Role In Allodynia Research Articles

Altered Intrinsic Properties and Inhibitory Connectivity in Aged Parvalbumin-Expressing Dorsal Horn Neurons.

The incidence of pain symptoms such as allodynia are known to increase with age. Parvalbumin expressing interneurons (PVINs) within the dorsal horn (DH) of the spinal cord play an important role in allodynia whereby their inhibitory connections prevent innocuous touch information from exciting nociceptive pathways. Here we ask whether the functional properties of PVINs are altered by aging, comparing their functional properties in adult (3–7 month) and aged mice (23–28 month). Patch clamp recordings were made from PVINs in laminae IIi-III of parasagittal spinal cord slices. The intrinsic excitability of PVINs changed with age. Specifically, AP discharge shifted from initial bursting to tonic firing, and firing duration during current injection increased. The nature of excitatory synaptic input to PVINs also changed with age with larger but less frequent spontaneous excitatory currents occurring in aged mice, however, the net effect of these differences produced a similar level of overall excitatory drive. Inhibitory drive was also remarkably similar in adult and aged PVINs. Photostimulation of ChR2 expressing PVINs was used to study inhibitory connections between PVINs and unidentified DH neurons and other PVINs. Based on latency and jitter, monosynaptic PVIN to unidentified-cell and PVIN-PVIN connections were compared in adult and aged mice, showing that PVIN to unidentified-cell connection strength increased with age. Fitting single or double exponentials to the decay phase of IPSCs showed there was also a shift from mixed (glycinergic and GABAergic) to GABAergic inhibitory transmission in aged animals. Overall, our data suggest the properties of PVIN neurons in aged animals enhance their output in spinal circuits in a manner that would blunt allodynia and help maintain normal sensory experience during aging.

Calmodulin Supports TRPA1 Channel Association with Opioid Receptors and Glutamate NMDA Receptors in the Nervous Tissue.

Transient receptor potential ankyrin member 1 (TRPA1) belongs to the family of thermo TRP cation channels that detect harmful temperatures, acids and numerous chemical pollutants. TRPA1 is expressed in nervous tissue, where it participates in the genesis of nociceptive signals in response to noxious stimuli and mediates mechanical hyperalgesia and allodynia associated with different neuropathies. The glutamate N-methyl-d-aspartate receptor (NMDAR), which plays a relevant role in allodynia to mechanical stimuli, is connected via histidine triad nucleotide-binding protein 1 (HINT1) and type 1 sigma receptor (σ1R) to mu-opioid receptors (MORs), which mediate the most potent pain relief. Notably, neuropathic pain causes a reduction in MOR antinociceptive efficacy, which can be reversed by blocking spinal NMDARs and TRPA1 channels. Thus, we studied whether TRPA1 channels form complexes with MORs and NMDARs that may be implicated in the aforementioned nociceptive signals. Our data suggest that TRPA1 channels functionally associate with MORs, delta opioid receptors and NMDARs in the dorsal root ganglia, the spinal cord and brain areas. These associations were altered in response to pharmacological interventions and the induction of inflammatory and also neuropathic pain. The MOR-TRPA1 and NMDAR-TRPA1 associations do not require HINT1 or σ1R but appear to be mediated by calcium-activated calmodulin. Thus, TRPA1 channels may associate with NMDARs to promote ascending acute and chronic pain signals and to control MOR antinociception.

Reciprocal Regulatory Interaction between TRPV1 and Kinin B1 Receptor in a Rat Neuropathic Pain Model.

Kinins are mediators of pain and inflammation and evidence suggests that the inducible kinin B1 receptor (B1R) is involved in neuropathic pain (NP). This study investigates whether B1R and TRPV1 are colocalized on nociceptors and/or astrocytes to enable regulatory interaction either directly or through the cytokine pathway (IL-1β, TNF-α) in NP. Sprague Dawley rats were subjected to unilateral partial sciatic nerve ligation (PSNL) and treated from 14 to 21 days post-PSNL with antagonists of B1R (SSR240612, 10 mg·kg−1, i.p.) or TRPV1 (SB366791, 1 mg·kg−1, i.p.). The impact of these treatments was assessed on nociceptive behavior and mRNA expression of B1R, TRPV1, TNF-α, and IL-1β. Localization on primary sensory fibers, astrocytes, and microglia was determined by immunofluorescence in the lumbar spinal cord and dorsal root ganglion (DRG). Both antagonists suppressed PSNL-induced thermal hyperalgesia, but only SB366791 blunted mechanical and cold allodynia. SSR240612 reversed PSNL-induced enhanced protein and mRNA expression of B1R and TRPV1 mRNA levels in spinal cord while SB366791 further increased B1R mRNA/protein expression. B1R and TRPV1 were found in non-peptide sensory fibers and astrocytes, and colocalized in the spinal dorsal horn and DRG, notably with IL-1β on astrocytes. IL-1β mRNA further increased under B1R or TRPV1 antagonism. Data suggest that B1R and TRPV1 contribute to thermal hyperalgesia and play a distinctive role in allodynia associated with NP. Close interaction and reciprocal regulatory mechanism are suggested between B1R and TRPV1 on astrocytes and nociceptors in NP.

A novel interaction between CX3CR1 and CCR2 signalling in monocytes constitutes an underlying mechanism for persistent vincristine-induced pain

BackgroundA dose-limiting side effect of chemotherapeutic agents such as vincristine (VCR) is neuropathic pain, which is poorly managed at present. Chemokine-mediated immune cell/neuron communication in preclinical VCR-induced pain forms an intriguing basis for the development of analgesics. In a murine VCR model, CX3CR1 receptor-mediated signalling in monocytes/macrophages in the sciatic nerve orchestrates the development of mechanical hypersensitivity (allodynia). CX3CR1-deficient mice however still develop allodynia, albeit delayed; thus, additional underlying mechanisms emerge as VCR accumulates. Whilst both patrolling and inflammatory monocytes express CX3CR1, only inflammatory monocytes express CCR2 receptors. We therefore assessed the role of CCR2 in monocytes in later stages of VCR-induced allodynia.MethodsMechanically evoked hypersensitivity was assessed in VCR-treated CCR2- or CX3CR1-deficient mice. In CX3CR1-deficient mice, the CCR2 antagonist, RS-102895, was also administered. Immunohistochemistry and Western blot analysis were employed to determine monocyte/macrophage infiltration into the sciatic nerve as well as neuronal activation in lumbar DRG, whilst flow cytometry was used to characterise monocytes in CX3CR1-deficient mice. In addition, THP-1 cells were used to assess CX3CR1-CCR2 receptor interactions in vitro, with Western blot analysis and ELISA being used to assess expression of CCR2 and proinflammatory cytokines.ResultsWe show that CCR2 signalling plays a mechanistic role in allodynia that develops in CX3CR1-deficient mice with increasing VCR exposure. Indeed, the CCR2 antagonist, RS-102895, proves ineffective in mice possessing functional CX3CR1 receptors but reduces VCR-induced allodynia in CX3CR1-deficient mice, in which CCR2+ monocytes are elevated by VCR. We suggest that a novel interaction between CX3CR1 and CCR2 receptors in monocytes accounts for the therapeutic effect of RS-102895 in CX3CR1-deficient mice. Indeed, we observe that CCR2, along with its ligand, CCL2, is elevated in the sciatic nerve in CX3CR1-deficient mice, whilst in THP-1 cells (human monocytes), downregulating CX3CR1 upregulates CCR2 expression via p38 MAP kinase signalling. We also show that the CX3CR1-CCR2 interaction in vitro regulates the release of pronociceptive cytokines TNF-α and IL1β.ConclusionsOur data suggests that CCL2/CCR2 signalling plays a crucial role in VCR-induced allodynia in CX3CR1-deficient mice, which arises as a result of an interaction between CX3CR1 and CCR2 in monocytes.

Reciprocal relationship between membrane type 1 matrix metalloproteinase and the algesic peptides of myelin basic protein contributes to chronic neuropathic pain

Myelin basic protein (MBP) is an auto-antigen able to induce intractable pain from innocuous mechanical stimulation (mechanical allodynia). The mechanisms provoking this algesic MBP activity remain obscure. Our present study demonstrates that membrane type 1 matrix metalloproteinase (MT1-MMP/MMP-14) releases the algesic MBP peptides from the damaged myelin, which then reciprocally enhance the expression of MT1-MMP in nerve to sustain a state of allodynia. Specifically, MT1-MMP expression and activity in rat sciatic nerve gradually increased starting at day 3 after chronic constriction injury (CCI). Inhibition of the MT1-MMP activity by intraneural injection of the function-blocking human DX2400 monoclonal antibody at day 3 post-CCI reduced mechanical allodynia and neuropathological signs of Wallerian degeneration, including axon demyelination, degeneration, edema and formation of myelin ovoids. Consistent with its role in allodynia, the MT1-MMP proteolysis of MBP generated the MBP69-86-containing epitope sequences in vitro. In agreement, the DX2400 therapy reduced the release of the MBP69-86 epitope in CCI nerve. Finally, intraneural injection of the algesic MBP69-86 and control MBP2-18 peptides differentially induced MT1-MMP and MMP-2 expression in the nerve. With these data we offer a novel, self-sustaining mechanism of persistent allodynia via the positive feedback loop between MT1-MMP and the algesic MBP peptides. Accordingly, short-term inhibition of MT1-MMP activity presents a feasible pharmacological approach to intervene in this molecular circuit and the development of neuropathic pain.

Mechanical allodynia

Mechanical allodynia (other pain) is a painful sensation caused by innocuous stimuli like light touch. Unlike inflammatory hyperalgesia that has a protective role, allodynia has no obvious biological utility. Allodynia is associated with nerve damage in conditions such as diabetes, and is likely to become an increasing clinical problem. Unfortunately, the mechanistic basis of this enhanced sensitivity is incompletely understood. In this review, we describe evidence for the involvement of candidate mechanosensitive channels such as Piezo2 and their role in allodynia, as well as the peripheral and central nervous system mechanisms that have also been implicated in this form of pain. Specific treatments that block allodynia could be very useful if the cell and molecular basis of the condition could be determined. There are many potential mechanisms underlying this condition ranging from alterations in mechanotransduction and sensory neuron excitability to the actions of inflammatory mediators and wiring changes in the CNS. As with other pain conditions, it is likely that the range of redundant mechanisms that cause allodynia will make therapeutic intervention problematic.

Antinociceptive effects of AS1069562, the (+)-isomer of indeloxazine, on spinal hypersensitivity induced by intrathecal injection of prostaglandin in mice: Comparison with duloxetine and amitriptyline

The (+)-isomer of indeloxazine AS1069562 exerts multiple pharmacological actions including the inhibition of serotonin (5-HT) and norepinephrine reuptake and analgesia in experimental animal pain models. Here, we evaluated the antinociceptive effects of AS1069562 and the antidepressants duloxetine and amitriptyline in mouse models of prostaglandin-induced spinal hypersensitivity. Prostaglandin E2 (PGE2) and F2α (PGF2α) were intrathecally administered to induce spinal hypersensitivity, causing tactile allodynia in mice. Allodynia induced by PGF2α but not by PGE2 was suppressed by desensitization of C-fibers with systemic pretreatment with resiniferatoxin. C-fiber hyperexcitability might therefore play a role in allodynia induced by PGF2α but not PGE2. In the PGE2-induced allodynia model, AS1069562 and duloxetine significantly suppressed allodynia, whereas amitriptyline did not. In the PGF2α-induced allodynia model, AS1069562 and amitriptyline significantly ameliorated allodynia, whereas duloxetine did not. To demonstrate the broad effects of AS1069562 compared to duloxetine, additional studies were conducted to elucidate other target mechanisms of AS1069562 beyond 5-HT and norepinephrine reuptake inhibition. AS1069562 exhibited affinity for both 5-HT1A and 5-HT3 receptors, and the analgesic effect of AS1069562 on PGF2α-induced allodynia was significantly blocked by the 5-HT1A receptor antagonist (S)–WAY100135 and the 5-HT3 receptor agonist SR57227. Taken together, these results indicate that AS1069562 inhibits both C-fiber- and non-C-fiber-dependent prostaglandin-induced allodynia, while duloxetine inhibits only non-C-fiber-triggered allodynia, and amitriptyline inhibits only C-fiber-triggered allodynia. These broad antinociceptive effects of AS1069562 may be due not only to 5-HT and norepinephrine reuptake inhibition but also to its effects on 5-HT receptors such as 5-HT1A and 5-HT3 receptors.

Icilin reduces voltage-gated calcium channel currents in naïve and injured DRG neurons in the rat spinal nerve ligation model

Recently, the transient receptor potential (TRP) channels TRPM8 and TRPA1 have been identified as molecular sensors for cold, and it has been suggested that they play a crucial role in allodynia by modulating voltage-gated calcium channel currents (ICa(V)). The aim of this study was to analyze the modulation of ICa(V) by the TRPM8-agonist icilin in vitro and to investigate the analgesic effect of icilin in a neuropathic pain model in vivo. Whole cell patch-clamp recordings were performed on isolated naïve and injured rat dorsal root ganglia (DRG) neurons, and the analgesic efficacy of icilin applied topically to the paws or intrathecally was tested in rats after spinal nerve ligation (SNL). ICa(V) (depolarization from −80 to 0mV) in naïve DRG neurons was reduced dose dependently (0.002–200µM) by icilin (18–80%). Subtype isolation of calcium channels show a marked reduction of L-type channel currents compared to N-type channel currents. The effects of icilin on ICa(V) were not significantly different in non-injured and SNL-injured DRG neurons. In vivo, neither topical (10–200µM) nor intrathecal application of icilin (0.1nM to 1µM) affected tactile allodynia or thermal hyperalgesia after SNL, but it increases cold allodynia 6h after application. We conclude that the icilin-induced modulation of ICa(V) in DRG neurons is unlikely to mediate analgesic effects or contribute directly to the pathogenesis of cold allodynia in the rat SNL model, but it is a potential mechanism for the analgesic effects of icilin in other pain models.

Spinal ERK activation contributes to the regulation of bladder function in spinal cord injured rats

The extracellular signal-regulated kinase 1 and 2 (ERK) pathway, regulated by phosphorylation on specific amino acids, is emerging as an important signaling cascade in neurones, transducing sensory input into cellular responses. In the mammalian nervous system, the ERK pathway has been found to mediate plasticity events. Particularly, in the spinal cord, ERK play an important role in allodynia and hyperalgesia. Recently, it was demonstrated that ERK activation is upregulated in the spinal cord of rats with chronic bladder inflammation and contributes to bladder overactivity. Thus, in this study we sought to assess the involvement of ERK in micturition reflexes associated to spinal cord injury (SCI) in the rat. Bladder function in chronic SCI rats was altered compared to spinal intact rats. PhosphoERK levels were upregulated in the L6 spinal cord segment, particularly after saline infusion for 2 h. The increase in spinal ERK phosphorylation was specifically restricted to L6 spinal segment. No variation in the levels of total ERK protein was observed. Intrathecal administration of PD98059, a specific inhibitor of ERK phosphorylation, reduced the frequency and amplitude of bladder contractions in SCI animals but not in spinal intact ones. Overall, our results demonstrate increased activation of the ERK pathway in the spinal cord from SCI rats, restricted to spinal segments that receive sensory input arising from the bladder. Since the use of PD98059 reduced the frequency and amplitude of bladder contractions, ERK inhibitors may provide a new therapeutic approach to the treatment of bladder overactivity after spinal injuries.

Mechanisms of central neuropathic pain: a combined psychophysical and fMRI study in syringomyelia

The pathophysiology of central pain syndromes is still poorly understood and their treatment remains a major challenge. It has long been suggested that lesions of the spinothalamic pathways are necessary for developing these pain syndromes. The recently proposed thermosensory disinhibition theory suggests that reduction of the inhibition of thermal sensory afferents that affect nociceptive systems may play a major pathophysiological role. Syringomyelia, which is frequently associated with central neuropathic pain, is characterized by a selective or preferential lesion of the spinothalamic tract resulting in thermosensory deficits of various extents and magnitudes. Thus, syringomyelia represents a unique 'pathological model' particularly suited to investigating the relationship between spinothalamic tract dysfunction, thermosensory deficits and pain. Here, we systematically compared the sensory loss (thermal and mechanical), using quantitative sensory testing, between 46 consecutive syringomyelia patients with or without neuropathic pain. We then further investigated the mechanisms of evoked pains in these patients, using functional MRI (fMRI) in a subgroup of patients with cold or brush-evoked allodynia, compared with patients without pain and healthy volunteers. We found no significant difference in the magnitude or extent of sensory deficits between patients with or without neuropathic pain, suggesting that lesions of the spinothalamic pathways are not sufficient for developing central pain. However, a different pattern of sensory deficits was observed between patients with spontaneous pain only (n = 11) and patients with both spontaneous pain and allodynia (n = 20), suggesting that the mechanisms of central pain are not univocal. In patients with spontaneous pain only, the thermal sensory loss was significantly more asymmetrical and there was a direct relationship between the extent of thermosensory deficits (i.e. deafferentation) and the intensity of burning pain. In contrast, patients with allodynia had reduced thermal deficits, in terms of both magnitude and extent. In addition, the sensory deficits were different between patients with cold or tactile allodynia, suggesting distinct pathophysiological mechanisms related to the sub-modalities of allodynia. Our fMRI study further confirmed this, showing that different sub-types of allodynia were associated with distinct patterns of brain activity, which do not necessarily correspond to the 'pain matrix' involved in acute physiological pain. The prefrontal cortex was the only area consistently activated by pathological evoked pains, suggesting that alteration of high-level pain modulatory mechanisms might play a major role in allodynia due to central lesion.

Neurotrophins from dorsal root ganglia trigger allodynia after spinal nerve injury in rats.

Injury to peripheral nerves often results in chronic pain which is difficult to relieve. The mechanism underlying the pain syndrome remains largely unknown. In previous studies we showed that neurotrophins are up-regulated in satellite cells around sensory neurons following sciatic nerve lesion. In the present study, we have examined whether the neurotrophins in the dorsal root ganglia play any role in allodynia after nerve injury. Antibodies to different neurotrophins, directly delivered to injured dorsal root ganglia, significantly reduced (with different time sequences) the percentage of foot withdrawal responses evoked by von Frey hairs. The antibodies to nerve growth factor acted during the early phase but antibodies to neurotrophin-3 and brain-derived neurotrophic factor were effective during the later phase. Exogenous nerve growth factor or brain-derived neurotrophic factor, but not neurotrophin-3, directly delivered to intact dorsal root ganglia, trigger a persistent mechanical allodynia. Our results showed that neurotrophins within the dorsal root ganglia after peripheral nerve lesion are involved in the generation of allodynia at different stages. These studies provide the first evidence that ganglia-derived neurotrophins are a source of nociceptive stimuli for neuropathic pain after peripheral nerve injury.