1-O-Acetylgeopyxin A, a derivative of a fungal metabolite, blocks tetrodotoxin-sensitive voltage-gated sodium, calcium channels and neuronal excitability which correlates with inhibition of neuropathic pain.

Chronic pain can be the result of an underlying disease or condition, medical treatment, inflammation, or injury. The number of persons experiencing this type of pain is substantial, affecting upwards of 50 million adults in the United States. Pharmacotherapy of most of the severe chronic pain patients includes drugs such as gabapentinoids, re-uptake blockers and opioids. Unfortunately, gabapentinoids are not effective in up to two-thirds of this population and although opioids can be initially effective, their long-term use is associated with multiple side effects. Therefore, there is a great need to develop novel non-opioid alternative therapies to relieve chronic pain. For this purpose, we screened a small library of natural products and their derivatives in the search for pharmacological inhibitors of voltage-gated calcium and sodium channels, which are outstanding molecular targets due to their important roles in nociceptive pathways. We discovered that the acetylated derivative of the ent-kaurane diterpenoid, geopyxin A, 1-O-acetylgeopyxin A, blocks voltage-gated calcium and tetrodotoxin-sensitive voltage-gated sodium channels but not tetrodotoxin-resistant sodium channels in dorsal root ganglion (DRG) neurons. Consistent with inhibition of voltage-gated sodium and calcium channels, 1-O-acetylgeopyxin A reduced reduce action potential firing frequency and increased firing threshold (rheobase) in DRG neurons. Finally, we identified the potential of 1-O-acetylgeopyxin A to reverse mechanical allodynia in a preclinical rat model of HIV-induced sensory neuropathy. Dual targeting of both sodium and calcium channels may permit block of nociceptor excitability and of release of pro-nociceptive transmitters. Future studies will harness the core structure of geopyxins for the generation of antinociceptive drugs.

Antiallodynic Effects of Cannabinoid Receptor 2 (CB2R) Agonists on Retrovirus Infection-Induced Neuropathic Pain.

The most common neurological complication in patients receiving successful combination antiretroviral therapy (cART) is peripheral neuropathic pain. Data show that distal symmetric polyneuropathy (DSP) also develops along with murine acquired immunodeficiency syndrome (MAIDS) after infection with the LP-BM5 murine retrovirus mixture. Links between cannabinoid receptor 2 (CB2R) and peripheral neuropathy have been established in animal models using nerve transection, chemotherapy-induced pain, and various other stimuli. Diverse types of neuropathic pain respond differently to standard drug intervention, and little is currently known regarding the effects of modulation through CB2Rs. In this study, we evaluated whether treatment with the exogenous synthetic CB2R agonists JWH015, JWH133, Gp1a, and HU308 controls neuropathic pain and neuroinflammation in animals with chronic retroviral infection. Hind-paw mechanical hypersensitivity in CB2R agonist-treated versus untreated animals was assessed using the MouseMet electronic von Frey system. Multicolor flow cytometry was used to determine the effects of CB2R agonists on macrophage activation and T-lymphocyte infiltration into dorsal root ganglia (DRG) and lumbar spinal cord (LSC). Results demonstrated that, following weekly intraperitoneal injections starting at 5 wk p.i., JWH015, JWH133, and Gp1a, but not HU308 (5 mg/kg), significantly ameliorated allodynia when assessed 2 h after ligand injection. However, these same agonists (2x/wk) did not display antiallodynic effects when mechanical sensitivity was assessed 24 h after ligand injection. Infection-induced macrophage activation and T-cell infiltration into the DRG and LSC were observed at 12 wk p.i., but this neuroinflammation was not affected by treatment with any CB2R agonist. Activation of JAK/STAT3 has been shown to contribute to development of neuropathic pain in the LSC and pretreatment of primary murine microglia (2 h) with JWH015-, JWH133-, or Gp1a-blocked IFN-gamma-induced phosphorylation of STAT1 and STAT3. Taken together, these data show that CB2R agonists demonstrate acute, but not long-term, antiallodynic effects on retrovirus infection-induced neuropathic pain.

Sex Differences in a Rodent Model of HIV-1-Associated Neuropathic Pain.

Worldwide, women account for approximately 51% of human immunodeficiency virus-1 (HIV) seropositive individuals. The prevalence of neuropathic pain among individuals with HIV and a lack of preclinical data characterizing sex differences prompted us to address this knowledge gap. C57BL/6 male and female mice received multiple intrathecal injections of HIV-glycoprotein 120 (gp120), followed by determination of mechanical allodynia and thermal hypersensitivity for four weeks. The influence of ovarian hormones in the gp120 pain model was evaluated by comparison of ovariectomized (OVX) mice versus sham control. We found that gp120-induced neuropathic pain-like behaviors are sex-dependent. Female mice showed both increased mechanical allodynia and increased cold sensitivity relative to their male counterparts. The OVX mice showed reduced pain sensitivity compared to sham, suggesting a role of the ovarian hormones in sex differences in pain sensitivity to gp120. Gp120-induced neuropathic pain caused a shift in estrous cycle toward the estrus phase. However, there is a lack of clear correlation between the estrous cycle and the development of neuropathic pain-like behaviors during the four week recording period. This data provided the first evidence for sex differences in a rodent model of HIV-related neuropathic pain, along with a potential role of ovarian hormones.

Enhanced facilitation and diminished inhibition characterizes the pronociceptive endogenous pain modulatory balance of persons living with HIV and chronic pain.

Chronic pain in persons living with HIV (PLWH) may be related to alterations in endogenous pain modulatory processes (e.g., high facilitation and low inhibition of nociception) that promote exaggerated pain responses, known as hyperalgesia, and central nervous system (CNS) sensitization. This observational study examined differences in endogenous pain modulatory processes between 59 PLWH with chronic pain, 51 PLWH without chronic pain, and 50 controls without HIV or chronic pain. Quantitative sensory testing for temporal summation (TS) of mechanical and heat pain as well as conditioned pain modulation (CPM) were used to assess endogenous pain facilitatory and inhibitory processes, respectively. Associations among TS, CPM, and self-reported clinical pain severity were also examined in PLWH with chronic pain. Findings demonstrated significantly greater TS of mechanical and heat pain for PLWH with chronic pain compared to PLWH without chronic pain and controls. CPM effects were present in controls, but not in either PLWH with or without chronic pain. Among PLWH with chronic pain, greater TS of mechanical pain was significantly associated with greater average clinical pain severity. Results of this study suggest that enhanced facilitation and diminished inhibition characterizes the pronociceptive endogenous pain modulatory balance of persons living with HIV and chronic pain.

P2Y12 receptor upregulation in satellite glial cells is involved in neuropathic pain induced by HIV glycoprotein 120 and 2',3'-dideoxycytidine.

The direct neurotoxicity of HIV and neurotoxicity of combination antiretroviral therapy medications both contribute to the development of neuropathic pain. Activation of satellite glial cells (SGCs) in the dorsal root ganglia (DRG) plays a crucial role in mechanical and thermal hyperalgesia. The P2Y12 receptor expressed in SGCs of the DRG is involved in pain transmission. In this study, we explored the role of the P2Y12 receptor in neuropathic pain induced by HIV envelope glycoprotein 120 (gp120) combined with ddC (2',3'-dideoxycytidine). A rat model of gp120+ddC-induced neuropathic pain was used. Peripheral nerve exposure to HIV-gp120+ddC increased mechanical and thermal hyperalgesia in gp120+ddC-treated model rats. The gp120+ddC treatment increased expression of P2Y12 receptor mRNA and protein in DRG SGCs. In primary cultured DRG SGCs treated with gp120+ddC, the levels of [Ca2+]i activated by the P2Y12 receptor agonist 2-(Methylthio) adenosine 5'-diphosphate trisodium salt (2-MeSADP) were significantly increased. P2Y12 receptor shRNA treatment inhibited 2-MeSADP-induced [Ca2+]i in primary cultured DRG SGCs treated with gp120+ddC. Intrathecal treatment with a shRNA against P2Y12 receptor in DRG SGCs reduced the release of pro-inflammatory cytokines, decreased phosphorylation of p38 MAPK in the DRG of gp120+ddC-treated rats. Thus, downregulating the P2Y12 receptor relieved mechanical and thermal hyperalgesia in gp120+ddC-treated rats.

Variations in herpes zoster manifestation.

Herpes zoster (HZ) is a neurocutaneous disorder due to endogenous reactivation of the varicella-zoster virus (VZV). The typical clinical manifestation is an acute segmental eruption of herpetiform umbilicated vesicles associated with malaise, pain, dysaesthesia, allodynia and probably fever. This review focuses on other possible clinical manifestations of the disease to sensitize physicians not to overlook HZ since only an early antiviral treatment can reduce the risk of post-zosteric neuralgia.

Detectable Viral Load May Be Associated with Increased Pain Sensitivity in Persons Living with HIV: Preliminary Findings.

OBJECTIVE: Animal models have previously shown that HIV is associated with hyperalgesia, or heightened sensitivity to painful stimuli. Efforts to determine whether this finding translates to humans are presently lacking. Among persons living with HIV (PLWH), those with detectable viral loads may be at greatest risk for heightened pain sensitivity. It was hypothesized that PLWH with detectable viral loads would be more sensitive to painful stimuli compared with PLWH without detectable viral loads and healthy controls without HIV. DESIGN: A total of 47 PLWH and 50 community-dwelling, healthy adults without HIV (controls) were recruited. Participants completed a quantitative sensory testing protocol to assess threshold, tolerance, and temporal summation in response to painful mechanical and heat stimuli. Most recent viral load was collected from medical records, and viral load was considered detectable if the count was greater than 50 copies/mL of blood. Of the 47 PLWH, 11 (23.4%) had detectable viral loads, the median viral load count was 10,200 copies/mL. RESULTS: PLWH with detectable viral loads demonstrated significantly lower pain thresholds for mechanical stimuli (F2,89 = 3.15, P = 0.049), significantly lower heat pain tolerances (F2,89 = 3.38, P = 0.039), and significantly greater temporal summation of heat pain at 48 °C (F2,89 = 10.66, P < 0.001) and 50 °C (F2,89 = 3.82, P = 0.026), compared with PLWH without detectable viral loads and healthy controls. CONCLUSIONS: These preliminary results tentatively suggest that the detectable presence of the virus may sensitize PLWH to painful mechanical and heat stimuli.

Inhibition of Mitochondrial Fission Protein Reduced Mechanical Allodynia and Suppressed Spinal Mitochondrial Superoxide Induced by Perineural Human Immunodeficiency Virus gp120 in Rats.

BACKGROUND: Mitochondria play an important role in many cellular and physiologic functions. Mitochondria are dynamic organelles, and their fusion and fission regulate cellular signaling, development, and mitochondrial homeostasis. The most common complaint of human immunodeficiency virus (HIV)-sensory neuropathy is pain on the soles in patients with HIV, but the exact molecular mechanisms of HIV neuropathic pain are not clear. In the present study, we investigated the role of mitochondrial dynamin-related protein 1 (Drp1, a GTPase that mediates mitochondrial fission) in the perineural HIV coat glycoprotein gp120-induced neuropathic pain state. METHODS: Neuropathic pain was induced by the application of recombinant HIV-1 envelope protein gp120 into the sciatic nerve. Mechanical threshold was tested using von Frey filaments. The mechanical threshold response was assessed over time using the area under curves. Intrathecal administration of antisense oligodeoxynucleotide (ODN) against Drp1, mitochondrial division inhibitor-1 (mdivi-1), or phenyl-N-tert-butylnitrone (a reactive oxygen species scavenger) was given. The expression of spinal Drp1 was examined using western blots. The expression of mitochondrial superoxide in the spinal dorsal horn was examined using MitoSox imaging. RESULTS: Intrathecal administration of either antisense ODN against Drp1 or mdivi-1 decreased mechanical allodynia (a sensation of pain evoked by nonpainful stimuli) in the gp120 model. Intrathecal ODN or mdivi-1 did not change basic mechanical threshold in sham surgery rats. Intrathecal Drp1 antisense ODN decreased the spinal expression of increased Drp1 protein induced by peripheral gp120 application. Intrathecal phenyl-N-tert-butylnitrone reduced mechanical allodynia. Furthermore, both intrathecal Drp1 antisense ODN and mdivi-1 reversed the upregulation of mitochondrial superoxide in the spinal dorsal horn in the gp120 neuropathic pain state. CONCLUSIONS: These data suggest that mitochondrial division plays a substantial role in the HIV gp120-related neuropathic pain state through mitochondrial reactive oxygen species and provides evidence for a novel approach to treating chronic pain in patients with HIV.

Gp120 in the pathogenesis of human immunodeficiency virus-associated pain.

OBJECTIVE: Chronic pain is a common neurological comorbidity of human immunodeficiency virus (HIV)-1 infection, but the etiological cause remains elusive. The objective of this study was to identify the HIV-1 causal factor that critically contributes to the pathogenesis of HIV-associated pain. METHODS: We first compared the levels of HIV-1 proteins in postmortem tissues of the spinal cord dorsal horn (SDH) from HIV-1/acquired immunodeficiency syndrome patients who developed chronic pain (pain-positive HIV-1 patients) and HIV-1 patients who did not develop chronic pain (pain-negative HIV-1 patients). Then we used the HIV-1 protein that was specifically increased in the pain-positive patients to generate mouse models. Finally, we performed comparative analyses on the pathological changes in the models and the HIV-1 patients. RESULTS: We found that HIV-1 gp120 was significantly higher in pain-positive HIV-1 patients (vs pain-negative HIV-1 patients). This finding suggested that gp120 was a potential causal factor of the HIV-associated pain. To test this hypothesis, we used a mouse model generated by intrathecal injection of gp120 and compared the pathologies of the model and the pain-positive human HIV-1 patients. The results showed that the mouse model and pain-positive human HIV-1 patients developed extensive similarities in their pathological phenotypes, including pain behaviors, peripheral neuropathy, glial reactivation, synapse degeneration, and aberrant activation of pain-related signaling pathways in the SDH. INTERPRETATION: Our findings suggest that gp120 may critically contribute to the pathogenesis of HIV-associated pain.

Varicella zoster virus-induced pain and post-herpetic neuralgia in the human host and in rodent animal models.

Pain and post-herpetic neuralgia (PHN) are common and highly distressing complications of herpes zoster that remain a significant public health concern and in need of improved therapies. Zoster results from reactivation of the herpesvirus varicella zoster virus (VZV) from a neuronal latent state established at the primary infection (varicella). PHN occurs in some one fifth to one third of zoster cases with severity, incidence, and duration of pain increasing with rising patient age. While VZV reactivation and the ensuing ganglionic damage trigger the pain response, the mechanisms underlying protracted PHN are not understood, and the lack of an animal model of herpes zoster (reactivation) makes this issue more challenging. A recent preclinical rodent model has developed that opens up the potential to allow the exploration of the underlying mechanisms and treatments for VZV-induced pain. Rats inoculated with live cell-associated human VZV into the hind paw reliably demonstrate thermal hyperalgesia and mechanical allodynia for extended periods and then spontaneously recover. Dorsal root ganglia express a limited VZV gene subset, including the IE62 regulatory protein, and upregulate expression of markers suggesting a neuropathic pain state. The model has been used to investigate treatment modalities and aspects of pain signaling and is under investigation by the authors to delineate VZV genetics involved in the induction of pain. This article compares human zoster-associated pain and PHN to the pain indicators in the rat and poses important questions that, if answered, could be the basis for new treatments.

Effect of a single dose of pregabalin on herpes zoster pain.

BACKGROUND: The effect of pregabalin on acute herpes zoster pain has not been previously evaluated. METHODS: In a randomized, double-blind, placebo-controlled, two-session crossover study the effect of a single oral dose of pregabalin (150 mg) on pain and allodynia was evaluated in 8 subjects with herpes zoster. RESULTS: Over 6 hours of observation, pain decreased by a mean of 33% with pregabalin and 14% with placebo (p < 0.10). Effects on allodynia and SF-MPQ were not significant. CONCLUSIONS: Compared to an earlier study of gabapentin 900 mg for acute zoster pain and allodynia that followed a nearly identical protocol, pregabalin had a similar effect on pain and was well tolerated, with no difference from placebo on sleepiness. Common side effects of light-headedness, unsteady gait, and slowed thinking were almost identical to that observed in the earlier study of gabapentin. Subject recruitment proved difficult in part due to the widespread off-label use of gabapentin and pregabalin for acute zoster pain in our region of the USA. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT00352651.

A virus-like particle-based anti-nerve growth factor vaccine reduces inflammatory hyperalgesia: potential long-term therapy for chronic pain.

Chronic pain resulting from inflammatory and neuropathic disorders causes considerable economic and social burden. For a substantial proportion of patients, conventional drug treatments do not provide adequate pain relief. Consequently, novel approaches to pain management, involving alternative targets and new therapeutic modalities compatible with chronic use, are being sought. Nerve growth factor (NGF) is a major mediator of chronic pain. Clinical testing of NGF antagonists is ongoing, and clinical proof of concept has been established with a neutralizing mAb. Active immunization, with the goal of inducing therapeutically effective neutralizing autoreactive Abs, is recognized as a potential treatment option for chronic diseases. We have sought to determine if such a strategy could be applied to chronic pain by targeting NGF with a virus-like particle (VLP)-based vaccine. A vaccine comprising recombinant murine NGF conjugated to VLPs from the bacteriophage Qß (NGFQß) was produced. Immunization of mice with NGFQß induced anti-NGF-specific IgG Abs capable of neutralizing NGF. Titers could be sustained over 1 y by periodic immunization but declined in the absence of boosting. Vaccination with NGFQß substantially reduced hyperalgesia in collagen-induced arthritis or postinjection of zymosan A, two models of inflammatory pain. Long-term NGFQß immunization did not change sensory or sympathetic innervation patterns or induce cholinergic deficits in the forebrain, nor did it interfere with blood-brain barrier integrity. Thus, autovaccination targeting NGF using a VLP-based approach may represent a novel modality for the treatment of chronic pain.

Pharmacological differences between static and dynamic allodynia in mice with herpetic or postherpetic pain.

In the present study, we investigated whether dynamic and static allodynia would be developed in the affected dermatome in murine models of herpetic pain and postherpetic neuralgia and pharmacologically characterized the allodynia. Inoculation with herpes simplex virus type-1 on the femur induced skin lesions in the dermatome including the plantar region of the hind paw from day 5 to day 21 after inoculation. Dynamic allodynia became apparent in the hind paw from day 3 to at least day 42. Static allodynia was not obvious during the stage of herpetic pain and gradually increased after the lesion healing. Mexiletine hydrochloride (30 mg/kg, p.o.) and ketamine hydrochloride (50 mg/kg, i.p.) produced a moderate attenuation of static but not dynamic allodynia. Diclofenac sodium (50 mg/kg, i.p.) did not affect both static and dynamic allodynia. Gabapentin (30 mg/kg, p.o.) markedly inhibited both static and dynamic allodynia. Developmental and pharmacological differences between static and dynamic allodynia suggest that independent mechanisms are responsible for dynamic and static allodynia. This murine model may be useful for the study of the mechanisms of dynamic allodynia of herpetic pain or postherpetic neuralgia and the development of new analgesics effective against the dynamic allodynia.

Different roles of nitric oxide synthase-1 and -2 between herpetic and postherpetic allodynia in mice.

We investigated using the mice role of nitric oxide synthase (NOS) in the spinal dorsal horn in herpetic and postherpetic pain, especially allodynia, which was induced by transdermal inoculation of the hind paw with herpes simplex virus type-1 (HSV-1). The virus inoculation induced NOS2 expression in the lumbar dorsal horn of mice with herpetic allodynia, but not postherpetic allodynia. There were no substantial alternations in the expression level of NOS1 at the herpetic and postherpetic stages. Herpetic allodynia was significantly inhibited by i.p. administration of the selective NOS2 inhibitor S-methylisothiourea, but not the selective NOS1 inhibitor 7-nitroindazole. NOS2 expression was observed around HSV-1 antigen-immunoreactive cells. On the other hand, postherpetic allodynia was significantly inhibited by i.p. administration of 7-nitroindazole, but not S-methylisothiourea. The activity of reduced nicotinamide adenine dinucleotide phosphate diaphorase, an index of NOS1 activity, significantly increased in the laminae I and II of the lumbar dorsal horn of mice with postherpetic allodynia, but not mice without postherpetic allodynia. The expression level of NOS1 mRNA in the dorsal root ganglia was similar between mice with and without postherpetic allodynia. The results suggest that herpetic and postherpetic allodynia is mediated by nitric oxide in the dorsal horn and that NOS2 and NOS1 are responsible for herpetic and postherpetic allodynia, respectively. It may be worth testing the effects of NOS2 and NOS1 inhibitors on herpetic pain and postherpetic neuralgia in human subjects, respectively.

Recombinant herpes vector-mediated analgesia in a primate model of hyperalgesia.

Some chronic pain syndromes are characterized by episodes of intense burning and hyperalgesia in localized areas of skin. These sensations are thought to be mediated, at least in part, by the activity of damaged, unmyelinated C nociceptors. These phenomena were modeled by assaying responses of macaques to thermal and chemical stimuli that produced periodic activation and sensitization of C nociceptors. Upon validation of this method, a recombinant herpes simplex vector encoding human preproenkephalin was topically applied to the dorsal surface of the feet of the monkeys. Immunohistochemistry and radioimmunoassay revealed that enkephalin peptides were being produced in releasable pools in sensory neurons innervating the treated skin area. Behavioral responses evoked by periodic sensitization and activation of C nociceptors innervating the vector-treated skin area revealed a substantial and long-lasting (at least 20 weeks) antihyperalgesic and analgesic effect limited to the areas to which the virus was applied. This approach may be a viable means of treating localized cutaneous burning pain and hyperalgesia.

Allodynia in rats infected with varicella zoster virus--a small animal model for post-herpetic neuralgia.

The most common complication of herpes zoster is post-herpetic neuralgia (PHN), which has been defined as severe pain occurring 1 month after rash onset or persisting for greater than 3 months. PHN is classed as a neuropathic pain that is associated with mechanical allodynia where normally innocuous tactile stimuli are perceived as painful. The development of therapies to treat PHN has been hampered by the lack of animal models, which mimic the clinical situation. We have previously reported that varicella zoster virus (VZV) infection in the rat results in mechanical allodynia and thermal hyperalgesia. Here, we report that following VZV infection of the left footpad rats develop a chronic mechanical allodynia, which is present for longer than 60 days post-infection and which resolves by 100 days PI. The model is robust and reproducible with animals consistently developing allodynia by 3 days PI and continuing to present with symptoms for at least 30 days. The reproducible nature of the induction and course of the allodynia allows the use of this model to determine the effect of various compounds on, and to investigate the pathogenic mechanisms underlying the development of VZV-induced allodynia. Comparative studies using HSV-1 show that the induction of the chronic allodynia is VZV-specific and is not a result is of virus replication-induced tissue damage or accompanying inflammation. Therefore, we propose that the rat VZV infection model could prove useful in studying the mechanisms underlying post-herpetic neuralgia.

Pharmacological and immunohistochemical characterization of a mouse model of acute herpetic pain.

We have recently found that the infection with herpes simplex virus type-1 (HSV-1) of primary sensory neurons induces nociceptive hypersensitivity to noxious mechanical (hyperalgesia) and tactile stimulation (allodynia) in mice. In the present experiments, we determined the distribution of HSV-1 in the dorsal root ganglia and examined the effects of four analgesic agents on hyperalgesia and allodynia. HSV-1 was inoculated on the unilateral shin. HSV-antigen-positive cells were detected in the L4 and L5 dorsal root ganglia on days 5 and 7, but not day 3, post-inoculation. About 80% of the positive cells were small in size. Allodynia and hyperalgesia appeared on day 5 post-inoculation. Antinociceptive effects of analgesic agents were examined on day 6 post-inoculation. Morphine (1-5 mg/kg, subcutaneous) and gabapentin (10-100 mg/kg, peroral) dose-dependently inhibited both allodynia and hyperalgesia. Diclofenac (10-100 mg/kg, intraperitoneal) also produced antinociceptive effects, but there was a ceiling for the effect on hyperalgesia. Amitriptyline (3, 10 mg/kg, subcutaneous) did not affect allodynia and hyperalgesia. The results suggest that mechanical allodynia and hyperalgesia appeared when HSV-1 proliferated in the sensory neurons. This mouse model may be useful for studying the mechanisms of acute herpetic pain and anti-neuropathic pain agents.