Gisela Segond von Banchet

Mechanisms of Pain in Arthritis

Abstract: Inflammation in the joint causes peripheral sensitization (increase of sensitivity of nociceptive primary afferent neurons) and central sensitization (hyperexcitability of nociceptive neurons in the central nervous system). The processes of sensitization are thought to be the basis of arthritic pain that appears as spontaneous pain (joints at rest) and hyperalgesia (augmented pain response on noxious stimulation and pain on normally nonpainful stimulation). Sensitization also facilitates efferent neuronal processes through which the nervous system influences the inflammatory process. Peripheral sensitization is produced by the action of inflammatory mediators such as bradykinin, prostaglandins, neuropeptides, and cytokines which activate corresponding receptors in proportions of nerve fibers. In addition, the expression of receptors, for example, bradykinin and neurokinin 1 receptors, is upregulated during inflammation. The development of hyperexcitability of spinal cord neurons is produced by various transmitter/receptor systems that constitute and modulate synaptic activation of the neurons. The key transmitter is glutamate that activates N‐methyl‐d‐aspartate (NMDA) and non‐NMDA receptors on spinal cord neurons. Blockade of these receptors prevents and reduces central sensitization. Excitatory neuropeptides (substance P and calcitonin gene‐related peptide) further central sensitization. Central sensitization also is facilitated by mediators that have complex actions (e.g., prostaglandin E2). Spinal PGE2 binds to receptors at presynaptic endings of primary afferent neurons (thus influencing synaptic release) and to receptors on postsynaptic spinal cord neurons. The administration of PGE2 to the spinal cord surface produces changes of responsiveness of spinal neurons similar to peripheral inflammation, and spinal indomethacin to the spinal cord attenuates development of hyperexcitability significantly.

Antinociceptive effects of tumor necrosis factor α neutralization in a rat model of antigen-induced arthritis: Evidence of a neuronal target

OBJECTIVE The reduction of pain in the course of antiinflammatory therapy can result from an attenuation of the inflammatory process and/or from the neutralization of endogenous mediators of inflammation that act directly on nociceptive neurons. The purpose of this study was to investigate whether analgesic effects of the neutralization of tumor necrosis factor alpha (TNFalpha) are due to an attenuation of inflammation or whether direct neuronal effects significantly contribute to pain relief in the course of therapy. METHODS Locomotor and pain-related behavior and histology were assessed in rats with chronic antigen-induced arthritis (AIA) in the knee joint, and the rats were treated with systemic saline, etanercept, or infliximab. The expression of TNF receptors (TNFRs) in dorsal root ganglia was measured using immunohistochemical analysis and polymerase chain reaction. Action potentials were recorded from afferent Adelta fibers and C fibers of the medial knee joint nerve, and etanercept and infliximab were injected intraarticularly into normal or inflamed knee joints (AIA or kaolin/carrageenan-induced inflammation). RESULTS In rats with AIA, both etanercept and infliximab significantly decreased inflammation-induced locomotor and pain-related behavior, while joint swelling was only weakly attenuated and histomorphology still revealed pronounced inflammation. A large proportion of dorsal root ganglion neurons showed TNFRI- and TNFRII-like immunoreactivity. Intraarticular injection of etanercept reduced the responses of joint afferents to mechanical stimulation of the inflamed joint starting 30 minutes after injection, but had no effect on responses to mechanical stimulation of the uninflamed joint. CONCLUSION Overall, these data show the pronounced antinociceptive effects of TNFalpha neutralization, thus suggesting that reduction of the effects of TNFalpha on pain fibers themselves significantly contributes to pain relief.

The cytokine TNFα increases the proportion of DRG neurones expressing the TRPV1 receptor via the TNFR1 receptor and ERK activation

TNFalpha is involved in the generation of hyperalgesia in pathological states such as neuropathy and inflammation. The pronociceptive action of TNFalpha may be mediated at least in part by activation of the TRPV1 receptor which transduces heat stimuli in primary nociceptive afferents and mediates thermal hyperalgesia. In the present study, we investigated in cultured dorsal root ganglion (DRG) neurones, the somata of primary afferent fibres, whether TNFalpha increases TRPV1 receptor expression. We found that long-term exposure of DRG neurones of both rat and mouse to TNFalpha significantly increased the proportion of DRG neurones expressing TRPV1 receptor-like immunoreactivity. This TNFalpha effect was abolished in mice DRG neurones when DRG cultures were obtained from tnfr1/2-/- and tnfr1-/-, but not from tnfr2-/- mice. Furthermore, we found that activation of ERK but not of p38 kinase or cyclooxygenases is critically involved in the TNFalpha-induced increase of TRPV1 receptor expression.

Experimental arthritis causes tumor necrosis factor-α-dependent infiltration of macrophages into rat dorsal root ganglia which correlates with pain-related behavior

ABSTRACT After peripheral nerve damage macrophages infiltrate the dorsal root ganglia (DRG) in which cell bodies of lesioned neurons are located. However, infiltration of macrophages into the DRGs was also reported in complete Freunds adjuvant (CFA)‐induced inflammation raising the question whether CFA inflammation induces nerve cell damage or whether peripheral inflammation may also trigger macrophage infiltration into DRGs. Related questions are, first, which signals trigger macrophage infiltration into DRGs and, second, is macrophage infiltration correlated with pain‐related behavior. Using the rat model of unilateral antigen‐induced arthritis (AIA) in the knee we found a massive infiltration of ED1+ macrophages into the ipsi‐ and contralateral lumbar DRGs but not into thoracic DRGs. At no time point of AIA DRG neurons showed expression of activating transcription factor‐3 (ATF3) indicating that macrophage infiltration is not explainable by nerve cell lesions in this model. During AIA, lumbar but not thoracic DRGs exhibited a bilateral de novo expression of vascular cell adhesion molecule‐1 (VCAM‐1) which is known to be involved in macrophage infiltration. Tumor necrosis factor‐&agr; (TNF‐&agr;) neutralization with etanercept or infliximab treatment after induction of AIA significantly reduced both macrophage infiltration and VCAM‐1 expression. It also decreased mechanical hyperalgesia at the inflamed joint although the joint inflammation itself was barely attenuated, and it reduced mechanical hyperalgesia at the non‐inflamed contralateral knee joint. Thus, bilateral segment‐specific infiltration of macrophages into DRGs is part of an unilateral inflammatory process in peripheral tissue and it may be involved in the generation of hyperalgesia in particular on the non‐inflamed side.

Calcitonin gene-related peptide enhances TTX-resistant sodium currents in cultured dorsal root ganglion neurons from adult rats

&NA; The neuropeptide calcitonin gene‐related peptide (CGRP) binds to a subpopulation of dorsal root ganglion (DRG) neurons, elevates intracellular calcium, and causes inward currents in about 30% of lumbar DRG neurons. Using whole‐cell patch clamp recordings, we found in the present study that application of CGRP to isolated and cultured DRG neurons from the adult rat enhances voltage‐gated TTX‐resistant (TTX‐R) Na+ inward currents in about 30% of small‐ to medium‐sized DRG neurons. During CGRP, peak densities of Na+ currents increased significantly. CGRP shifted the membrane conductance of the CGRP‐responsive cells towards hyperpolarization without changing the slope of the peak conductance curve. The effect of CGRP was blocked by coadministration of CGRP8–37, an antagonist at the CGRP receptor. The effect of CGRP was also blocked after bath application of PKA14–22, a membrane‐permeant blocker of protein kinase A, and PKC19–31, a PKC inhibitor, in the recording pipette. These data show pronounced facilitatory effects of CGRP on TTX‐R Na+ currents in DRG neurons which are mediated through CGRP receptors and intracellular pathways involving protein kinases A and C. Thus, in addition to prostaglandins, CGRP is another mediator that affects TTX‐R Na+ currents which are thought to occur mainly in nociceptive DRG neurons.

Interleukin-17 sensitizes joint nociceptors to mechanical stimuli and contributes to arthritic pain through neuronal interleukin-17 receptors in rodents

OBJECTIVE Interleukin-17 (IL-17) is considered a proinflammatory cytokine, but whether neuronal IL-17 receptors contribute to the generation of arthritic pain is unknown. This study was undertaken to explore whether IL-17A acts on neurons, whether it sensitizes joint nociceptors, and whether neutralization of IL-17 is antinociceptive. METHODS We recorded action potentials from rat joint nociceptors after intraarticular injection of IL-17A. We studied the expression of the IL-17A receptor in the rat dorsal root ganglia (DRG), explored the effect of IL-17A on signaling pathways in cultured rat DRG neurons, and using patch clamp recordings, monitored changes of excitability by IL-17A. We tested whether an antibody to IL-17 influences pain behaviors in mice with antigen-induced arthritis (AIA). RESULTS A single injection of IL-17A into the rat knee joint elicited a slowly developing and long-lasting sensitization of nociceptive C fibers of the joint to mechanical stimuli, which was not attenuated by neutralizing tumor necrosis factor α or IL-6. The IL-17A receptor was visualized in most rat DRG neurons, the cell bodies of primary sensory neurons. In isolated and cultured rat DRG neurons, IL-17A caused rapid phosphorylation of protein kinase B and ERK, and it rapidly enhanced excitability. In mice with unilateral AIA in the knee, an antibody against IL-17 improved the guarding score and reduced secondary mechanical hyperalgesia at the ipsilateral paw. CONCLUSION Our findings indicate that IL-17A has the potential to act as a pain mediator by targeting IL-17 receptors in nociceptive neurons, and these receptors are particularly involved in inflammation-evoked mechanical hyperalgesia.

Neuronal IL-17 receptor upregulates TRPV4 but not TRPV1 receptors in DRG neurons and mediates mechanical but not thermal hyperalgesia.

In addition to the proinflammatory cytokines tumor necrosis factor-α, interleukin-6 and interleukin-1ß, the cytokine interleukin-17 (IL-17) is considered an important mediator of autoimmune diseases such as rheumatoid arthritis. Because tumor necrosis factor-α and interleukin-1ß have the potential to influence the expression of transduction molecules such as transient receptor potential vanilloid 1 (TRPV1) in dorsal root ganglion (DRG) neurons and thus to contribute to pain we explored in the present study whether IL-17A activates DRG neurons and influences the expression of TRPV1. The IL-17A receptor was visualized in most neurons in dorsal root ganglion (DRG) sections as well as in cultured DRG neurons. Upon long-term exposure to IL-17A, isolated and cultured rat DRG neurons showed a significant upregulation of extracellular-regulated kinase (ERK) and nuclear factor κB (NFκB). Long-term exposure of neurons to IL-17A did not upregulate the expression of TRPV1. However, we found a pronounced upregulation of transient receptor potential vanilloid 4 (TRPV4) which is considered a candidate transduction molecule for mechanical hyperalgesia. Upon the injection of zymosan into the paw, IL-17A-deficient mice showed less mechanical hyperalgesia than wild type mice but thermal hyperalgesia was not attenuated in IL-17A-deficient mice. These data show, therefore, a particular role of IL-17 in mechanical hyperalgesia, and they suggest that this effect is linked to an activation and upregulation of TRPV4.

Acute and long-term effects of IL-6 on cultured dorsal root ganglion neurones from adult rat.

IL‐6 contributes to pain and hyperalgesia in inflamed tissue. We have investigated short‐ and long‐term effects of IL‐6 on dorsal root ganglion (DRG) neurones. Glycoprotein 130‐like immunoreactivity (the signal transduction receptor subunit) was found in almost all neurones in DRG sections and in cultured DRG neurones from adult rat. In calcium‐imaging studies bath application of IL‐6 caused an increase of intracellular calcium in about one‐third of the DRG neurones suggesting functional IL‐6 receptors in a proportion of neurones. Long‐term but not short‐term exposure of DRG neurones to IL‐6 in vitro significantly enhanced the proportion of DRG neurones expressing neurokinin 1 receptor‐like immunoreactivity from 10% to up to 40%. This up‐regulation was dependent on the activation of mitogen‐activated protein kinase kinase (MEK) in the neurones, suggesting that the mitogen‐activated protein kinase (MAPK) pathway is important for this effects of IL‐6. Calcium‐imaging studies demonstrated that previous exposure of DRG neurones to IL‐6 enhanced the proportion of neurones that exhibit a substance P‐induced rise in intracellular calcium. These data show that IL‐6 has short‐ and long‐term effects on a proportion of DRG neurones. These effects are likely to contribute to pro‐nociceptive effects of IL‐6.

The role of interleukin‐1β in arthritic pain: Main involvement in thermal, but not mechanical, hyperalgesia in rat antigen‐induced arthritis

OBJECTIVE Interleukin-1β (IL-1β) is considered a pronociceptive cytokine, but its role in the generation of arthritic pain is unknown. The aim of this study was to investigate the role of IL-1β in arthritic pain and to explore the antinociceptive potential of the IL-1 receptor type I (IL-1RI) antagonist anakinra. METHODS Antigen-induced arthritis (AIA) was induced in rats. Expression of IL-1RI in the dorsal root ganglia (DRGs) was determined, and the effects of anakinra on inflammation, pain-related behavior, and receptor expression were assessed. In cultured DRG neurons, the effect of IL-1β on the expression of the transient receptor potential vanilloid 1 (TRPV-1) ion channel was examined. Recordings of action potentials from joint nociceptors were made after intraarticular injection of IL-1β into the rat knee joints. RESULTS AIA generated pronounced and persistent mechanical and thermal hyperalgesia, and IL-1RI expression in the lumbar DRGs was significantly up-regulated. Treatment with anakinra did not significantly reduce the severity of arthritis or mechanical hyperalgesia, but did result in a pronounced reduction in thermal hyperalgesia. In cultured DRG neurons, IL-1β up-regulated the expression of TRPV-1, a major transduction molecule involved in thermal hyperalgesia. During AIA, anakinra treatment down-regulated the expression of TRPV-1, consistent with the pronounced reduction in thermal hyperalgesia. IL-1β increased the mechanosensitivity of C-fibers of the joint, but reduced the mechanosensitivity of Aδ-fibers, thus having opposite effects on these mechanonociceptive nerve fibers. CONCLUSION In the context of arthritic knee pain, IL-1β and IL-1 receptors appear to be involved in thermal, rather than mechanical, hyperalgesia. Therefore, neutralization of IL-1β may be mainly antinociceptive in disease states characterized by thermal hyperalgesia, but not in disease states mainly characterized by mechanical hyperalgesia.

Spinal interleukin-6 is an amplifier of arthritic pain in the rat

OBJECTIVE Significant joint pain is usually widespread beyond the affected joint, which results from the sensitization of nociceptive neurons in the central nervous system (central sensitization). This study was undertaken to explore whether the proinflammatory cytokine interleukin-6 (IL-6) in the joint induces central sensitization, whether joint inflammation causes the release of IL-6 from the spinal cord, and whether spinal IL-6 contributes to central sensitization. METHODS In anesthetized rats, electrophysiologic recordings of spinal cord neurons with sensory input from the knee joint were made. Neuronal responses to mechanical stimulation of the rat knee and leg were monitored. IL-6 and soluble IL-6 receptor (sIL-6R) were applied to the knee joint or the spinal cord. Spinal release of IL-6 was measured by enzyme-linked immunosorbent assay. Soluble gp130, which neutralizes IL-6/sIL-6R, was spinally applied during the development of joint inflammation or during established inflammation. RESULTS A single injection of IL-6/sIL-6R into the rat knee joint as well as application of IL-6/sIL-6R to the rat spinal cord significantly increased the responses of spinal neurons to mechanical stimulation of the knee and ankle joint, i.e., induced central sensitization. Application of soluble gp130 to the rat spinal cord attenuated this effect of IL-6. The development of knee inflammation in the rat caused spinal release of IL-6. Spinal application of soluble gp130 attenuated the development of inflammation-evoked central sensitization but did not reverse it. CONCLUSION Our findings indicate that the generation of joint pain in the rat involves not only IL-6 in the joint but also IL-6 released from the spinal cord. Spinal IL-6 contributes to central sensitization and thus promotes the widespread hyperalgesia observed in the course of joint disease.