Jon D Levine

Publications of Jon D Levine

• Generation of a pain memory in the primary afferent nociceptor triggered by PKCε activation of CPEB.

  Authors: Oliver Bogen, Nicole Alessandri-Haber, Carissa Chu, Robert W Gear, Jon D Levine

  The Journal of neuroscience : the official journal of the Society for Neuroscience. 02/2012; 32(6):2018-26.

  Isolectin B(4)-positive [IB(4)(+)] primary afferent nociceptors challenged with an inflammatory or neuropathic insult develop a PKCε-dependent long-lasting hyperalgesic response to a subsequentIsolectin B(4)-positive [IB(4)(+)] 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 E(2) (PGE(2)), 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 PGE(2) do not show the enhanced and prolonged hyperalgesic response by which primed IB(4)(+)-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 IB(4)(+)-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.

• IB4-saporin attenuates acute and eliminates chronic muscle pain in the rat.

  Authors: Pedro Alvarez, Robert W Gear, Paul G Green, Jon D Levine

  Experimental neurology. 12/2011; 233(2):859-65.

  The function of populations of nociceptors in muscle pain syndromes remain poorly understood. We compared the contribution of two major classes, isolectin B4-positive (IB4(+)) and IB4-negativeThe function of populations of nociceptors in muscle pain syndromes remain poorly understood. We compared the contribution of two major classes, isolectin B4-positive (IB4(+)) and IB4-negative (IB4(-)) nociceptors, in acute and chronic inflammatory and ergonomic muscle pain. Baseline mechanical nociceptive threshold was assessed in the gastrocnemius muscle of rats treated with IB4-saporin, which selectively destroys IB4(+) nociceptors. Rats were then submitted to models of acute inflammatory (intramuscular carrageenan)- or ergonomic intervention (eccentric exercise or vibration)-induced muscle pain, and each of the three models also evaluated for the transition from acute to chronic pain, manifest as prolongation of prostaglandin E2 (PGE(2))-induced hyperalgesia, after recovery from the hyperalgesia induced by acute inflammation or ergonomic interventions. IB4-saporin treatment did not affect baseline mechanical nociceptive threshold. However, compared to controls, IB4-saporin treated rats exhibited shorter duration mechanical hyperalgesia in all three models and attenuated peak hyperalgesia in the ergonomic pain models. And, IB4-saporin treatment completely prevented prolongation of PGE(2)-induced mechanical hyperalgesia. Thus, IB4(+) and IB4(-) neurons contribute to acute muscle hyperalgesia induced by diverse insults. However, only IB4+ nociceptors participate in the long term consequence of acute hyperalgesia.

• Sexual dimorphism in endothelin-1 induced mechanical hyperalgesia in the rat.

  Authors: Elizabeth K Joseph, Jon D Levine

  Experimental neurology. 12/2011; 233(1):505-12.

  While the onset of mechanical hyperalgesia induced by endothelin-1 was delayed in female rats, compared to males, the duration was much longer. Given that the repeated test stimulus used to assessWhile the onset of mechanical hyperalgesia induced by endothelin-1 was delayed in female rats, compared to males, the duration was much longer. Given that the repeated test stimulus used to assess nociceptive threshold enhances hyperalgesia, a phenomenon we have referred to as stimulus-induced enhancement of hyperalgesia, we also evaluated for sexual dimorphism in the impact of repeated application of the mechanical test stimulus on endothelin-1 hyperalgesia. In male and female rats, endothelin-1 induced hyperalgesia is already maximal at 30 min. At this time stimulus-induced enhancement of hyperalgesia, which is observed only in male rats, persisted for 3-4h. In contrast, in females, it develops only after a very long (15 day) delay, and is still present, without attenuation, at 45 days. Ovariectomy eliminated these differences between male and female rats. These findings suggest marked, ovarian-dependent sexual dimorphism in endothelin-1 induced mechanical hyperalgesia and its enhancement by repeated mechanical stimulation.

• Early-life stress produces muscle hyperalgesia and nociceptor sensitization in the adult rat.

  Authors: Paul G Green, Xiaojie Chen, Pedro Alvarez, Luiz F Ferrari, Jon D Levine

  Pain. 08/2011; 152(11):2549-56.

  Chronic pain in adults has been associated with early-life stress. To examine the pronociceptive effect of early-life stress, we evaluated cutaneous and muscle nociception and activity in muscleChronic pain in adults has been associated with early-life stress. To examine the pronociceptive effect of early-life stress, we evaluated cutaneous and muscle nociception and activity in muscle nociceptors in an animal model of neonatal stress, limited bedding, in the rat. In this neonatal limited bedding (NLB) model, litters are exposed to limited bedding between postnatal days 2 and 9, and controls to standard bedding. In adult NLB-treated rats, mechanical nociceptive threshold in skeletal muscle was significantly lower (~22%) than in controls. Furthermore, administration of prostaglandin E(2) in skin as well as muscle produced markedly prolonged hyperalgesia, an effect prevented by spinal intrathecal injection of oligodeoxynucleotide antisense to protein kinase Cε (PKCε), a second messenger in nociceptors that has been implicated in the induction and maintenance of chronic pain. In electrophysiological studies, mechanical threshold of muscle nociceptors was reduced by ~31% and conduction velocity significantly increased (~28%). These findings indicate that neonatal stress induces a persistent hyperalgesia and nociceptor sensitization manifest in the adult and that the second messenger PKCε may be a target against which therapies might be directed to treat a chronic pain syndrome that is associated with early-life traumatic stress.

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

  Authors: Luiz F Ferrari, Adrienne Chum, Oliver Bogen, David B Reichling, Jon D Levine

  The Journal of neuroscience : the official journal of the Society for Neuroscience. 08/2011; 31(31):11404-10.

  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 Drp1While 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 rat's 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.

• Muscle pain in models of chemotherapy-induced and alcohol-induced peripheral neuropathy.

  Authors: Pedro Alvarez, Luiz F Ferrari, Jon D Levine

  Annals of neurology. 07/2011; 70(1):101-9.

  While inflammatory pain is well described in skeletal muscle, neuropathic muscle pain remains to be clarified. We used 3 well-established rodent models of peripheral neuropathy to evaluate for muscleWhile inflammatory pain is well described in skeletal muscle, neuropathic muscle pain remains to be clarified. We used 3 well-established rodent models of peripheral neuropathy to evaluate for muscle pain. In rats exposed to either of 2 neurotoxic cancer chemotherapies, paclitaxel or oxaliplatin, or to alcohol consumption, we assessed the evolution of mechanical hyperalgesia in skeletal muscle and skin, in the same animal. To explore the involvement of protein kinase C epsilon (PKCε), a second messenger implicated in some forms of neuropathic pain, antisense oligodeoxynucleotides (AS-ODNs) or mismatch ODNs (MM-ODNs) for PKCε were administered intrathecally. Rats submitted to models of chemotherapy-induced and alcohol-induced neuropathy developed persistent muscle hyperalgesia, which evolved in parallel in muscle and skin. The administration of PKCε AS, which has been shown to mediate cutaneous hyperalgesia in paclitaxel and ethanol models of neuropathic pain, also inhibited muscle hyperalgesia induced by these agents. Stopping AS-ODN was associated with the reappearance of hyperalgesia at both sites. The AS-ODN to PKCε treatment was devoid of effect in both muscle and skin in the oxaliplatin neuropathy model. Our results support the suggestion that neuropathic muscle pain may be a greater clinical problem than generally appreciated.

• Estrogen destabilizes microtubules through an ion-conductivity-independent TRPV1 pathway.

  Authors: Chandan Goswami, Julia Kuhn, Olayinka A Dina, Gregorio Fernández-Ballester, Jon D Levine, Antonio Ferrer-Montiel, Tim Hucho

  Journal of neurochemistry. 06/2011; 117(6):995-1008.

  Recently, we described estrogen and agonists of the G-protein coupled estrogen receptor GPR30 to induce protein kinase C (PKC)ε-dependent pain sensitization. PKCε phosphorylates the ion channelRecently, we described estrogen and agonists of the G-protein coupled estrogen receptor GPR30 to induce protein kinase C (PKC)ε-dependent pain sensitization. PKCε phosphorylates the ion channel transient receptor potential, vanilloid subclass I (TRPV1) close to a novel microtubule-TRPV1 binding site. We now modeled the binding of tubulin to the TRPV1 C-terminus. The model suggests PKCε phosphorylation of TRPV1-S800 to abolish the tubulin-TRPV1 interaction. Indeed, in vitro PKCε phosphorylation of TRPV1 hindered tubulin-binding to TRPV1. In vivo, treatment of sensory neurons and F-11 cells with estrogen and the GPR30 agonist, G-1, resulted in microtubule destabilization and retraction of microtubules from filopodial structures. We found estrogen and G-1 to regulate the stability of the microtubular network via PKC phosphorylation of the PKCε-phosphorylation site TRPV1-S800. Microtubule disassembly was not, however, dependent on TRPV1 ion conductivity. TRPV1 knock-down in rats inverted the effect of the microtubule-modulating drugs, Taxol and Nocodazole, on estrogen-induced and PKCε-dependent mechanical pain sensitization. Thus, we suggest the C-terminus of TRPV1 to be a signaling intermediate downstream of estrogen and PKCε, regulating microtubule-stability and microtubule-dependent pain sensitization.

• Mitochondrial dependence of nerve growth factor-induced mechanical hyperalgesia.

  Authors: Carissa Chu, Emma Levine, Robert W Gear, Oliver Bogen, Jon D Levine

  Pain. 05/2011; 152(8):1832-7.

  Mitochondria are present at high concentration at the site of sensory transduction in the peripheral terminals of nociceptors. Because nerve growth factor (NGF), which induces nociceptorMitochondria are present at high concentration at the site of sensory transduction in the peripheral terminals of nociceptors. Because nerve growth factor (NGF), which induces nociceptor sensitization by acting on the high-affinity tropomyosin receptor kinase A (TrkA) receptor, also produces local recruitment of mitochondria in DRG neurons, we evaluated the role of mitochondria in NGF-induced mechanical hyperalgesia. Inhibition of 3 major mitochondrial functions-oxidation of nutrients, adenosine triphosphate (ATP) production, and generation of reactive oxygen species--markedly attenuated NGF-induced mechanical hyperalgesia in the rat. Disruption of microtubules, which are required for the trafficking and subcellular localization of mitochondria, also attenuated NGF-induced hyperalgesia. Our results suggest a contribution of mitochondrial localization and function to NGF-dependent pain syndromes.

• Further validation of a model of fibromyalgia syndrome in the rat.

  Authors: Paul G Green, Pedro Alvarez, Robert W Gear, Dennis Mendoza, Jon D Levine

  The journal of pain : official journal of the American Pain Society. 04/2011; 12(7):811-8.

  We have recently developed an animal model of fibromyalgia syndrome in the rat. In this model, rats exposed to unpredictable sound stress develop a delayed onset enhancement and prolongation ofWe have recently developed an animal model of fibromyalgia syndrome in the rat. In this model, rats exposed to unpredictable sound stress develop a delayed onset enhancement and prolongation of cytokine-induced mechanical hyperalgesia in muscle and skin. In this study, we tested the hypothesis that our model also manifests symptoms of common comorbid diagnoses: irritable bowel syndrome, temporomandibular disorder, and anxiety. Both visceral sensitivity and cytokine hyperalgesia in masseter muscle were present in the stressed rats. Furthermore, in an established model of irritable bowel syndrome-water avoidance-we observed significant muscle hyperalgesia. Finally, using the elevated plus maze to assess for anxiety level, we observed a significantly higher anxiety level in sound stress-exposed rats. Thus, unpredictable sound stress produces a condition in the rat with several features-delayed onset visceral and temporomandibular hyperalgesia and increased anxiety, as well as cutaneous and muscle hyperalgesia-commonly found in patients with fibromyalgia syndrome. PERSPECTIVE: A stress model-unpredictable sound-in the rat exhibits several features (cutaneous, musculoskeletal, and visceral hyperalgesia, as well as anxiety) that are found in patients with fibromyalgia syndrome. Thus, this model may be used to test hypotheses about the underlying mechanisms and response to therapy in patients with fibromyalgia.

• Enhanced cytokine-induced mechanical hyperalgesia in skeletal muscle produced by a novel mechanism in rats exposed to unpredictable sound stress.

  Authors: Olayinka A Dina, Jon D Levine, Paul G Green

  European journal of pain (London, England). 03/2011; 15(8):796-800.

  Stress exacerbates both experimental and clinical pain, most well-characterized in irritable bowel and fibromyalgia syndromes. Since it has been hypothesized that cytokines play an etiopathogenicStress exacerbates both experimental and clinical pain, most well-characterized in irritable bowel and fibromyalgia syndromes. Since it has been hypothesized that cytokines play an etiopathogenic role in fibromyalgia and other chronic widespread pain conditions, we investigated the relationship between stress and cytokines in a model of stress-induced chronic somatic pain. A series of experiments were performed to evaluate the impact of stress on the hyperalgesia-induced by endotoxin (lipopolysaccharide, LPS) and the role of two pro-inflammatory cytokines, interleukin-6 (IL-6) and tumor necrosis α (TNFα). Fourteen days after exposure to a 4-day protocol of unpredictable sound stress, the ability of systemic LPS (100 μg/kg, i.p) to elicit cytokine-mediated mechanical hyperalgesia was measured in gastrocnemius muscle. LPS-induced hyperalgesia was significantly greater in stressed rats, but when rats were treated intrathecally with antisense oligodeoxynucleotide (ODN), to decrease either the gp130 subunit of the IL-6 receptor or the TNFα receptor, in nociceptors, skeletal muscle hyperalgesia in sound stressed, but not control, rats was prevented. These data suggest that chronic stress alters signaling in the primary afferent nociceptor for the hyperalgesia induced by endogenously produced pro-inflammatory cytokines.

• Nucleus accumbens facilitates nociception.

  Authors: Robert W Gear, Jon D Levine

  Experimental neurology. 03/2011; 229(2):502-6.

  We have previously demonstrated an opioid link in nucleus accumbens (NAc) that mediates antinociception produced by a novel ascending pain modulation pathway. For example, noxious stimulation inducesWe have previously demonstrated an opioid link in nucleus accumbens (NAc) that mediates antinociception produced by a novel ascending pain modulation pathway. For example, noxious stimulation induces heterosegmental antinociception that is mediated by both mu- and delta-opioid receptors in NAc. However, spinal intrathecal administration of the mu-receptor agonist [D-Ala(2), N-Me-Phe(4),Gly(5)-ol]-enkephalin (DAMGO) also induces heterosegmental antinociception. The aim of the present study in the rat was to identify the intra-NAc opioid receptors that mediate the antinociceptive effects of spinally administered DAMGO and also to determine the effect of NAc efferent activity on nociception. Intra-NAc administration of either the mu-opioid receptor antagonist Cys(2),Tyr(3), Orn(5),Pen(7)amide (CTOP) or the delta-opioid receptor antagonist naltrindole blocked the antinociceptive effect of spinally administered DAMGO on the jaw-opening reflex (JOR). Injection of quaternary lidocaine (QX-314) attenuated the JOR, suggesting that the output of NAc is pronociceptive. In support of this, intra-NAc injection of the excitatory amino acid agonist kainate enhanced the JOR. Thus, it is possible to modulate activity in NAc to bidirectionally attenuate or enhance nociception, suggesting a potential role for NAc in setting nociceptive sensitivity.

• Pain and death: neurodegenerative disease mechanisms in the nociceptor.

  Authors: David B Reichling, Jon D Levine

  Annals of neurology. 01/2011; 69(1):13-21.

  Chronic peripheral neuropathic pain is the result of abnormal activity in sensory nerves. It is well recognized that this sensory nerve dysfunction can be caused by traumatic, toxic, or metabolicChronic peripheral neuropathic pain is the result of abnormal activity in sensory nerves. It is well recognized that this sensory nerve dysfunction can be caused by traumatic, toxic, or metabolic insult to the nerve. In addition, there is growing recognition that neuropathic pain is a frequent manifestation of neurodegenerative diseases. In this regard, important clues to the cellular mechanisms of neuropathic pain may be found by close examination of neurodegenerative diseases (including Parkinson's disease) in which neuropathic pain is often an underappreciated but important clinical manifestation. This approach identifies specific mitochondrial and cytoskeletal mechanisms, previously implicated in the pathophysiology of neurodegenerative diseases in the central nervous system, that might contribute to neuropathic dysfunction in peripheral sensory nerve fibers. Investigations in preclinical models of common peripheral neuropathic pain conditions have supported the idea that a subset of these cellular mechanisms of neurodegeneration can produce painful hyperactivity in primary afferent nociceptors. Importantly, this emerging concept of neurodegenerative disease mechanisms in the primary afferent nociceptor identifies novel molecular targets for the treatment neuropathic pain.

• Neuropathic pain-like alterations in muscle nociceptor function associated with vibration-induced muscle pain.

  Authors: Xiaojie Chen, Paul G Green, Jon D Levine

  Pain. 11/2010; 151(2):460-6.

  We recently developed a rodent model of the painful muscle disorders induced by occupational exposure to vibration. In the present study we used this model to evaluate the function of sensory neuronsWe recently developed a rodent model of the painful muscle disorders induced by occupational exposure to vibration. In the present study we used this model to evaluate the function of sensory neurons innervating the vibration-exposed gastrocnemius muscle. Activity of 74 vibration-exposed and 40 control nociceptors, with mechanical receptive fields in the gastrocnemius muscle, were recorded. In vibration-exposed rats ∼15% of nociceptors demonstrated an intense and long-lasting barrage of action potentials in response to sustained suprathreshold mechanical stimulation (average of 2635 action potentials with frequency of ∼44Hz during a 1min suprathreshold stimulus) much greater than that has been reported to be produced even by potent inflammatory mediators. While these high-firing nociceptors had lower mechanical thresholds than the remaining nociceptors, exposure to vibration had no effect on conduction velocity and did not induce spontaneous activity. Hyperactivity was not observed in any of 19 neurons from vibration-exposed rats pretreated with intrathecal antisense for the IL-6 receptor subunit gp130. Since vibration can injure peripheral nerves and IL-6 has been implicated in painful peripheral neuropathies, we suggest that the dramatic change in sensory neuron function and development of muscles pain, induced by exposure to vibration, reflects a neuropathic muscle pain syndrome.

• Attenuation of activity in an endogenous analgesia circuit by ongoing pain in the rat.

  Authors: Luiz F Ferrari, Robert W Gear, Jon D Levine

  The Journal of neuroscience : the official journal of the Society for Neuroscience. 10/2010; 30(41):13699-706.

  Analgesic efficacy varies depending on the pain syndrome being treated. One reason for this may be a differential effect of individual pain syndromes on the function of the endogenous pain controlAnalgesic efficacy varies depending on the pain syndrome being treated. One reason for this may be a differential effect of individual pain syndromes on the function of the endogenous pain control circuits at which these drugs act to produce analgesia. To test this hypothesis, we examined the effects of diverse (i.e., ongoing inflammatory, neuropathic, or chronic widespread) pain syndromes on analgesia induced by activation of an opioid-mediated, noxious stimulus-induced endogenous pain control circuit. This circuit was activated by subdermal capsaicin injection at a site remote from the site of nociceptive testing. Analgesia was not affected by carrageenan-induced inflammatory pain or the early phase of oxaliplatin neuropathy (a complication of cancer chemotherapy). However, the duration of analgesia was markedly shorter in the late phase of oxaliplatin neuropathy and in alcoholic neuropathy. A model of fibromyalgia syndrome produced by chronic unpredictable stress and proinflammatory cytokines also shortened analgesia duration, but so did the same stress alone. Therefore, since chronic pain can activate neuroendocrine stress axes, we tested whether they are involved in the attenuation of analgesic duration induced by these pain syndromes. Rats in which the sympathoadrenal axis was ablated by adrenal medullectomy showed normal duration pain-induced analgesia in groups with either late-phase oxaliplatin neuropathy, alcoholic neuropathy, or exposure to sound stress. These results support the suggestion that pain syndromes can modulate activity in endogenous pain control circuits and that this effect is sympathoadrenal dependent.

• Eccentric exercise induces chronic alterations in musculoskeletal nociception in the rat.

  Authors: Pedro Alvarez, Jon D Levine, Paul G Green

  The European journal of neuroscience. 09/2010; 32(5):819-25.

  Eccentric muscle exercise is a common cause of acute and chronic (lasting days to weeks) musculoskeletal pain. To evaluate the mechanisms involved, we have employed a model in the rat, in whichEccentric muscle exercise is a common cause of acute and chronic (lasting days to weeks) musculoskeletal pain. To evaluate the mechanisms involved, we have employed a model in the rat, in which eccentric hind limb exercise produces both acute mechanical hyperalgesia as well as long-term changes characterized by enhanced hyperalgesia to subsequent exposure to an inflammatory mediator. Eccentric exercise of the hind limb produced mechanical hyperalgesia, measured in the gastrocnemius muscle, which returned to baseline at 120 h post-exercise. When nociceptive thresholds had returned to baseline, intramuscular injection of prostaglandin E(2) (PGE(2) ) induced hyperalgesia that was unattenuated 240 h later, much longer than PGE(2) -induced hyperalgesia in unexercised rats (4 h). This marked prolongation of PGE(2) hyperalgesia induced by eccentric exercise was prevented by the spinal intrathecal injection of oligodeoxynucleotide antisense to protein kinase Cε, a second messenger in nociceptors implicated in the induction of chronic pain. Exercise-induced hyperalgesia and prolongation of PGE(2) hyperalgesia were inhibited by the spinal intrathecal administration of antisense for the interleukin-6 but not the tumor necrosis factor α type 1 receptor. These findings provide further insight into the mechanism underlying exercise-induced chronic muscle pain, and suggest novel approaches for the prevention and treatment of exercise- or work-related chronic musculoskeletal pain syndromes.

• Alcohol consumption enhances antiretroviral painful peripheral neuropathy by mitochondrial mechanisms.

  Authors: Luiz F Ferrari, Jon D Levine

  The European journal of neuroscience. 09/2010; 32(5):811-8.

  A major dose-limiting side effect of human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) chemotherapies, such as the nucleoside reverse transcriptase inhibitors (NRTIs), is aA major dose-limiting side effect of human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) chemotherapies, such as the nucleoside reverse transcriptase inhibitors (NRTIs), is a small-fiber painful peripheral neuropathy, mediated by its mitochondrial toxicity. Co-morbid conditions may also contribute to this dose-limiting effect of HIV/AIDS treatment. Alcohol abuse, which alone also produces painful neuropathy, is one of the most important co-morbid risk factors for peripheral neuropathy in patients with HIV/AIDS. Despite the prevalence of this problem and its serious impact on the quality of life and continued therapy in HIV/AIDS patients, the mechanisms by which alcohol abuse exacerbates highly active antiretroviral therapy (HAART)-induced neuropathic pain has not been demonstrated. In this study, performed in rats, we investigated the cellular mechanism by which consumed alcohol impacts antiretroviral-induced neuropathic pain. NRTI 2',3'-dideoxycytidine (ddC; 50 mg/kg) neuropathy was mitochondrial-dependent and PKCε-independent, and alcohol-induced painful neuropathy was PKCε-dependent and mitochondrial-independent. At low doses, ddC (5 mg/kg) and alcohol (6.5% ethanol diet for 1 week), which alone do not affect nociception, together produce profound mechanical hyperalgesia. This hyperalgesia is mitochondrial-dependent but PKCε-independent. These experiments, which provide the first model for studying the impact of co-morbidity in painful neuropathy, support the clinical impression that alcohol consumption enhances HIV/AIDS therapy neuropathy, and provide evidence for a role of mitochondrial mechanisms underlying this interaction.

• Multiple PKCε-dependent mechanisms mediating mechanical hyperalgesia.

  Authors: Elizabeth K Joseph, Jon D Levine

  Pain. 04/2010; 150(1):17-21.

  We have recently implicated mitochondrial mechanisms in models of neuropathic and inflammatory pain, in some of which a role of protein kinase Cepsilon (PKCepsilon) has also been implicated. SinceWe have recently implicated mitochondrial mechanisms in models of neuropathic and inflammatory pain, in some of which a role of protein kinase Cepsilon (PKCepsilon) has also been implicated. Since mitochondria contain several proteins that are targets of PKCepsilon, we evaluated the role of mitochondrial mechanisms in mechanical hyperalgesia induced by proinflammatory cytokines that induce PKCepsilon-dependent nociceptor sensitization, and by a direct activator of PKCepsilon (psiepsilonRACK), in the rat. Prostaglandin E(2) (PGE(2))-induced hyperalgesia is short lived in naïve rats, while it is prolonged in psiepsilonRACK pre-treated rats, a phenomenon referred to as priming. Inhibitors of two closely related mitochondrial functions, electron transport (complexes I-V) and oxidative stress (reactive oxygen species), attenuated mechanical hyperalgesia induced by intradermal injection of psiepsilonRACK. In marked contrast, in a PKCepsilon-dependent form of mechanical hyperalgesia induced by prostaglandin E(2) (PGE(2)), inhibitors of mitochondrial function failed to attenuate hyperalgesia. These studies support the suggestion that at least two downstream signaling pathways can mediate the hyperalgesia induced by activating PKCepsilon.

• Shared mechanisms for opioid tolerance and a transition to chronic pain.

  Authors: Elizabeth K Joseph, David B Reichling, Jon D Levine

  The Journal of neuroscience : the official journal of the Society for Neuroscience. 03/2010; 30(13):4660-6.

  Clinical pain conditions may remain responsive to opiate analgesics for extended periods, but such persistent acute pain can undergo a transition to an opiate-resistant chronic pain state thatClinical pain conditions may remain responsive to opiate analgesics for extended periods, but such persistent acute pain can undergo a transition to an opiate-resistant chronic pain state that becomes a much more serious clinical problem. To test the hypothesis that cellular mechanisms of chronic pain in the primary afferent also contribute to the development of opiate resistance, we used a recently developed model of the transition of from acute to chronic pain, hyperalgesic priming. Repeated intradermal administration of the potent and highly selective mu-opioid agonist, [d-Ala(2),N-MePhe(4),gly-ol]-enkephalin (DAMGO), to produce tolerance for its inhibition of prostaglandin E(2) hyperalgesia, simultaneously produced hyperalgesic priming. Conversely, injection of an inflammogen, carrageenan, used to produce priming produced DAMGO tolerance. Both effects were prevented by inhibition of protein kinase Cepsilon (PKCepsilon). Carrageenan also induced opioid dependence, manifest as mu-opioid receptor antagonist (d-Phe-Cys-Tyr-d-Trp-Orn-Thr-Pen-Thr-NH(2))-induced hyperalgesia that, like priming, was PKCepsilon and G(i) dependent. These findings suggest that the transition from acute to chronic pain, and development of mu-opioid receptor tolerance and dependence may be linked by common cellular mechanisms in the primary afferent.

• Quantitative automated microscopy (QuAM) elucidates growth factor specific signalling in pain sensitization.

  Authors: Christine Andres, Sonja Meyer, Olayinka A Dina, Jon D Levine, Tim Hucho

  Molecular pain. 01/2010; 6:98.

  Dorsal root ganglia (DRG)-neurons are commonly characterized immunocytochemically. Cells are mostly grouped by the experimenter's eye as "marker-positive" and "marker-negative" according to theirDorsal root ganglia (DRG)-neurons are commonly characterized immunocytochemically. Cells are mostly grouped by the experimenter's eye as "marker-positive" and "marker-negative" according to their immunofluorescence intensity. Classification criteria remain largely undefined. Overcoming this shortfall, we established a quantitative automated microscopy (QuAM) for a defined and multiparametric analysis of adherent heterogeneous primary neurons on a single cell base.The growth factors NGF, GDNF and EGF activate the MAP-kinase Erk1/2 via receptor tyrosine kinase signalling. NGF and GDNF are established factors in regeneration and sensitization of nociceptive neurons. If also the tissue regenerating growth factor, EGF, influences nociceptors is so far unknown. We asked, if EGF can act on nociceptors, and if QuAM can elucidate differences between NGF, GDNF and EGF induced Erk1/2 activation kinetics. Finally, we evaluated, if the investigation of one signalling component allows prediction of the behavioral response to a reagent not tested on nociceptors such as EGF. We established a software-based neuron identification, described quantitatively DRG-neuron heterogeneity and correlated measured sample sizes and corresponding assay sensitivity. Analysing more than 70,000 individual neurons we defined neuronal subgroups based on differential Erk1/2 activation status in sensory neurons. Baseline activity levels varied strongly already in untreated neurons. NGF and GDNF subgroup responsiveness correlated with their subgroup specificity on IB4(+)- and IB4(-)-neurons, respectively. We confirmed expression of EGF-receptors in all sensory neurons. EGF treatment induced STAT3 translocation into the nucleus. Nevertheless, we could not detect any EGF induced Erk1/2 phosphorylation. Accordingly, intradermal injection of EGF resulted in a fundamentally different outcome than NGF/GDNF. EGF did not induce mechanical hyperalgesia, but blocked PGE2-induced sensitization. QuAM is a suitable if not necessary tool to analyze activation of endogenous signalling in heterogeneous cultures. NGF, GDNF and EGF stimulation of DRG-neurons shows differential Erk1/2 activation responses and a corresponding differential behavioral phenotype. Thus, in addition to expression-markers also signalling-activity can be taken for functional subgroup differentiation and as predictor of behavioral outcome. The anti-nociceptive function of EGF is an intriguing result in the context of tissue damage but also for understanding pain resulting from EGF-receptor block during cancer therapy.

• Neurotoxic catecholamine metabolite in nociceptors contributes to painful peripheral neuropathy.

  Authors: Olayinka A Dina, Sachia G Khasar, Nicole Alessandri-Haber, Oliver Bogen, Xiaojie Chen, Paul G Green, David B Reichling, Robert O Messing, Jon D Levine

  The European journal of neuroscience. 12/2009; 30(11):2235.