Neurobiology of Pain最新文献

Globally, it is estimated that one in five people suffer from chronic pain, with prevalence increasing with age. The pathophysiology of chronic pain encompasses complex sensory, immune, and inflammatory interactions within both the central and peripheral nervous systems. Microglia, the resident macrophages of the central nervous system (CNS), are critically involved in the initiation and persistence of chronic pain. Microglia respond to local signals from the CNS but are also modulated by signals from the gastrointestinal tract. Emerging data from preclinical and clinical studies suggest that communication between the gut microbiome, the community of bacteria residing within the gut, and microglia is involved in producing chronic pain. Targeted strategies that manipulate or restore the gut microbiome have been shown to reduce microglial activation and alleviate symptoms associated with inflammation. These data indicate that manipulations of the gut microbiome in chronic pain patients might be a viable strategy in improving pain outcomes. Herein, we discuss the evidence for a connection between microglia and the gut microbiome and explore the mechanisms by which commensal bacteria might influence microglial reactivity to drive chronic pain.

Previous studies have shown that oral administration of the NMDAR modulator NYX-2925 alleviates pain in several animal models of neuropathic pain and this appears to be through mPFC, but not spinal, mediated mechanisms. While much is known about the impact of neuropathic pain on NMDAR-mediated signaling in the spinal cord, limited studies have focused on the brain. In the current study, we assess signaling changes associated with NMDAR-mediated plasticity in the mPFC and the impact of NYX-2925 administration on the normalization of these signaling changes. We found a decrease in activated Src levels in the mPFC of animals with chronic constriction injury (CCI) of the sciatic nerve. While Src mediated activation of NMDARs was also decreased in CCI animals, the main NMDAR phosphorylation site of CAMKII was not affected. This is in opposition to what has been found in the spinal cord, where both Src and CAMKII activation are increased. Oral administration of NYX-2925 restored levels of activated Src and Src phosphorylation sites on GluN2A and GluN2B in the mPFC, with no effect on activated CAMKII levels. The analgesic effect of NYX-2925 appears dependent on this restoration of Src activation in the mPFC, as co-administering Src activation inhibitors prevented the NYX-2925 analgesic effect. Overall, these data suggest that NMDAR-mediated signaling plays a key role in neuropathic pain, albeit in different directions in the spinal cord vs. the mPFC. Furthermore, the analgesic effect of NYX-2925 appears to involve a restoration of NMDAR-mediated signaling in the mPFC.

Endogenous lipid mediators are proposed to contribute to headache and facial pain by activating trigeminal neurons (TN). We recently identified 11-hydroxy-epoxide- and 11-keto-epoxide derivatives of linoleic acid (LA) that are present in human skin and plasma and potentially contribute to nociception. Here we expand upon initial findings by examining the effects of 11-hydroxy- and 11-keto-epoxide-LA derivatives on TN activation in comparison to LA, the LA derivative [9-hydroxy-octadecadienoic acid (9-HODE)] and prostaglandin E₂ (PGE₂). 11-hydroxy- and 11-keto-epoxide-LA derivatives elicited Ca²⁺ transients in TN subpopulations. The proportion of neurons responding to test compounds (5 μM, 5 min) ranged from 16.2 ± 3.8 cells (11 K-9,10E-LA) to 34.1 ± 2.4 cells (11H-12,13E-LA). LA and 9-HODE (5 μM, 5 min) elicited responses in 11.6 ± 3.1% and 9.7 ± 3.4% of neurons, respectively. 11H-12,13E-LA, 11K-12,13E-LA, and 11H-9,10E-LA produced Ca²⁺ responses in significantly higher proportions of neurons compared to either LA or 9-HODE (F (6, 36) = 5.12, P = 0.0007). 11H-12,13E-LA and 11H-9,10E-LA increased proportions of responsive neurons in a concentration-dependent fashion, similar to PGE₂. Most sensitive neurons responded to additional algesic agents (32.9% to capsaicin, 40.1% to PGE₂, 58.0% to AITC), however 20.6% did not respond to any other agent. In summary, 11-hydroxy-epoxide derivatives of LA increase trigeminal neuron excitability, suggesting a potential role in headache or facial pain.

Chronic pain following spinal cord injury (SCI) is associated with electrical hyperactivity (spontaneous and evoked) in primary nociceptors. Cyclic adenosine monophosphate (cAMP) signaling is an important contributor to nociceptor excitability, and knockdown of the cAMP effector, exchange protein activated by cAMP (EPAC), has been shown to relieve pain-like responses in several chronic pain models. To examine potentially distinct roles of each EPAC isoform (EPAC1 and 2) in maintaining chronic pain, we used rat and mouse models of contusive spinal cord injury (SCI). Pharmacological inhibition of EPAC1 or 2 in a rat SCI model was sufficient to reverse SCI-induced nociceptor hyperactivity, indicating that EPAC1 and 2 signaling activity are complementary, with both required to maintain hyperactivity. However, EPAC activation was not sufficient to induce similar hyperactivity in nociceptors from naïve rats, and we observed no change in EPAC protein expression after SCI. In the mouse SCI model, inhibition of both EPAC isoforms through a combination of pharmacological inhibition and genetic deletion was required to reverse SCI-induced nociceptor hyperactivity. This was consistent with our finding that neither EPAC1^−/− nor EPAC2^−/− mice were protected against SCI-induced chronic pain as assessed with an operant mechanical conflict test. Thus, EPAC1 and 2 activity may play a redundant role in mouse nociceptors, although no corresponding change in EPAC protein expression levels was detected after SCI. Despite some differences between these species, our data demonstrate a fundamental role for both EPAC1 and EPAC2 in mechanisms maintaining nociceptor hyperactivity and chronic pain after SCI.

The evolution of peripheral and central changes following a peripheral nerve injury imply the onset of afferent signals that affect the brain. Changes to inflammatory processes may contribute to peripheral and central alterations such as altered psychological state and are not well characterized in humans. We focused on four elements that change peripheral and central nervous systems following ankle injury in 24 adolescent patients and 12 age-sex matched controls. Findings include (a) Changes in tibial, fibular, and sciatic nerve divisions consistent with neurodegeneration; (b) Changes within the primary motor and somatosensory areas as well as higher order brain regions implicated in pain processing; (c) Increased expression of fear of pain and pain reporting; and (d) Significant changes in cytokine profiles relating to neuroinflammatory signaling pathways. Findings address how changes resulting from peripheral nerve injury may develop into chronic neuropathic pain through changes in the peripheral and central nervous system.

Background and purpose

Calcitonin gene-related peptide (CGRP) plays an important role in migraine pathophysiology. CGRP acts primarily by activating a receptor composed of 3 proteins: calcitonin receptor-like receptor (CLR), receptor activity-modifying protein 1 (RAMP1), and receptor component protein (RCP). We tested the hypothesis that sex differences exist in protein levels of two key components of this CGRP receptor: CLR and RCP.

Methods

We used specific antibodies to assess baseline protein levels of CLR and RCP in the spinal trigeminal nucleus caudalis (SpVc) and upper cervical spinal cord of both male and female rats. We also tested if manipulations that knock-down the expression of RCP in SpVc, using locally-mediated gene transfer of short hairpin RNA (shRNA), ameliorate pain in an animal model of intracranial migraine-like pain induced by chemical noxious stimulation of the meninges. To assess pain, we used tests of ongoing pain (rat face grimace test and freezing behavior) and tests of facial mechanical hypersensitivity and allodynia.

Results

There was no difference in CLR levels between male and female animals (p > 0.11) in SpVc and the upper cervical cord. However, female animals exhibited greater baseline levels of RCP (up to 3-fold higher) compared to males (p < 0.002). The knock-down of RCP expression in SpVc attenuated mechanical facial allodynia induced by chemical noxious stimulation of the meninges, but had little effect on ongoing pain behaviors in female and male animals.

Conclusions

RCP is an integral component of the CGRP receptor and may play a key role in mediating CGRP induced central sensitization after noxious stimulation of the meninges. RCP expression in the SpVc and upper cervical cord is sexually dimorphic, with higher levels of expression in females. This dimorphism may be related to the increased incidence of migraines in females–a hypothesis that should be tested in the future.

The cyclic nucleotide signaling, including cAMP-PKA and cGMP-PKG pathways, has been well known to play critical roles in regulating cellular growth, metabolism and many other intracellular processes. In recent years, more and more studies have uncovered the roles of cAMP and cGMP in the nervous system. The cAMP and cGMP signaling mediates chronic pain induced by different forms of injury and stress. Here we summarize the roles of cAMP-PKA and cGMP-PKG signaling pathways in the pathogenesis of chronic pain after nerve injury. In addition, acute dissociation and chronic compression of the dorsal root ganglion (DRG) neurons, respectively, leads to neural hyperexcitability possibly through PAR2 activation-dependent activation of cAMP-PKA pathway. Clinically, radiotherapy can effectively alleviate bone cancer pain at least partly through inhibiting the cancer cell-induced activation of cAMP-PKA pathway. Roles of cyclic nucleotide signaling in neuropathic and inflammatory pain are also seen in many other animal models and are involved in many pro-nociceptive mechanisms including the activation of hyperpolarization-activated cyclic nucleotide (HCN)-modulated ion channels and the exchange proteins directly activated by cAMP (EPAC). Further understanding the roles of cAMP and cGMP signaling in the pathogenesis of chronic pain is theoretically significant and clinically valuable for treatment of chronic pain.

Fibroblast Growth Factor Homologous Factors (FHF) constitute a subfamily of FGF proteins with four prototypes (FHF1-4; also known as FGF11-14). FHF proteins have been shown to bind directly to the membrane-proximal segment of the C-terminus in voltage-gated sodium channels (Nav), and regulate current density, availability, and frequency-dependent inhibition of sodium currents. Members of the FHF2 subfamily, FHF2A and FHF2B, differ in the length and sequence of their N-termini, and, importantly, differentially regulate Nav1.6 gating properties. Using immunohistochemistry, we show that FHF2 isoforms are expressed in adult dorsal root ganglion (DRG) neurons where they co-localize with Nav1.6 and Nav1.7. FHF2A and FHF2B show differential localization in neuronal compartments in DRG neurons, and levels of expression of FHF2 factors are down-regulated following sciatic nerve axotomy. Because Nav1.7 in nociceptors plays a critical role in pain, we reasoned that its interaction with FHF2 isoforms might regulate its current properties. Using whole-cell patch clamp in heterologous expression systems, we show that the expression of FHF2A in HEK293 cell line stably expressing Nav1.7 channels causes no change in activation, whereas FHF2B depolarizes activation. Both FHF2 isoforms depolarize fast-inactivation. Additionally, FHF2A causes an accumulation of inactivated channels at all frequencies tested due to a slowing of recovery from inactivation, whereas FHF2B has little effect on these properties of Nav1.7. Measurements of the Nav1.7 current in DRG neurons in which FHF2 levels are knocked down confirmed the effects of FHF2A on repriming, and FHF2B on activation, however FHF2A and B did not have an effect on fast inactivation. Our data demonstrates that FHF2 does indeed regulate the current properties of Nav1.7 and does so in an isoform and cell-specific manner.

Chronic itch is a debilitating condition characterised by excessive scratching and is a symptom frequently reported in skin diseases such as atopic dermatitis. It has been proposed that release of the cysteine protease Cathepsin S (CatS) from skin keratinocytes or immune cells resident in or infiltrating the skin could act as a pruritogen in chronic itch conditions. CatS is known to activate protease-activated receptor 2 (PAR2). We therefore hypothesised that enzymatic activation of neuronally expressed PAR2 by CatS was responsible for activation of sensory neurons and transmission of itch signals. Intradermally-injected human recombinant (hr)-CatS or the PAR2 agonist, SLIGRL-NH₂ behaved as pruritogens by causing scratching behaviour in mice. Hr-CatS-induced scratching behaviour was prevented by CatS inhibitors and PAR2 antagonists and reduced by 50% in TRPV1^−/− mice compared with wild-type mice, whilst no significant reduction in scratching behaviour was observed in TRPA1^−/− mice. Cultured dorsal root ganglion (DRG) cells showed an increase in [Ca²⁺]_i following incubation with hr-CatS, and the percentage of neurons that responded to hr-CatS decreased in the presence of a PAR2 antagonist or in cultures of neurons from TRPV1^−/− mice. Taken together, our results indicate CatS acts as a pruritogen via PAR2 activation in TRPV1-expressing sensory neurons.

Objective

Disability and movement-related pain are major symptoms of joint disease, motivating the development of methods to quantify motor behaviour in rodent joint pain models. We compared effects on behaviour, assessed the levels of biochemical mediators and made a detailed histopathological evaluation after induction of rat monoiodoacetate (MIA) monoarthritis into the ankle or knee joint.

Design

Twenty-seven male Lewis rats were used. Before and up to 28 days after induction, they were tested for weight bearing during walking (dynamic), and standing (static), and for mechanical sensitivity. At termination synovial fluid was taken from ankle and/or knee joints for analysis of monocyte chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6), macrophage inflammatory protein 3 alpha (MIP-3α), keratinocyte chemoattractant (KC)/human growth-regulated oncogene (GRO) and L(+)-lactate, and from separate rats joints were collected for histopathological assessment.

Results

MIA ankle joint injection gave a marked reduction of dynamic weight bearing during the first days, not seen in rats with knee joint injection. At three weeks, it was decreased in the group with knee injection, but not in those with ankle injection. However, the different injection sites caused similar reductions in static weight bearing during the early phase, which was normalized in the group with ankle injection but continued and was strengthened with time in the knee injected group. Histopathological assessment, biochemical mediators and joint swelling confirmed the disparate profiles.

Conclusions

This work shows that ankle versus knee joint injection of MIA resulted in different profiles in rats, which may mirror what has been found in human patients with osteoarthritis.