results indicate that the increased ROS in spinal cord may induce pain by reducing GABA inhibitory influence on SG neurons that are involved in pain transmission.     

RhoA/Rho kinase pathway contributes to the pathogenesis of thermal hyperalgesia in diabetic mice

Diabetic neuropathy is one of the most common complications of diabetes and causes various problems in daily life. Several investigations have noted that many factors in the spinal cord are involved in the symptoms of painful diabetic neuropathy, and there are very few effective therapeutic regimens. In the present study, we sought to elucidate the role of the RhoA/Rho kinase (ROCK) pathway in thermal hyperalgesia in diabetic mice. The intracellular localization of RhoA and the expression of eNOS were measured by western blotting. Thermal hyperalgesia was assessed by the tail-flick test and mechanical allodynia was assessed by automated von Frey filament test in streptozotocin(STZ)-induced diabetic mice. The spinal cord of STZ-treated diabetic mice showed increased membrane-bound RhoA compared to non-diabetic control. Treatment with the RhoA inhibitor exoenzyme C3, Clostridium botulinum, and the ROCK inhibitor Y27632 attenuated thermal hyperalgesia and mechanical allodynia in diabetic mice. Moreover, daily treatment with simvastatin attenuated all of those changes in diabetic mice. The expression of eNOS and NO metabolite contents in the spinal cord was decreased in diabetic mice, and these changes were normalized by treatment with simvastatin. The present results show that HMG-CoA reductase inhibitors have an inhibitory effect on thermal hyperalgesia in diabetic mice, which is mediated by an increase in NO production through the inhibition of RhoA/ROCK pathways. These results suggest that ROCK inhibitors and HMG-CoA inhibitors may be attractive compounds to relieve the symptoms of painful diabetic neuropathies.     

The induction of Fos-like immunoreactivity by noxious thermal, mechanical and chemical stimuli in the lumbar spinal cord of infant rats

The present study examined the maturation of nociceptive primary afferents using expression of Fos-like immunoreactivity in the second-order spinal cord neurons as an anatomical and functional marker. Pinch, immersion in hot water, or formalin injection applied to the hindpaw was used as the peripheral noxious stimulus in awake 0-, 1-, 2-, 3-, and 14-day-old rat pups. On the day of birth, all 3 stimuli elicited expression of the Fos protein in dorsal horn cells indicating that nociceptive primary afferents are functional at this age. The expression of the Fos protein was related to the intensity of stimulation since greater injection volumes of formalin or prolonged application of the thermal stimulus increased the number of stained nuclei. The number of stained nuclei was age dependent and older pups exhibited a greater number of stained nuclei. The results of this study are consistent with electrophysiological studies that have demonstrated that the primary nociceptive afferents continue to mature during the rats' postnatal life. Furthermore, the number of Fos immunoreactive neurons in immature rats are age and stimulus-intensity dependent.     

Peripheral and spinal GABAergic regulation of incisional pain in rats

Impairment of spinal GABAergic inhibition is demonstrated to contribute to pathologic chronic pain states. We investigated spinal and peripheral GABAergic regulation of incisional pain in rats. We found that intrathecal but not peripheral administration of muscimol (GABA-A receptor agonist) and baclofen (GABA-B receptor agonist) reduced mechanical and thermal hyperalgesia after plantar incision in rats. Nonevoked pain behavior after incision was unaffected by these agonists. Similarly, nociception in unincised rats was not reduced by the same dose of agonists. Thus, GABA-A and GABA-B receptors are involved in mediating incision-induced hyperalgesia (but not nonevoked pain). Intrathecal and systemic application of L-838,417, a subtype-selective benzodiazepine site agonist (α2, α3, α5), reduced mechanical and heat hyperalgesia after incision, indicating a role of these subunits in mediating incision-induced hyperalgesia. Interestingly, the effects of all agonists were more intense and prolonged on the day after surgery than on the day of incision. Similarly, spinally administered GABA-A and GABA-B antagonists increased pain behavior, again with a greater effect 1 day after incision. One possible explanation for this finding might be that an incision modulates GABA-mediated inhibition 1 day after incision. However, expression of GABA-A receptor subunits α2 and α3 and GABA-B receptor subunits within the dorsal horn of the spinal cord were unchanged after incision, indicating that receptor expression cannot explain a possible modulation of GABAergic inhibition after incision. Thus, other mechanisms need to be considered. In conclusion, GABA-A and GABA-B receptors are promising targets for postoperative, incisional pain in humans.     

Effect of antioxidant treatment on spinal GABA neurons in a neuropathic pain model in the mouse

One feature of neuropathic pain is a reduced spinal gamma-aminobutyric acid (GABA)-ergic inhibitory function. However, the mechanisms behind this attenuation remain to be elucidated. This study investigated the involvement of reactive oxygen species in the spinal GABA neuron loss and reduced GABA neuron excitability in spinal nerve ligation (SNL) model of neuropathic pain in mice. The importance of spinal GABAergic inhibition in neuropathic pain was tested by examining the effects of intrathecally administered GABA receptor agonists and antagonists in SNL and naïve mice, respectively. The effects of SNL and antioxidant treatment on GABA neuron loss and functional changes were examined in transgenic GAD67-enhanced green fluorescent protein positive (EGFP+) mice. GABA receptor agonists transiently reversed mechanical hypersensitivity of the hind paw in SNL mice. On the other hand, GABA receptor antagonists made naïve mice mechanically hypersensitive. Stereological analysis showed that the numbers of enhanced green fluorescent protein positive (EGFP+) GABA neurons were significantly decreased in the lateral superficial laminae (I–II) on the ipsilateral L5 spinal cord after SNL. Repeated antioxidant treatments significantly reduced the pain behaviors and prevented the reduction in EGFP+ GABA neurons. The response rate of the tonic firing GABA neurons recorded from SNL mice increased with antioxidant treatment, whereas no change was seen in those recorded from naïve mice, which suggested that oxidative stress impaired some spinal GABA neuron activity in the neuropathic pain condition. Together the data suggest that neuropathic pain, at least partially, is attributed to oxidative stress, which induces both a GABA neuron loss and dysfunction of surviving GABA neurons.     

运动训练对脊髓损伤后痉挛大鼠脊髓内钾-氯离子协同转运体2表达的影响

目的:研究运动训练对脊髓损伤(SCI)后痉挛大鼠行为学表现及脊髓内钾-氯离子协同转运体2(KCC2)表达的影响,探讨KCC2在运动训练的解痉效应中的作用。方法:将30只雌性SD大鼠随机分为假手术组、对照组、实验组。采用改良Allen撞击法建立脊髓损伤后痉挛模型,使用BBB评分、Ashworth评分来评估三组大鼠术后行为学变化,并于术后5周用免疫组化技术检测各组大鼠损伤远段脊髓KCC2表达。结果:假手术组术后行为学评分一直保持正常。术后1d—5周,对照组和实验组的BBB评分均持续升高,且2组组内不同时间点的BBB评分差异均有显著性意义(P0.05)。术后2—5周,实验组评分均高于对照组,且差异均有显著性意义(P0.05)。对照组和实验组在术后1周左右出现痉挛,后痉挛程度逐渐加重,至术后5周左右降低,2组组内不同时间点的Ashworth评定结果差异均有显著性意义(P0.05)。术后3—5周,对照组评定等级均高于实验组,差异有显著性意义(P0.05)。三组大鼠脊髓运动神经元胞膜上KCC2表达水平比较,组间差异有显著性意义(P0.05)。与假手术组相比,其余两组大鼠KCC2蛋白表达下调(P0.05);与对照组相比,实验组能够明显上调KCC2蛋白的表达(P0.05),但仍低于假手术组(P0.05)。结论:运动训练可以促进脊髓损伤后运动功能恢复,有效缓解痉挛,并可抑制KCC2表达的下调,运动训练的解痉效应可能是通过增加KCC2表达实现的。     

Early loss of peptidergic intraepidermal nerve fibers in an STZ-induced mouse model of insensate diabetic neuropathy

Peptidergic and nonpeptidergic nociceptive neurons represent parallel yet distinct pathways of pain transmission, but the functional consequences of such specificity are not fully understood. Here, we quantified the progression of peptidergic and nonpeptidergic axon loss within the epidermis in the setting of a dying-back neuropathy induced by diabetes. STZ-induced diabetic MrgD mice heterozygous for green fluorescent protein (GFP) in nonpeptidergic DRG neurons were evaluated for sensitivity to mechanical and noxious thermal and chemogenic stimuli 4 or 8 weeks post-STZ. Using GFP expression in conjunction with PGP9.5 staining, nonpeptidergic (PGP+/GFP+) and peptidergic (PGP+/GFP-) intraepidermal nerve fibers (IENFs) were quantified at each time point. At 4 weeks post-STZ, nonpeptidergic epidermal innervation remained unchanged while peptidergic innervation was reduced by 40.6% in diabetic mice. By 8 weeks post-STZ, both nonpeptidergic innervation and peptidergic innervation were reduced in diabetic mice by 34.1% and 43.8%, respectively, resulting in a 36.5% reduction in total epidermal IENFs. Behavioral deficits in mechanical, thermal, and chemogenic sensitivity were present 4 weeks post-STZ, concomitant with the reduction in peptidergic IENFs, but did not worsen over the next 4 weeks as nonpeptidergic fibers were lost, suggesting that the early reduction in peptidergic fibers may be an important driving force in the loss of cutaneous sensitivity. Furthermore, behavioral responses were correlated at the 4 week time point with peptidergic, but not nonpeptidergic, innervation. These results reveal that peptidergic and nonpeptidergic nociceptive neurons are differentially damaged by diabetes, and behavioral symptoms are more closely related to the losses in peptidergic epidermal fibers.     

Descending facilitation from the brainstem determines behavioural and neuronal hypersensitivity following nerve injury and efficacy of pregabalin

Various mechanisms at peripheral, spinal and/or supraspinal levels may underlie neuropathic pain. The nervous system's capacity for long-term reorganisation and chronic pain may result from abnormalities in RVM facilitatory On cells. Hence, via brainstem injections of the toxic conjugate dermorphin-saporin, which specifically lesions facilitatory cells expressing the mu-opioid receptor (MOR), we sought to determine the influence of these cells in normal and spinal nerve-ligated (SNL) rats. We combined behavioural, electrophysiological and pharmacological techniques to show that the supraspinal facilitatory drive is essential for neuronal processing of noxious stimuli in normal and neuropathic states, and that descending facilitatory neurones maintain behavioural hypersensitivities to mechanical stimuli during the late stages of nerve injury. Furthermore, we showed that these neurones are essential for the state-dependent inhibitory actions of pregabalin (PGB), a drug used in the treatment of neuropathic pain. During the early stages of nerve injury, or following medullary MOR cell ablation, PGB is ineffective at inhibiting spinal neuronal responses possibly due to quiescent spinal 5HT(3) receptors. This can however be overcome, and PGB's efficacy restored, by pharmacologically mimicking the descending drive at the spinal level with a 5HT(3) receptor agonist. Since RVM facilitatory neurones are integral to a spino-bulbo-spinal loop that reaches brain areas co-ordinating the sensory and affective components of pain, we propose that activity therein may influence painful outcome following nerve injury, and responsiveness to treatment.     

Electroacupuncture attenuates bone cancer-induced hyperalgesia and inhibits spinal preprodynorphin expression in a rat model

Cancer pain impairs the quality of life of cancer patients, but opioid intervention can cause significant side effects that further decrease quality of life. Although electroacupuncture (EA) has been used to treat cancer pain, its mechanisms are largely unknown. To examine its effects and underlying mechanisms on cancer pain, we injected AT-3.1 prostate cancer cells into the tibia to induce bone cancer in the male Copenhagen rat. The resulting pain was treated with 10 Hz/2 mA/0.4 ms pulse EA for 30 min daily at the point equivalent to the human acupoint GB30 (Huantiao) between days 14 and 18 after the injection. For sham control, EA needles were inserted into GB30 without stimulation. Thermal hyperalgesia, a decrease in paw withdrawal latency (PWL) to a noxious thermal stimulus, and mechanical hyperalgesia, a decrease in paw withdrawal pressure threshold (PWPT), was measured at baseline and 20 min after the EA treatment. Preprodynorphin mRNA and dynorphin were determined by RT-PCR and immunohistochemistry, respectively. Thermal and mechanical hyperalgesia developed ipsilaterally between days 12 and 18 after cancer cell inoculation. EA significantly (P < 0.05) attenuated this hyperalgesia, as shown by increased PWL and PWPT, and inhibited up-regulation of preprodynorphin mRNA and dynorphin compared to sham control. Intrathecal injection of antiserum against dynorphin A (1–17) also significantly inhibited the cancer-induced hyperalgesia.These results suggest that EA alleviates bone cancer pain at least in part by suppressing dynorphin expression, and they support the clinical use of EA in the treatment of cancer pain.     

Psychosocial subgroups in persons with spinal cord injuries and chronic pain

Widerström-Noga EG, Felix ER, Cruz-Almeida Y, Turk DC. Psychosocial subgroups in persons with spinal cord injuries and chronic pain.

Objectives

To define adaptational subgroups in people with chronic pain and spinal cord injury (SCI), and to compare these subgroups with respect to demographic factors, level of injury, functional independence, pain disability, depressed mood, social support, and life satisfaction.

Design

Interviews.

Setting

Veterans Affairs medical center and The Miami Project to Cure Paralysis.

Participants

Persons with SCI and chronic pain (N=190).

Interventions

Not applicable.

Main Outcome Measure

The Multidimensional Pain Inventory, SCI version.

Results

Cluster analysis revealed 3 subgroups: (1) dysfunctional (34.6% of all participants), characterized by higher pain severity, life interference, and affective distress scores, and lower levels of life control and activities scores; (2) interpersonally supported (33.0% of participants), characterized by moderately high pain severity, and higher life control, support from significant others, distracting responses, solicitous responses, and activities scores; and (3) adaptive copers (32.4% of participants), characterized by lower pain severity, life interference, affective distress, support from significant others, distracting responses, solicitous responses, activities and higher life control scores. Compared with the dysfunctional subgroup, the interpersonally supported subgroup reported significantly greater social support and life satisfaction and less pain disability and emotional distress, despite moderately high pain severity.

Conclusion

Three subgroups, independent of sex, pain duration, and functional status, were identified. Although severe pain significantly decreases life satisfaction after SCI, its impact is moderated by perceived social support.     

The effects of GABA(B) agonists and gabapentin on mechanical hyperalgesia in models of neuropathic and inflammatory pain in the rat

We have examined the effects of a novel GABA(B) agonist, CGP35024, in models of chronic neuropathic (partial sciatic ligation) and inflammatory (Freund's complete adjuvant) pain in the rat, and its inhibitory action on spinal transmission in vitro. The effects of CGP35024 were compared with L-baclofen and gabapentin. CGP35024 and L-baclofen reversed neuropathic mechanical hyperalgesia following single subcutaneous or intrathecal administration, but did not affect inflammatory mechanical hyperalgesia. Gabapentin only moderately affected neuropathic hyperalgesia following a single administration by either route, but produced significant reversal following daily administration for 5 days. It was only weakly active against inflammatory hyperalgesia following single or repeated administration. The antihyperalgesic effects of L-baclofen and CGP35024, but not gabapentin, were blocked by the selective GABA(B) receptor antagonist CGP56433A. CGP35024 was seven times more potent against neuropathic hyperalgesia than in the rotarod test for motor co-ordination, whilst L-baclofen was approximately equipotent in the two tests. In the isolated hemisected spinal cord from the rat, CGP35024, L-baclofen and gabapentin all inhibited capsaicin-evoked ventral root potentials (VRPs). CGP35024 and L-baclofen, but not gabapentin, also inhibited the polysynaptic and monosynaptic phases of electrically-evoked VRPs, as well as the 'wind-up' response to repetitive stimulation. These data indicate that CGP35024 and L-baclofen modulate nociceptive transmission in the spinal cord to inhibit neuropathic hyperalgesia, and that CGP35024 has a therapeutic window for antihyperalgesia over spasmolysis.     

大鼠脊髓损伤后NG2胶质细胞的活化增殖研究

目的观察大鼠损伤脊髓中NG2胶质细胞的活化。方法 60只雄性Wistar大鼠随机分为两组(n=15):手术组用改良Allen`s打击法制作脊髓损伤模型;假手术组只切除椎板,不损伤脊髓。在术后3 d、7 d、30 d,用免疫组化方法检测NG2细胞活化。结果相比较假手术组,手术组脊髓损伤后NG2细胞活化明显,细胞增殖活化在术后3 d升高,7 d达高峰,持续至30 d(P<0.05)。结论大鼠脊髓损伤后NG2大量活化增殖,参与了胶质瘢痕形成,对脊髓损伤后脊髓功能的恢复产生一定影响。     

大鼠脊髓损伤模型中硫氧还蛋白还原酶的表达及其作用

目的：探讨大鼠脊髓损伤模型中硫氧还蛋白还原酶（thioredoxin reductase，TR）的动态表达及其作用。方法：成年Wister大鼠36只，随机分为6组，每组6只。建立大鼠脊髓压迫伤模型，以原位杂交法测定脊髓损伤后不同时间TR的动态变化。结果：脊髓压迫伤后2hTR mRNA表达开始增高，6—12h达到高峰，24h开始下降，48h降到正常水平。结论：脊髓损伤后TR合成增加，通过抗氧化、清除自由基及抑制细胞凋亡，对脊髓损伤有保护作用。     

Internal consistency, stability, and validity of the spinal cord injury version of the multidimensional pain inventory

OBJECTIVE: To evaluate the internal consistency, stability, and construct validity of a spinal cord injury (SCI) version of the Multidimensional Pain Inventory (MPI-SCI). DESIGN: Interview. SETTING: Veterans Affairs medical center and university-based institute. PARTICIPANTS: Community sample of persons with SCI and chronic pain (N=161). INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURE: The MPI-SCI. RESULTS: The internal consistency of the MPI-SCI subscales ranged from fair (.60) for affective distress to substantial (.94) for pain interference with activities. The subscales of the MPI-SCI (ie, life interference [r=.81], affective distress [r=.71], solicitous responses [r=.86], distracting responses [r=.85], general activity [r=.69], pain interference with activities [r=.78], pain severity [r=.69], negative responses [r=.69]) showed adequate stability. In contrast, the stability of the support (r=.59) and the life control subscales (r=.31) was unacceptably low. All MPI-SCI subscales with the exception of the perceived responses by significant others subscales showed good convergent, discriminant, and concurrent validity. CONCLUSIONS: The MPI-SCI appears to be a reasonable measure for evaluating chronic pain impact after SCI. In clinical trials, however, supplementary instruments should be included to assess changes in affect, social support, and perceptions of life control.     

Lycopene attenuates thermal hyperalgesia in a diabetic mouse model of neuropathic pain

Diabetic neuropathic pain, an important microvascular complication of diabetes mellitus is recognized as one of the most difficult types of pain to treat. The development of tolerance, inadequate relief and potential toxicity of classical antinociceptives warrant the investigation of the newer agents to relieve this pain. The aim of the present study was to explore the antinociceptive effect of lycopene and its effect on tumour necrosis factor‐α (TNF‐α) and nitric oxide (NO) release in streptozotocin induced diabetic mice. Four weeks after a single intraperitoneal injection of streptozotocin (200mg/kg), mice were tested in the tail immersion and hot‐plate assays. Diabetic mice exhibited significant hyperalgesia alongwith increased plasma glucose and decreased body weights as compared with control mice. Lycopene (1, 2 and 4mg/kg body weight; per oral) treatment, from the 4th to 8th week after streptozotocin injection, significantly attenuated thermal hyperalgesia and the hot‐plate latencies. Lycopene also inhibited the TNF‐α and NO release in a dose dependent manner. These results indicate an antinociceptive activity of lycopene possibly through its inhibitory action on NO and TNF‐α release and point towards its potential to attenuate diabetic neuropathic pain.     

Gender and minority differences in the pain experience of people with spinal cord injury

OBJECTIVES: To examine gender and minority differences in the prevalence and severity of pain in people with traumatic-onset spinal cord injury (SCI) during follow-up, and to determine the relation of those differences to demographic characteristics, etiology of injury, and level and extent of the lesion. DESIGN: Survey and analysis of cross-sectional data using case-control methodology and multiple regression methods. SETTING: Model Spinal Cord Injury Systems (MSCIS). PARTICIPANTS: A total of 7379 individuals with traumatic-onset SCI from 16 MSCIS entered in the National Spinal Cord Injury Statistical Center database between 1998 and 2002. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Prevalence and severity of pain as reported in follow-up surveys. RESULTS: Pain prevalence remained fairly stable over time, ranging from 81% at 1 year postinjury to 82.7% at 25 years. Pain was no more common in women than in men, nor did pain severity scores differ significantly. However, pain prevalence was significantly lower among nonwhites, although they tended to report a higher average pain severity score when pain was present. Also, people with SCI who were employed when injured, who had more than a high school education, and who were not tetraplegic reported a higher prevalence of pain. Pain interfered with work more often for women and nonwhites during some, but not all, follow-up years, and for those who were not employed at the time of interview, for those whose SCI was caused by violence, for those with paraplegia, and for those with incomplete SCI. CONCLUSIONS: Pain is a common and significant problem for the majority of people with SCI. It may interfere less frequently with work over time, which suggests that an adaptive process may be occurring. Gender differences in the pain experience did not emerge, but nonwhites tended to have a lower prevalence of pain. If pain was present, nonwhites tended to report more severe pain than did whites. Further research is needed to delineate the possible psychosocial and biomedical causes of these findings.     

Spinal interleukin-17 promotes thermal hyperalgesia and NMDA NR1 phosphorylation in an inflammatory pain rat model

It is known that interleukin-17 (IL-17) is associated with autoimmune disorders and that peripheral IL-17 plays a role in arthritis and neuropathic pain. The present study investigated the possibility of spinal cell expression of IL-17 during inflammatory pain and possible IL-17 involvement in such pain. Hyperalgesia was induced by injecting complete Freund adjuvant (CFA, 0.08 mL, 40 μg Mycobacterium tuberculosis) into one hind paw of the rat. Paw withdrawal latency (PWL) was tested before (−48 h) and 2 and 24 h after CFA injection to assess hyperalgesia. IL-17 antibody (0.2–2 μg/rat) was given intrathecally (i.t.) 24 h before CFA to block the action of basal IL-17 and 2 h before each of 2 PWL tests to block CFA-induced IL-17. I.t. recombinant IL-17 (10–400 ng per rat) was administered to naive rats to determine its effects on PWL and phosphorylated NR1 (p-NR1). p-NR1 modulates N-methyl-d-aspartate receptor (NMDAR) activity to facilitate pain. Spinal cords were removed for IL-17 immunostaining, double immunostaining of IL-17/cell markers and IL-17 receptor A (IL-17RA)/NR1, for Western blot testing of IL-17, p-NR1, IL-17RA, and GFAP, for in situ IL-17RA hybridization, and for real time polymerase chain reaction of IL-17RA. The data reveal that IL-17 is up-regulated in activated and nonactivated astrocytes; that IL-17RA is localized in NR1-immunoreactive neurons and up-regulated; and that IL-17 antibody at 2 μg/rat significantly increased PWL (P < .05) and decreased p-NR1 and IL-17RA compared to control in CFA- and IL-17-injected rats. The results suggest that spinal IL-17 is produced by astrocytes and enhances p-NR1 to facilitate pain.     

Differential distribution of activated spinal neurons containing glycine and/or GABA and expressing c-fos in acute and chronic pain models

The inhibitory transmitters GABA and glycine play an important role in modulating pain transmission, both in normal and in pathological situations. In the present study we have combined in situ hybridization for identifying spinal neurons that use the transmitter(s) glycine and/or GABA (Gly/GABA neurons) with immunohistochemistry for c-fos, a marker for neuronal activation. This procedure was used with acute pain models induced by the injection of capsaicin or formalin; and chronic pain models using Complete Freund’s Adjuvant (CFA, chronic inflammation), and the spared nerve injury (SNI) model (neuropathic pain). In all models Gly/GABA neurons were activated as indicated by their expression of c-fos. The pattern of Gly/GABA neuronal activation was different for every model, both anatomically and quantitatively. However, the averaged percentage of activated neurons that were Gly/GABA in the chronic phase (?20 h survival, 46%) was significantly higher than in the acute phase (?2 h survival, 34%). In addition, the total numbers of activated Gly/GABA neurons were similar in both phases, showing that the activation of non-Gly/GABA (presumed excitatory) neurons in the chronic phase decreased. Finally, morphine application equally decreased the total number of activated neurons and activated Gly/GABA neurons. This showed that morphine did not specifically activate Gly/GABA neurons to achieve nociceptive inhibition. The present study shows an increased activity of Gly/GABA neurons in acute and chronic models. This mechanism, together with mechanisms that antagonize the effects of GABA and glycine at the receptor level, may determine the sensitivity of our pain system during health and disease.