Oliver Bogen

Oxaliplatin Acts on IB4-Positive Nociceptors to Induce an Oxidative Stress-Dependent Acute Painful Peripheral Neuropathy

UNLABELLED The toxicity profile of oxaliplatin, a platinum derivative currently used in the treatment of colorectal cancer, differs from those of the other platinum compounds, cisplatin and carboplatin. Oxaliplatin treatment induces an acute neurotoxicity characterized by a rapid onset of cold-induced distal dysesthesia and a chronic sensory peripheral neuropathy. A single intravenous dose of oxaliplatin produced a dose-dependent mechanical hyperalgesia and heat and cold allodynia; repeated administration intensified symptoms. A single intradermal dose of oxaliplatin produced a dose-dependent mechanical hyperalgesia. A single dose intravenous oxaliplatin also lowered thresholds and increased responses of C-fiber nociceptors to mechanical stimulation, confirming a peripheral site of action. Whereas peripheral administration of inhibitors of second messengers implicated in models of other painful peripheral neuropathies (PKA, PKC, NO, Ca(2+), and caspase) had no effect; both systemic and local administration of antioxidants (acetyl-L-carnitine, alpha-lipoic acid or vitamin C), all markedly inhibited oxaliplatin-induced hyperalgesia. Intrathecal administration of the neurotoxin for IB4-positive nociceptors, IB4-saporin, markedly attenuated IB4 staining in the dorsal horn of the spinal cord and completely prevented oxaliplatin-induced hyperalgesia. We suggest that oxaliplatin acts on IB4 (+)-nociceptors to induce oxidative stress-dependent acute peripheral sensory neuropathy. PERSPECTIVE Many drugs used to treat cancer produce pain as their dose-limiting side effect. We used a model of this pain syndrome induced by oxaliplatin to demonstrate that pain is produced by action on a subset of nociceptors, the IB4-positive DRG neurons. This information could help define cellular targets against which protective therapies could be developed.

Stress Induces a Switch of Intracellular Signaling in Sensory Neurons in a Model of Generalized Pain

Stress dramatically exacerbates pain in diseases such as fibromyalgia and rheumatoid arthritis, but the underlying mechanisms are unknown. We tested the hypothesis that stress causes generalized hyperalgesia by enhancing pronociceptive effects of immune mediators. Rats exposed to nonhabituating sound stress exhibited no change in mechanical nociceptive threshold, but showed a marked increase in hyperalgesia evoked by local injections of prostaglandin E2 or epinephrine. This enhancement, which developed more than a week after exposure to stress, required concerted action of glucocorticoids and catecholamines at receptors located in the periphery on sensory afferents. The altered response to pronociceptive mediators involved a switch in coupling of their receptors from predominantly stimulatory to inhibitory G-proteins (Gs to Gi), and for prostaglandin E2, emergence of novel dependence on protein kinase Cε. Thus, an important mechanism in generalized pain syndromes may be stress-induced coactivation of the hypothalamo-pituitary-adrenal and sympathoadrenal axes, causing a long-lasting alteration in intracellular signaling pathways, enabling normally innocuous levels of immune mediators to produce chronic hyperalgesia.

Proinflammatory cytokines mediating burn-injury pain

&NA; Thermal burns induce pain at the site of injury, mechanical hyperalgesia, associated with a complex time‐dependent inflammatory response. To determine the contribution of inflammatory mediators to burn injury‐induced mechanical hyperalgesia, we measured dynamic changes in the levels of three potent hyperalgesic cytokines, interleukin IL‐1β, IL‐6, and tumor necrosis factor‐α (TNFα), in skin of the rat, following a partial‐thickness burn injury. Only IL‐6 demonstrated a sustained increase ipsilateral but not contralateral to the burn, correlating with the prolonged ipsilateral mechanical hyperalgesia. Spinal intrathecal injection of oligodeoxynucleotides antisense for gp130, a receptor subunit shared by members of the IL‐6 family of cytokines, attenuated both burn‐ and intradermal IL‐6‐induced hyperalgesia, as did intradermal injection of anti‐IL‐6 function blocking antibodies. These studies suggest that IL‐6 is an important mediator of burn‐injury pain.

Generation of a Pain Memory in the Primary Afferent Nociceptor Triggered by PKCε Activation of CPEB

Isolectin B4-positive [IB4(+)] primary afferent nociceptors challenged with an inflammatory or neuropathic insult develop a PKCε-dependent long-lasting hyperalgesic response to a subsequent challenge by the proinflammatory cytokine prostaglandin E2 (PGE2), a phenomenon known as hyperalgesic priming. Here we demonstrate that the neuroplasticity underlying nociceptor priming requires 72 h to be established; rats that have been challenged with the inflammatory mediator TNFα 24 or 48 h ahead of PGE2 do not show the enhanced and prolonged hyperalgesic response by which primed IB4(+)-nociceptors are being characterized. Moreover, as the underlying plasticity can be interrupted by the peripheral administration of the protein translation inhibitor anisomycin it is reflected by changes in the peripheral protein expression pattern. Finally, the induction of priming by the selective PKCε agonist, psi ε receptor for activated c kinase (ψεRACK) can be prevented, but not reversed by intrathecal injections of antisense oligodeoxynucleotides for the cytoplasmic polyadenylation element binding protein (CPEB) mRNA, a master regulator of protein translation that coimmunoprecipitated with PKCε and is almost exclusively expressed by IB4(+)-nociceptors. Our results suggest that CPEB is downstream of PKCε in the cellular signaling cascade responsible for the induction of priming, raising the intriguing possiblity that prion-like misfolding could be a responsible mechanism for the chronification of pain.

GDNF hyperalgesia is mediated by PLCγ, MAPK/ERK, PI3K, CDK5 and Src family kinase signaling and dependent on the IB4‐binding protein versican

The function of the isolectin B4 (IB4+)‐binding and GDNF‐dependent Ret (Ret+)‐expressing non‐peptidergic subpopulation of nociceptors remain poorly understood. We demonstrate that acute administration of GDNF sensitizes nociceptors and produces mechanical hyperalgesia in the rat. Intrathecal IB4–saporin, a selective toxin for IB4+/Ret+‐nociceptors, attenuates GDNF but not NGF hyperalgesia. Conversely, intrathecal antisense to Trk A attenuated NGF but not GDNF hyperalgesia. Intrathecal administration of antisense oligodeoxynucleotides targeting mRNA for versican, the molecule that renders the Ret‐expressing nociceptors IB4‐positive (+), also attenuated GDNF but not NGF hyperalgesia, as did ADAMTS‐4, a matrix metalloprotease known to degrade versican. Finally, inhibitors for all five signaling pathways known to be activated by GDNF at GFRα1/Ret: PLCγ, CDK5, PI3K, MAPK/ERK and Src family kinases, attenuated GDNF hyperalgesia. Our results demonstrate a role of the non‐peptidergic nociceptors in pain produced by the neurotrophin GDNF and suggest that the IB4‐binding protein versican functions in the expression of this phenotype.

Nociceptor Subpopulations Involved in Hyperalgesic Priming

We have previously developed a model in the rat for the transition from acute to chronic pain, hyperalgesic priming, in which a long-lasting neuroplastic change in signaling pathways mediates a prolongation of proinflammatory cytokine-induced nociceptor sensitization and mechanical hyperalgesia, induced at the site of a previous inflammatory insult. Induction of priming is mediated by activation of protein kinase C(epsilon) (PKC(epsilon)) in the peripheral terminal of the primary afferent nociceptor. Given that hyperalgesic mediator-induced PKC(epsilon) translocation occurs in isolectin B4 (IB4)(+)-nonpeptidergic but not in receptor tyrosine kinase (TrkA)(+)-peptidergic nociceptors, we tested the hypothesis that hyperalgesic priming was restricted to the IB4(+) subpopulation of nociceptors. After recovery from nerve growth factor (NGF)- and GDNF-induced hyperalgesia, a proinflammatory cytokine, prostaglandin E(2) (PGE(2)) induced, PKC(epsilon)-dependent, markedly prolonged hyperalgesia, two features that define the development of the primed state. Thus, hyperalgesic priming occurs in both the IB4(+)-nonpeptidergic and TrkA(+)-peptidergic subpopulations of nociceptive afferents. Of note, however, while attenuation of PKC(epsilon) prevented NGF-induced priming, the hyperalgesia induced by NGF is PKC(epsilon) independent. We propose that separate intracellular pools of PKC(epsilon), in the peripheral terminals of nociceptors, mediate nociceptor sensitization and the induction of hyperalgesic priming.

PLC-β3 signals upstream of PKCε in acute and chronic inflammatory hyperalgesia

Abstract While protein kinase C&egr; has been shown to contribute to acute and chronic mechanical hyperalgesia, its upstream signaling pathway has received little attention. Since phospholipase C can signal to PKC&egr; and has been implicated in nociceptor sensitization, we tested if it is upstream of PKC&egr; in mechanisms underlying primary mechanical hyperalgesia. In the rat, the PKC&egr;‐dependent mechanical hyperalgesia and hyperalgesic priming (i.e., a form of chronic latent enhanced hyperalgesia) induced by carrageenan were attenuated by a non‐selective PLC inhibitor U‐73122. A lipid mediator of PLC signaling, l‐α‐lysophosphatidylcholine produced dose‐dependent mechanical hyperalgesia and hyperalgesic priming, which was attenuated by EAVSLKPT, a selective PKC&egr; inhibitor. However, U‐73122 did not attenuate hyperalgesia induced by ψ&egr;RACK, a selective PKC&egr; activator. Antisense to PLC‐β3 isoform, which was found in small‐diameter dorsal root ganglion neurons, also attenuated carrageenan‐induced acute and chronic‐latent hyperalgesia. These studies support the suggestion that PLC‐β3 is an important upstream signaling molecule for PKC&egr;‐mediated acute and chronic inflammatory pain.

Role of Nociceptor αCaMKII in Transition from Acute to Chronic Pain (Hyperalgesic Priming) in Male and Female Rats

We have previously shown that activation of protein kinase Cε (PKCε) in male rats induces a chronic, long-lasting change in nociceptors such that a subsequent exposure to proinflammatory mediators produces markedly prolonged mechanical hyperalgesia. This neuroplastic change, hyperalgesic priming, is dependent on activation of cytoplasmic polyadenylation element-binding protein (CPEB), downstream of PKCε, and consequent translation of mRNAs in the peripheral terminal of the nociceptor. Since α calmodulin-dependent protein kinase II (αCaMKII), a molecule implicated in neuroplasticity, is a target of CPEB and can also affect CPEB function, we investigated its role in the transition from acute to chronic pain. Priming induced by direct activation of PKCε can be prevented by inhibition of αCaMKII. In addition, direct activation of αCaMKII induces priming, which was not prevented by pretreatment with PKCε antisense, suggesting that αCaMKII is downstream of PKCε in the induction of priming. Activation of ryanodine receptors (RyRs), which can lead to activation of αCaMKII, also induced priming, in a calcium- and αCaMKII-dependent manner. Similarly, inhibition of the RyR and a calcium buffer prevented induction of priming by PKCε. Unlike activation of PKCε, ryanodine and αCaMKII induced priming in female as well as male rats. Our results demonstrate a contribution of αCaMKII to induction of hyperalgesic priming, a phenomenon implicated in the transition from acute to chronic pain.

Role of Drp1, a key mitochondrial fission protein, in neuropathic pain.

While oxidative stress has been implicated in small-fiber painful peripheral neuropathies, antioxidants are only partially effective to treat patients. We have tested the hypothesis that Drp1 (dynamin-related protein 1), a GTPase that catalyzes the process of mitochondrial fission, which is a mechanism central for the effect and production of reactive oxygen species (ROS), plays a central role in these neuropathic pain syndromes. Intrathecal administration of oligodeoxynucleotide antisense against Drp1 produced a decrease in its expression in peripheral nerve and markedly attenuated neuropathic mechanical hyperalgesia caused by HIV/AIDS antiretroviral [ddC (2′,3′-dideoxycytidine)] and anticancer (oxaliplatin) chemotherapy in male Sprague Dawley rats. To confirm the role of Drp1 in these models of neuropathic pain, as well as to demonstrate its contribution at the site of sensory transduction, we injected a highly selective Drp1 inhibitor, mdivi-1, at the site of nociceptive testing on the dorsum of the rats hindpaw. mdivi-1 attenuated both forms of neuropathic pain. To evaluate the role of Drp1 in hyperalgesia induced by ROS, we demonstrated that intradermal hydrogen peroxide produced dose-dependent hyperalgesia that was inhibited by mdivi-1. Finally, mechanical hyperalgesia induced by diverse pronociceptive mediators involved in inflammatory and neuropathic pain—tumor necrosis factor α, glial-derived neurotrophic factor, and nitric oxide—was also inhibited by mdivi-1. These studies provide support for a substantial role of mitochondrial fission in preclinical models of inflammatory and neuropathic pain.

Peripheral Administration of Translation Inhibitors Reverses Increased Hyperalgesia in a Model of Chronic Pain in the Rat

UNLABELLED Chronic pain is extremely difficult to manage, in part due to lack of progress in reversing the underlying pathophysiology. Since translation of messenger ribonucleic acids (mRNAs) in the peripheral terminal of the nociceptor plays a role in the transition from acute to chronic pain, we tested the hypothesis that transient inhibition of translation in the peripheral terminal of the nociceptor could reverse hyperalgesic priming, a model of transition from acute to chronic pain. We report that injection of translation inhibitors rapamycin and cordycepin, which inhibit translation by different mechanisms, at the peripheral terminal of the primed nociceptor produces reversal of priming in the rat that outlasted the duration of action of these drugs to prevent the development of priming. These data support the suggestion that interruption of translation in the nociceptor can reverse a preclinical model of at least 1 form of chronic pain. PERSPECTIVE This study provides evidence that ongoing protein translation in the sensory neuron terminals is involved in pain chronification, and local treatment that transiently interrupts this translation may be a useful therapy to chronic pain.