Activation of NK₁ receptors in the locus coeruleus induces analgesia through noradrenergic-mediated descending inhibition in a rat model of neuropathic pain

BACKGROUND AND PURPOSE

The locus coeruleus (LC) is a major source of noradrenergic projections to the dorsal spinal cord, and thereby plays an important role in the modulation of nociceptive information. The LC receives inputs from substance P (SP)-containing fibres from other regions, and expresses the NK₁ tachykinin receptor, a functional receptor for SP. In the present study, we investigated the roles of SP in the LC in neuropathic pain.

EXPERIMENTAL APPROACH

Chronic constriction injury (CCI) of the left sciatic nerve was performed in rats to induce neuropathic pain. After development of neuropathic pain, SP was injected into the LC and the nocifensive behaviours were assessed. The involvement of noradrenergic descending inhibition in SP-induced analgesia was examined by i.t. administration of yohimbine, an α₂-adrenoceptor antagonist. NK₁ receptor expression in the LC was examined by immunohistochemistry.

KEY RESULTS

In CCI rats, mechanical allodynia was alleviated by SP injection into the LC. These effects were abolished by prior injection of WIN 51708, an NK₁ receptor antagonist, into the LC or i.t. treatment with yohimbine. NK₁ receptor-like immunoreactivity was observed in noradrenergic neurons throughout the LC in intact rats, and remained unchanged after CCI.

CONCLUSION AND IMPLICATIONS

SP in the LC exerted analgesic effects on neuropathic pain through NK₁ receptor activation and resulted in facilitation of spinal noradrenergic transmission. Accordingly, manipulation of the SP/NK₁ receptor signalling pathway in the LC may be a promising strategy for effective treatment of neuropathic pain.     

Glial cell line-derived neurotrophic factormediated enhancement of noradrenergic descending inhibition in the locus coeruleus exerts prolonged analgesia in neuropathic pain

Background and Purpose

The locus coeruleus (LC) is the principal nucleus containing the noradrenergic neurons and is a major endogenous source of pain modulation in the brain. Glial cell line-derived neurotrophic factor (GDNF), a well-established neurotrophic factor for noradrenergic neurons, is a major pain modulator in the spinal cord and primary sensory neurons. However, it is unknown whether GDNF is involved in pain modulation in the LC.

Experimental Approach

Rats with chronic constriction injury (CCI) of the left sciatic nerve were used as a model of neuropathic pain. GDNF was injected into the left LC of rats with CCI for 3 consecutive days and changes in mechanical allodynia and thermal hyperalgesia were assessed. The α₂-adrenoceptor antagonist yohimbine was injected intrathecally to assess the involvement of descending inhibition in GDNF-mediated analgesia. The MEK inhibitor U0126 was used to investigate whether the ERK signalling pathway plays a role in the analgesic effects of GDNF.

Key Results

Both mechanical allodynia and thermal hyperalgesia were attenuated 24 h after the first GDNF injection. GDNF increased the noradrenaline content in the dorsal spinal cord. The analgesic effects continued for at least 3 days after the last injection. Yohimbine abolished these effects of GDNF. The analgesic effects of GDNF were partly, but significantly, inhibited by prior injection of U0126 into the LC.

Conclusions and Implications

GDNF injection into the LC exerts prolonged analgesic effects on neuropathic pain in rats by enhancing descending noradrenergic inhibition.     

Cytokine activin C ameliorates chronic neuropathic pain in peripheral nerve injury rodents by modulating the TRPV1 channel

Background and PurposeThe cytokine activin C is mainly expressed in small‐diameter dorsal root ganglion (DRG) neurons and suppresses inflammatory pain. However, the effects of activin C in neuropathic pain remain elusive.Experimental ApproachMale rats and wild‐type and TRPV1 knockout mice with peripheral nerve injury ‐ sciatic nerve axotomy and spinal nerve ligation in rats; chronic constriction injury (CCI) in mice – provided models of chronic neuropathic pain. Ipsilateral lumbar (L)4–5 DRGs were assayed for activin C expression. Chronic neuropathic pain animals were treated with intrathecal or locally pre‐administered activin C or the vehicle. Nociceptive behaviours and pain‐related markers in L4–5 DRGs and spinal cord were evaluated. TRPV1 channel modulation by activin C was measured.Key ResultsFollowing peripheral nerve injury, expression of activin βC subunit mRNA and activin C protein was markedly up‐regulated in L4–5 DRGs of animals with axotomy, SNL or CCI. [Correction added on 26 November 2020, after first online publication: The preceding sentence has been corrected in this current version.] Intrathecal activin C dose‐dependently inhibited neuropathic pain in spinal nerve ligated rats. Local pre‐administration of activin C decreased neuropathic pain, macrophage infiltration into ipsilateral L4–5 DRGs and microglial reaction in L4–5 spinal cords of mice with CCI. In rat DRG neurons, activin C enhanced capsaicin‐induced TRPV1 currents. Pre‐treatment with activin C reduced capsaicin‐evoked acute hyperalgesia and normalized capsaicin‐evoked persistent hypothermia in mice. Finally, the analgesic effect of activin C was abolished in TRPV1 knockout mice with CCI.Conclusion and ImplicationsActivin C inhibits neuropathic pain by modulating TRPV1 channels, revealing potential analgesic applications in chronic neuropathic pain therapy.     

Capnellene,a natural marine compound derived from soft coral,attenuates chronic constriction injury-induced neuropathic pain in rats

Background and purpose:

Natural compounds obtained from marine organisms have received considerable attention as potential sources of novel drugs for treatment of human inflammatory diseases. Capnellene, isolated from the marine soft coral Capnella imbricate, 4,4,6a-trimethyl-3-methylene-decahydro-cyclopenta[]pentalene-2,3a-diol (GB9) exhibited anti-inflammatory actions on activated macrophages in vitro. Here we have assessed the anti-neuroinflammatory properties of GB9 and its acetylated derivative, acetic acid 3a-hydroxy-4,4,6a-trimethyl-3-methylene-decahydro-cyclopenta[]pentalen-2-yl ester (GB10).

Experimental approach:

Effects of GB9 or GB10 on the expression of inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) in interferon-γ (IFN-γ)-stimulated mouse microglial BV2 cells were measured by Western blot. The in vivo effects of these compounds were examined in the chronic constriction injury (CCI) rat model of neuropathic pain, measuring thermal hyperalgesia, and microglial activation and COX-2 protein in lumbar spinal cord, by immunohistochemistry.

Key results:

In BV2 cells, GB9 and GB10 inhibited the expression of iNOS and COX-2, stimulated by IFN-γ. Intrathecal administration of GB9 and GB10 inhibited CCI-induced nociceptive sensitization and thermal hyperalgesia in a dose-dependent manner. Intraperitoneal injection of GB9 inhibited CCI-induced thermal hyperalgesia and also inhibited CCI-induced activation of microglial cells and up-regulation of COX-2 in the dorsal horn of the lumbar spinal cord ipsilateral to the injury.

Conclusion and implications:

Taken together, these data indicate that the marine-derived capnellenes, GB9 and GB10, had anti-neuroinflammatory and anti-nociceptive properties in IFN-γ-stimulated microglial cells and in neuropathic rats respectively. Therefore, capnellene may serve as a useful lead compound in the search for new therapeutic agents for treatment of neuroinflammatory diseases.     

Central and peripheral sites of action for CB₂ receptor mediated analgesic activity in chronic inflammatory and neuropathic pain models in rats

BACKGROUND AND PURPOSE

Cannabinoid CB₂ receptor activation by selective agonists has been shown to produce analgesic effects in preclinical models of inflammatory and neuropathic pain. However, mechanisms underlying CB₂-mediated analgesic effects remain largely unknown. The present study was conducted to elucidate the CB₂ receptor expression in ‘pain relevant’ tissues and the potential sites of action of CB₂ agonism in rats.

EXPERIMENTAL APPROACH

Expression of cannabinoid receptor mRNA was evaluated by quantitative RT-PCR in dorsal root ganglia (DRGs), spinal cords, paws and several brain regions of sham, chronic inflammatory pain (CFA) and neuropathic pain (spinal nerve ligation, SNL) rats. The sites of CB₂ mediated antinociception were evaluated in vivo following intra-DRG, intrathecal (i.t.) or intraplantar (i.paw) administration of potent CB₂-selective agonists A-836339 and AM1241.

KEY RESULTS

CB₂ receptor gene expression was significantly up-regulated in DRGs (SNL and CFA), spinal cords (SNL) or paws (CFA) ipsilateral to injury under inflammatory and neuropathic pain conditions. Systemic A-836339 and AM1241 produced dose-dependent efficacy in both inflammatory and neuropathic pain models. Local administration of CB₂ agonists also produced significant analgesic effects in SNL (intra-DRG and i.t.) and CFA (intra-DRG) pain models. In contrast to A-836339, i.paw administration of AM-1241 dose-relatedly reversed the CFA-induced thermal hyperalgesia, suggesting that different mechanisms may be contributing to its in vivo properties.

CONCLUSIONS AND IMPLICATIONS

These results demonstrate that both DRG and spinal cord are important sites contributing to CB₂ receptor-mediated analgesia and that the changes in CB₂ receptor expression play a crucial role for the sites of action in regulating pain perception.     

Effects of milnacipran, a 5-HT and noradrenaline reuptake inhibitor, on C-fibre-evoked field potentials in spinal long-term potentiation and neuropathic pain

BACKGROUND AND PURPOSE

The analgesic action of 5-HT and noradrenaline reuptake inhibitors (SNRIs) on nociceptive synaptic transmission in the spinal cord is poorly understood. We investigated the effects of milnacipran, an SNRI, on C-fibre-evoked field potentials (FPs) in spinal long-term potentiation (LTP), a proposed synaptic mechanism of hypersensitivity, and on the FPs in a neuropathic pain model.

EXPERIMENTAL APPROACH

C-fibre-evoked FPs by electrical stimulation of the sciatic nerve fibres were recorded in the spinal dorsal horn of anaesthetized adult rats, and LTP was induced by high-frequency stimulation of the sciatic nerve fibres. A rat model of neuropathic pain was produced by L5 spinal nerve ligation and transection.

KEY RESULTS

Milnacipran produced prolonged inhibition of C-fibre-evoked FPs when applied spinally after the establishment of LTP of C-fibre-evoked FPs in naïve animals. In the neuropathic pain model, spinal administration of milnacipran clearly reduced the basal C-fibre-evoked FPs. These inhibitory effects of milnacipran were blocked by spinal administration of methysergide, a 5-HT_1/2 receptor antagonist, and yohimbine or idazoxan, α₂-adrenoceptor antagonists. However, spinal administration of milnacipran in naïve animals did not affect the basal C-fibre-evoked FPs and the induction of spinal LTP.

CONCLUSION AND IMPLICATIONS

Milnacipran inhibited C-fibre-mediated nociceptive synaptic transmission in the spinal dorsal horn after the establishment of spinal LTP and in the neuropathic pain model, by activating both spinal 5-hydroxytryptaminergic and noradrenergic systems. The condition-dependent inhibition of the C-fibre-mediated transmission by milnacipran could provide novel evidence regarding the analgesic mechanisms of SNRIs in chronic pain.     

The activation of supraspinal GPR40/FFA1 receptor signalling regulates the descending pain control system

Background and Purpose

The ω-3 polyunsaturated fatty acids exert antinociceptive effects in inflammatory and neuropathic pain; however, the underlying mechanisms remain unclear. Docosahexaenoic acid-induced antinociception may be mediated by the orphan GPR40, now identified as the free fatty acid receptor 1 (FFA1 receptor). Here, we examined the involvement of supraspinal FFA1 receptor signalling in the regulation of inhibitory pain control systems consisting of serotonergic and noradrenergic neurons.

Experimental Approach

Formalin-induced pain behaviours were measured in mice. Antinociception induced by FFA1 receptor agonists was examined by intrathecal injections of a catecholaminergic toxin, 5-HT lowering drug or these antagonists. The expression of FFA1 receptor protein and c-Fos was estimated by immunohistochemistry, and the levels of noradrenaline and 5-HT in the spinal cord were measured by LC-MS/MS.

Key Results

FFA1 receptors colocalized with NeuN (a neuron marker) in the medulla oblongata and with tryptophan hydroxylase (TPH; a serotonergic neuron marker) and dopamine β-hydroxylase (DBH; a noradrenergic neuron marker). A single i.c.v. injection of GW9508, a FFA1 receptor agonist, increased the number of c-Fos-positive cells and the number of neurons double-labelled for c-Fos and TPH and/or DBH. It decreased formalin-induced pain behaviour. This effect was inhibited by pretreatment with 6-hydroxydopamine, DL-p-chlorophenylalanine, yohimbine or WAY100635. Furthermore, GW9508 facilitated the release of noradrenaline and 5-HT in the spinal cord. In addition, GW1100, a FFA1 receptor antagonist, significantly increased formalin-induced pain-related behaviour.

Conclusion and Implications

Activation of the FFA1 receptor signalling pathway may play an important role in the regulation of the descending pain control system.     

Combined inhibition of monoacylglycerol lipase and cyclooxygenases synergistically reduces neuropathic pain in mice

Background and Purpose

Neuropathic pain is commonly treated with GABA analogues, steroids or non-steroidal anti-inflammatory drugs (NSAIDs). NSAIDs inhibit one or more COX isozymes but chronic COX inhibition paradoxically increases gastrointestinal inflammation and risk of unwanted cardiovascular events. The cannabinoids also have analgesic and anti-inflammatory properties and reduce neuropathic pain in animal models. The present study investigated the analgesic effects of inhibiting both monoacylglycerol lipase (MAGL) and COX enzymes, using low doses of both inhibitors.

Experimental Approach

Mice subjected to chronic constriction injury (CCI) were tested for mechanical and cold allodynia after administration of the MAGL inhibitor, JZL184, or the non-selective COX inhibitor diclofenac. Then, both drugs were co-administered at fixed dose proportions of 1:3, 1:1 and 3:1, based on their ED₅₀ values. PGs, endocannabinoids and related lipids were quantified in lumbar spinal cord.

Key Results

Combining low doses of JZL184 and diclofenac synergistically attenuated mechanical allodynia and additively reduced cold allodynia. The cannabinoid CB₁ receptor antagonist, rimonabant, but not the CB₂ receptor antagonist, SR144528, blocked the analgesic effects of the JZL184 and diclofenac combination on mechanical allodynia, implying that CB₁ receptors were primarily responsible for the anti-allodynia. Diclofenac alone and with JZL184 significantly reduced PGE₂ and PGF_2α in lumbar spinal cord tissue, whereas JZL184 alone caused significant increases in the endocannabinoid metabolite, N-arachidonoyl glycine.

Conclusions and Implications

Combining COX and MAGL inhibition is a promising therapeutic approach for reducing neuropathic pain with minimal side effects.     

Spinal actions of ω-conotoxins,CVID, MVIIA and related peptides in a rat neuropathic pain model

BACKGROUND AND PURPOSE

Antagonists of the N-type voltage gated calcium channel (VGCC), Ca_v2.2, have a potentially important role in the treatment of chronic neuropathic pain. ω-conotoxins, such MVIIA and CVID are effective in neuropathic pain models. CVID is reported to have a greater therapeutic index than MVIIA in neuropathic pain models, and it has been suggested that this is due to faster reversibility of binding, but it is not known whether this can be improved further.

EXPERIMENTAL APPROACH

We examined the potency of CVID, MVIIA and two intermediate hybrids ([K10R]CVID and [R10K]MVIIA) to reverse signs of neuropathic pain in a rat nerve ligation model in parallel with production of side effects. We also examined the potency and reversibility to inhibit primary afferent synaptic neurotransmission in rat spinal cord slices.

KEY RESULTS

All ω-conotoxins produced dose-dependent reduction in mechanical allodynia. They also produced side effects on the rotarod test and in a visual side-effect score. CVID displayed a marginally better therapeutic index than MVIIA. The hybrids had a lesser effect in the rotarod test than either of their parent peptides. Finally, the conotoxins all presynaptically inhibited excitatory synaptic neurotransmission into the dorsal horn and displayed recovery that was largely dependent upon the magnitude of inhibition and not the conotoxin type.

CONCLUSIONS AND IMPLICATIONS

These findings indicate that CVID provides only a marginal improvement over MVIIA in a preclinical model of neuropathic pain, which appears to be unrelated to reversibility from binding. Hybrids of these conotoxins might provide viable alternative treatments.     

The non-peptide GLP-1 receptor agonist WB4-24 blocks inflammatory nociception by stimulating β-endorphin release from spinal microglia

BACKGROUND AND PURPOSE

Two peptide agonists of the glucagon-like peptide-1 (GLP-1) receptor, exenatide and GLP-1 itself, exert anti-hypersensitive effects in neuropathic, cancer and diabetic pain. In this study, we have assessed the anti-allodynic and anti-hyperalgesic effects of the non-peptide agonist WB4-24 in inflammatory nociception and the possible involvement of microglial β-endorphin and pro-inflammatory cytokines.

EXPERIMENTAL APPROACH

We used rat models of inflammatory nociception induced by formalin, carrageenan or complete Freund''s adjuvant (CFA), to test mechanical allodynia and thermal hyperalgesia. Expression of β-endorphin and pro-inflammatory cytokines was measured using real-time quantitative PCR and fluorescent immunoassays.

KEY RESULTS

WB4-24 displaced the specific binding of exendin (9–39) in microglia. Single intrathecal injection of WB4-24 (0.3, 1, 3, 10, 30 and 100 μg) exerted dose-dependent, specific, anti-hypersensitive effects in acute and chronic inflammatory nociception induced by formalin, carrageenan and CFA, with a maximal inhibition of 60–80%. Spinal WB4-24 was not effective in altering nociceptive pain. Subcutaneous injection of WB4-24 was also antinociceptive in CFA-treated rats. WB4-24 evoked β-endorphin release but did not inhibit expression of pro-inflammatory cytokines in either the spinal cord of CFA-treated rats or cultured microglia stimulated by LPS. WB4-24 anti-allodynia was prevented by a microglial inhibitor, β-endorphin antiserum and a μ-opioid receptor antagonist.

CONCLUSIONS AND IMPLICATIONS

Our results suggest that WB4-24 inhibits inflammatory nociception by releasing analgesic β-endorphin rather than inhibiting the expression of proalgesic pro-inflammatory cytokines in spinal microglia, and that the spinal GLP-1 receptor is a potential target molecule for the treatment of pain hypersensitivity including inflammatory nociception.     

Gonadectomy affects brain derived neurotrophic factor in rats after chronic constriction nerve injury

INTRODUCTION The development of animal models of neuropathicpainhas beenof value in characterizingthe neuralmecha-nisms involved in neuropathic pain syndromes and as-sociated processes following peripheral nerve injury.The chronic constriction injury (CCI) model[1], producedby loose ligation of the sciatic nerve with four chromicgut sutures, has been known as one of the most reliablemodels of neuropathic pain. With CCI of the rat sciaticnerve, animals show symptoms similar to the clinic…     

Chemo-nociceptive signalling from the colon is enhanced by mild colitis and blocked by inhibition of transient receptor potential ankyrin 1 channels

Background and purpose:

Transient receptor potential ankyrin 1 (TRPA1) channels are expressed by primary afferent neurones and activated by irritant chemicals including allyl isothiocyanate (AITC). Here we investigated whether intracolonic AITC causes afferent input to the spinal cord and whether this response is modified by mild colitis, morphine or a TRPA1 channel blocker.

Experimental approach:

One hour after intracolonic administration of AITC to female mice, afferent signalling was visualized by expression of c-Fos in laminae I–II_o of the spinal dorsal horn at sacral segment S1. Mild colitis was induced by dextran sulphate sodium (DSS) added to drinking water for 1 week.

Key results:

Relative to vehicle, AITC (2%) increased expression of c-Fos in the spinal cord. Following induction of mild colitis by DSS (2%), spinal c-Fos responses to AITC, but not vehicle, were augmented by 41%. Colonic inflammation was present (increased myeloperoxidase content and disease activity score), whereas colonic histology, locomotion, feeding and drinking remained unchanged. Morphine (10 mg·kg⁻¹) or the TRPA1 channel blocker HC-030031 (300 mg·kg⁻¹) inhibited the spinal c-Fos response to AITC, in control and DSS-pretreated animals, whereas the response to intracolonic capsaicin (5%) was blocked by morphine but not HC-030031.

Conclusions and implications:

Activation of colonic TRPA1 channels is signalled to the spinal cord. Mild colitis enhanced this afferent input that, as it is sensitive to morphine, is most likely of a chemonociceptive nature. As several irritant chemicals can be present in chyme, TRPA1 channels may mediate several gastrointestinal pain conditions.     

Dexmedetomidine blocks thermal hyperalgesia and spinal glial activation in rat model of monoarthritis

Aim:

To investigate the effect of systemic administration dexmedetomidine, a selective alpha 2 adrenergic receptor (α₂AR) agonist, on thermal hyperalgesia and spinal glial activation evoked by monoarthritis (MA).

Methods:

MA was induced by an intra-articular injection of complete Freund''s adjuvant (CFA). Thermal hyperalgesia was measured by Hargreaves'' test. The spinal glial activation status was analyzed by GFAP (an astrocytic marker) and Iba-1 (a microglial marker) immunohistochemistry or immunoblotting.

Results:

Unilateral intra-articular injection of CFA produced a robust glial activation of astrocytes and microglia in the spinal cord, which was associated with the development and maintenance of thermal hyperalgesia. Intraperitoneal (ip) injection of dexmedetomidine (2.5 and 10 μg/kg) was repeatedly given once daily for 5 days with the first injection 60 min before intra-articular CFA. At the dose of 10 μg/kg, dexmedetomidine significantly attenuated MA-induced ipsilateral hyperalgesia from day 2 to day 5. MA-induced up-regulation of GFAP expression on both sides of the spinal dorsal horn was significantly suppressed by day 5 post-MA following dexmedetomidine application, whereas MA-induced Iba-1 up-regulation was only partially suppressed.

Conclusion:

Systemic dexmedetomidine inhibits the activation of spinal glia, which is possibly associated with its antihyperalgesia in monoarthritic rats.     

Antinociceptive effects of NCX-701 (nitro-paracetamol) in neuropathic rats: enhancement of antinociception by co-administration with gabapentin

Background and purpose:

Neuropathic pain is characterized by a poor response to classic analgesics. In the present study, we have assessed the antinociceptive activity of NCX-701 (nitro-paracetamol) in neuropathic rats, after systemic and intrathecal (i.t.) administration. In addition, we analysed the possible benefit of the combination of NCX-701 and gabapentin, a well-known potent analgesic, in the treatment of neuropathic pain

Experimental approach:

The antinociceptive effects of i.v. and i.t. NCX-701 and paracetamol were studied in spinal cord neuronal responses from neuropathic adult male Wistar rats, using the recording of single motor units technique. The effect of i.v. and i.t. NCX-701 in combination with i.v. gabapentin was studied by isobolographic analysis.

Key results:

The experiments showed that NCX-701, but not paracetamol, dose-dependently reduced the nociceptive responses evoked by noxious mechanical and electrical stimulation, after i.v. (ID₅₀ 542 ± 5 µmol·kg⁻¹ for noxious mechanical stimulation) or i.t. (ID₅₀ 932 ± 16 nmol·kg⁻¹) administration. The combined administration of i.v. or i.t. NCX-701 and i.v. gabapentin induced a more intense antinociceptive effect than any of the two drugs given alone. The isobolographic analysis showed a synergistic effect.

Conclusions and implications:

NCX-701 is an effective antinociceptive compound in situations of neuropathy-induced sensitization, with an action mainly located in the spinal cord. The combination of NCX-701 and gabapentin induces a synergistic enhancement of the depression of nociceptive responses evoked by natural noxious stimulation. The use of NCX-701 alone or in combination with gabapentin might open up new and promising perspectives in the treatment of neuropathic pain.     

Effects of intrathecal epigallocatechin gallate, an inhibitor of Toll-like receptor 4, on chronic neuropathic pain in rats

Numerous studies revealed that spinal inflammation and immune response play an important role in neuropathic pain. In this study, we investigated the effects of intrathecal injection of a Toll-like receptor (TLR4) inhibitor epigallocatechin gallate (EGCG) on neuropathic pain induced by chronic constriction injury of the sciatic nerve (CCI). A total of 120 rats were randomly assigned into 4 groups: sham-operated group, CCI group, CCI plus normal saline group and CCI plus EGCG group. CCI and sham surgeries were performed and both thermal hyperalgesia and mechanical allodynia were tested. Lumbar spinal cord was sampled and the mRNA and protein expressions of TLR4 and High Mobility Group 1 protein (HMGB1) were detected, the contents of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-10 (IL-10) were measured by ELISA, and immunohistochemistry for nuclear factor kappa B (NF-κB) was also carried out. When compared with the sham group, both mechanical and heat pain thresholds were significantly decreased, and the mRNA and protein expressions of TLR4 and HMGB1, the contents of TNF-α, IL-1β and IL-10 in the spinal cords and NF-κB expression in the spinal dorsal horn were markedly increased in CCI rats (P<0.05). After intrathecal injection of EGCG (1mg/kg) once daily from 1day before to 3days after CCI surgery, the expressions of TLR4, NF-κB, HMGB1, TNF-α and IL-1β were markedly decreased while the content of IL-10 in the spinal cord increased significantly accompanied by dramatical improvement of pain behaviors in CCI rats (P<0.05). These results show that the TLR4 signaling pathway plays an important role in the occurrence and development of neuropathic pain, and the therapy targeting TLR4 might be a novel strategy in the treatment of neuropathic pain.     

Antihyperalgesic effects of imidazoline I2 receptor ligands in rat models of inflammatory and neuropathic pain

Background and Purpose

A new imidazoline I₂ receptor ligand, CR4056, is effective for chronic inflammatory pain and diabetic neuropathy. However, it is unclear whether other I₂ receptor ligands have similar effects and whether antinociceptive tolerance develops with repeated treatment.

Experimental Approach

The Von Frey filament test was used to measure mechanical hyperalgesia and the plantar test to measure thermal hyperalgesia in rats injected with complete Freund''s adjuvant (CFA) treatment or had undergone surgery to induce chronic constriction injury (CCI), models of inflammatory pain and peripheral neuropathic pain respectively. The effects of morphine and I₂ receptor ligands, 2-BFI, BU224, tracizoline and CR4056, 3.2–32 mg·kg−1, i.p., on hyperalgesia or affective pain (as measured by a place escape/avoidance paradigm) were studied in separate experiments.

Key Results

Morphine and the I₂ receptor ligands (2-BFI, BU224 and tracizoline) all dose-dependently attenuated mechanical and thermal hyperalgesia in CFA-treated rats. The anti-hyperalgesic effects of 2-BFI in CFA-treated and CCI rats were attenuated by the I₂ receptor antagonist idazoxan. The combination of 2-BFI and morphine produced additive effects against mechanical hyperalgesia in CFA-treated rats. Repeated treatment (daily for 7–9 days) with 2-BFI or CR4056 did not produce antinociceptive tolerance in CFA-treated or CCI rats. Morphine and the I₂ receptor ligands (2-BFI, BU224 and CR4056) were all effective at attenuating place escape/avoidance behaviour in CFA-treated rats.

Conclusions and Implications

Imidazoline I₂ receptor ligands have antihyperalgesic effects in rat models of inflammatory and neuropathic pain and may represent a new class of pharmacotherapeutics for the management of chronic pain.     

Effect of Mas-related gene (Mrg) receptors on hyperalgesia in rats with CFA-induced inflammation via direct and indirect mechanisms

Background and Purpose

Mas oncogene-related gene (Mrg) receptors are exclusively distributed in small-sized neurons in trigeminal and dorsal root ganglia (DRG). We investigated the effects of MrgC receptor activation on inflammatory hyperalgesia and its mechanisms.

Experimental Approach

A selective MrgC receptor agonist, bovine adrenal medulla peptide 8-22 (BAM8-22) or melanocyte-stimulating hormone (MSH) or the μ-opioid receptor (MOR) antagonist CTAP was administered intrathecally (i.t.) in rats injected with complete Freund''s adjuvant (CFA) in one hindpaw. Thermal and mechanical nociceptive responses were assessed. Neurochemicals were measured by immunocytochemistry, Western blot, ELISA and RT-PCR.

Key Results

CFA injection increased mRNA for MrgC receptors in lumbar DRG. BAM8-22 or MSH, given i.t., generated instant short and delayed long-lasting attenuations of CFA-induced thermal hyperalgesia, but not mechanical allodynia. These effects were associated with decreased up-regulation of neuronal NOS (nNOS), CGRP and c-Fos expression in the spinal dorsal horn and/or DRG. However, i.t. administration of CTAP blocked the induction by BAM8-22 of delayed anti-hyperalgesia and inhibition of nNOS and CGRP expression in DRG. BAM8-22 also increased mRNA for MORs and pro-opiomelanocortin, along with β-endorphin content in the lumbar spinal cord and/or DRG. MrgC receptors and nNOS were co-localized in DRG neurons.

Conclusions and Implications

Activation of MrgC receptors suppressed up-regulation of pronociceptive mediators and consequently inhibited inflammatory pain, because of the activation of up-regulated MrgC receptors and subsequent endogenous activity at MORs. The uniquely distributed MrgC receptors could be a novel target for relieving inflammatory pain.     

Glycyrrhizin attenuates rat ischemic spinal cord injury by suppressing inflammatory cytokines and HMGB1

Aim:

To investigate the neuroprotective effect of glycyrrhizin (Gly) against the ischemic injury of rat spinal cord and the possible role of the nuclear protein high-mobility group box 1 (HMGB1) in the process.

Methods:

Male Sprague-Dawley rats were subjected to 45 min aortic occlusion to induce transient lumbar spinal cord ischemia. The motor functions of the animals were assessed according to the modified Tarlov scale. The animals were sacrificed 72 h after reperfusion and the lumbar spinal cord segment (L2–L4) was taken out for histopathological examination and Western blotting analysis. Serum inflammatory cytokine and HMGB1 levels were analyzed using ELISA.

Results:

Gly (6 mg/kg) administered intravenously 30 min before inducing the transient lumbar spinal cord ischemia significantly improved the hind-limb motor function scores, and reduced the number of apoptotic neurons, which was accompanied by reduced levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in the plasma and injured spinal cord. Moreover, the serum HMGB1 level correlated well with the serum TNF-α, IL-1β and IL-6 levels during the time period of reperfusion.

Conclusion:

The results suggest that Gly can attenuate the transient spinal cord ischemic injury in rats via reducing inflammatory cytokines and inhibiting the release of HMGB1.     

Pharmacological properties of S1RA, a new sigma-1 receptor antagonist that inhibits neuropathic pain and activity-induced spinal sensitization

BACKGROUND AND PURPOSE

The sigma-1 (σ₁) receptor is a ligand-regulated molecular chaperone that has been involved in pain, but there is limited understanding of the actions associated with its pharmacological modulation. Indeed, the selectivity and pharmacological properties of σ₁ receptor ligands used as pharmacological tools are unclear and the demonstration that σ₁ receptor antagonists have efficacy in reversing central sensitization-related pain sensitivity is still missing.

EXPERIMENTAL APPROACH

The pharmacological properties of a novel σ₁ receptor antagonist (S1RA) were first characterized. S1RA was then used to investigate the effect of pharmacological antagonism of σ₁ receptors on in vivo nociception in sensitizing conditions and on in vitro spinal cord sensitization in mice. Drug levels and autoradiographic, ex vivo binding for σ₁ receptor occupancy were measured to substantiate behavioural data.

KEY RESULTS

Formalin-induced nociception (both phases), capsaicin-induced mechanical hypersensitivity and sciatic nerve injury-induced mechanical and thermal hypersensitivity were dose-dependently inhibited by systemic administration of S1RA. Occupancy of σ₁ receptors in the CNS was significantly correlated with the antinociceptive effects. No pharmacodynamic tolerance to the antiallodynic and antihyperalgesic effect developed following repeated administration of S1RA to nerve-injured mice. As a mechanistic correlate, electrophysiological recordings demonstrated that pharmacological antagonism of σ₁ receptors attenuated the wind-up responses in spinal cords sensitized by repetitive nociceptive stimulation.

CONCLUSIONS AND IMPLICATIONS

These findings contribute to evidence identifying the σ₁ receptor as a modulator of activity-induced spinal sensitization and pain hypersensitivity, and suggest σ₁ receptor antagonists as potential novel treatments for neuropathic pain.     

Lack of effect of chronic pre-treatment with the FAAH inhibitor URB597 on inflammatory pain behaviour: evidence for plastic changes in the endocannabinoid system

BACKGROUND AND PURPOSE

Elevating levels of endocannabinoids with inhibitors of fatty acid amide hydrolase (FAAH) is a major focus of pain research, purported to be a safer approach devoid of cannabinoid receptor-mediated side effects. Here, we have determined the effects of sustained pharmacological inhibition of FAAH on inflammatory pain behaviour and if pharmacological inhibition of FAAH was as effective as genetic deletion of FAAH on pain behaviour.

EXPERIMENTAL APPROACH

Effects of pre-treatment with a single dose, versus 4 day repeated dosing with the selective FAAH inhibitor, URB597 (i.p. 0.3 mg·kg⁻¹), on carrageenan-induced inflammatory pain behaviour and spinal pro-inflammatory gene induction were determined in rats. Effects of pain induction and of the drug treatments on levels of arachidonoyl ethanolamide (AEA), palmitoyl ethanolamide (PEA) and oleolyl ethanolamide (OEA) in the spinal cord were determined.

KEY RESULTS

Single, but not repeated, URB597 treatment significantly attenuated the development of inflammatory hyperalgesia (P < 0.001, vs. vehicle-treated animals). Neither mode of URB597 treatment altered levels of AEA, PEA and OEA in the hind paw, or carrageenan-induced paw oedema. Single URB597 treatment produced larger increases in AEA, PEA and OEA in the spinal cord, compared with those after repeated administration. Single and repeated URB597 treatment decreased levels of immunoreactive N-acylphosphatidylethanolamine phospholipase D (NAPE-PLD) in the spinal cord and attenuated carrageenan-induced spinal pro-inflammatory gene induction.

CONCLUSION AND IMPLICATIONS

Changes in the endocannabinoid system may contribute to the loss of analgesic effects following repeated administration of low dose URB597 in this model of inflammatory pain.