Mechanical Pain Hypersensitivity Research Articles

Neonatal vincristine administration modulates intrinsic neuronal excitability in the rat dorsal root ganglion and spinal dorsal horn during adolescence.

Our recent work has shown that the early-life administration of vincristine (VNC), commonly used to treat pediatric cancers, evokes mechanical pain hypersensitivity in rats that emerges during adolescence and persists into adulthood. However, the underlying mechanisms remain unclear, as nothing is known about how neonatal VNC treatment influences peripheral and central nociceptive processing at the cellular level. Here, we used in vitro intracellular microelectrode and whole-cell patch-clamp recordings to evaluate the consequences of early-life VNC administration on the intrinsic membrane properties of adolescent dorsal root ganglion and spinal superficial dorsal horn neurons. The results demonstrate that VNC treatment increased the prevalence and rate of repetitive firing in both large- and medium-diameter sensory neurons, while reducing repetitive firing in small-diameter neurons, in comparison with vehicle-treated littermate controls. By contrast, passive membrane properties and peripheral conduction velocities were similar between experimental groups across all classes of primary afferents. Within the adolescent superficial dorsal horn, neonatal VNC exposure significantly enhanced the intrinsic membrane excitability of lamina I spinoparabrachial neurons, as evidenced by a decrease in rheobase and elevation of repetitive firing frequency compared with controls. Meanwhile, putative interneurons within lamina I exhibited a reduction in repetitive action potential discharge after early-life chemotherapy. Collectively, these findings suggest that neonatal VNC treatment evokes cell type-specific changes in intrinsic excitability at multiple levels of the ascending pain pathway. Overall, this work lays an essential foundation for the future exploration of the ionic mechanisms that drive chemotherapy-induced chronic pain in children and adolescents.

TRPA1 Antagonists for Pain Relief.

Here, we review the literature assessing the role of transient receptor potential ankyrin 1 (TRPA1), a calcium-permeable non-selective cation channel, in various types of pain conditions. In the nervous system, TRPA1 is expressed in a subpopulation of nociceptive primary sensory neurons, astroglia, oligodendrocytes and Schwann cells. In peripheral terminals of nociceptive primary sensory neurons, it is involved in the transduction of potentially harmful stimuli and in their central terminals it is involved in amplification of nociceptive transmission. TRPA1 is a final common pathway for a large number of chemically diverse pronociceptive agonists generated in various pathophysiological pain conditions. Thereby, pain therapy using TRPA1 antagonists can be expected to be a superior approach when compared with many other drugs targeting single nociceptive signaling pathways. In experimental animal studies, pharmacological or genetic blocking of TRPA1 has effectively attenuated mechanical and cold pain hypersensitivity in various experimental models of pathophysiological pain, with only minor side effects, if any. TRPA1 antagonists acting peripherally are likely to be optimal for attenuating primary hyperalgesia (such as inflammation-induced sensitization of peripheral nerve terminals), while centrally acting TRPA1 antagonists are expected to be optimal for attenuating pain conditions in which central amplification of transmission plays a role (such as secondary hyperalgesia and tactile allodynia caused by various types of peripheral injuries). In an experimental model of peripheral diabetic neuropathy, prolonged blocking of TRPA1 has delayed the loss of nociceptive nerve endings and their function, thereby promising to provide a disease-modifying treatment.

No evidence of widespread mechanical pressure hyperalgesia after experimentally induced central sensitization through skin nociceptors.

An increasing number of clinical studies involving a range of chronic pain conditions report widespread mechanical pressure pain hypersensitivity, which is commonly interpreted as resulting from central sensitization (CS). Secondary hyperalgesia (increased pinprick sensitivity surrounding the site of injury) is considered to be a manifestation of CS. However, it has not been rigorously tested whether CS induced by peripheral nociceptive input involves widespread mechanical pressure pain hypersensitivity. The aim of this study was to assess whether high-frequency electrical stimulation (HFS), which induces a robust secondary hyperalgesia, also induces a widespread decrease of pressure pain thresholds (PPTs). We measured PPTs bilaterally on the temples (temporalis muscles), on the legs (tibialis anterior muscles), and on the ventral forearm (flexor carpi radialis muscles) before, 20 minutes after, and 45 minutes after applying HFS on the ventral forearm of sixteen healthy young volunteers. To evaluate the presence of secondary hyperalgesia, mechanical pinprick sensitivity was assessed on the skin surrounding the site where HFS was applied and also on the contralateral arm. HFS induced a significant increase in mechanical pinprick sensitivity on the HFS-treated arm. However, HFS did not decrease PPTs neither in the area of increased pinprick sensitivity nor at more distant sites. This study provides no evidence for the hypothesis that CS, induced after intense activation of skin nociceptors, involves a widespread decrease of PPTs.

Intrathecal administration of adrenomedullin induces mechanical allodynia and neurochemical changes in spinal cord and DRG

This study investigated the effect of adrenomedullin (AM) on mechanical pain sensitivity and its possible mechanisms. Intrathecal injection of AM receptor agonist AM1-50 (20 μg) once per day briefly reduced mechanical pain threshold on days 1 and 2 but induced prolonged mechanical allodynia on day 3. However, AM1-50 did not change mechanical pain sensation when the AM receptor antagonist AM22-52 (20 μg) was intrathecally co-administered. Daily administration of AM1-50 (20 μg) for 3 days increased expression of phosphorylated extracellular signal-regulated protein kinase (pERK) and neuronal nitric oxide synthase (nNOS) in the spinal dorsal horn. The AM-induced increase in pERK and nNOS was inhibited by the co-administration of AM22-52. The chronic administration of AM1-50 also increased expression of microglial maker Iba1 and astrocytic marker GFAP (glial fibrillary acidic protein) in the spinal dorsal horn in an AM22-52-sensitive manner. Furthermore, the application of AM1-50 (10 nM, 3 h) to dorsal root ganglion (DRG) explant cultures induced an increase in the expression of transient receptor potential vanilloid 1 (TRPV1). The treatment with AM1-50 did not change TRPV1 expression in DRG in the presence of AM22-52 (2 μM). These results suggest that the increased AM bioactivity induced mechanical allodynia and may contribute to the mechanical pain hypersensitivity under pathological conditions. The mechanisms may involve the activation of ERK signaling pathway and spinal glia as well as the recruitment of nNOS and TRPV1 in the spinal dorsal horn or DRG. The present study indicates that inhibition of the activation AM receptor might provide a fruitful strategy to relieving chronic pain.

Validating Rat Model of Empathy for Pain: Effects of Pain Expressions in Social Partners.

Pain can be socially transferred between familiar rats due to empathic responses. To validate rat model of empathy for pain, effects of pain expressions in a cagemate demonstrator (CD) in pain on empathic pain responses in a naïve cagemate observer (CO) after 30 min priming dyadic social interactions (PDSI) were evaluated. The CD rats were prepared with four pain models: bee venom (BV), formalin, complete Freund's adjuvant (CFA), and spared nerve injury (SNI). Both BV and formalin tests are characterized by displayable and eye-identifiable spontaneous pain-related behaviors (SPRB) immediately after treatment, while CFA and SNI models are characterized by delayed occurrence of evoked pain hypersensitivity but with less eye-identifiable SPRB. After 30 min PDSI with a CD immediately after BV and formalin, respectively, the empathic mechanical pain hypersensitivity (EMPH) could be identified at both hind paws in CO rats. The BV—or formalin-induced EMPH in CO rats lasted for 4–5 h until full recovery. However, EMPH failed to develop in CO after socially interacting with a CD immediately after CFA, or 2 h after BV when SPRB completely disappeared. The CO's EMPH was partially relieved when socially interacting with an analgecized CD whose SPRB had been significantly suppressed. Moreover, repeated exposures to a CD in pain could enhance EMPH in CO. Finally, social transfer of pain hypersensitivity was also identified in CO who was being co-housed in pairs with a conspecific treated with CFA or SNI. The results suggest that development of EMPH in CO rats would be determined not only by extent of familiarity but also by visually identifiable pain expressions in the social partners during short period of PDSI. However, the visually unidentifiable pain can also be transferred to naïve cagemate when being co-housed in pairs with a distressed conspecific. In summary, the vicariously social contagion of pain between familiar rats is dependent upon not only expressions of pain in social partners but also the time that dyads spent in social communications. The rat model of empathy for pain is a highly stable, reproducible and valid model for studying the neural mechanisms of empathy in lower animals.

Macrophage angiotensin II type 2 receptor triggers neuropathic pain

Peripheral nerve damage initiates a complex series of structural and cellular processes that culminate in chronic neuropathic pain. The recent success of a type 2 angiotensin II (Ang II) receptor (AT2R) antagonist in a phase II clinical trial for the treatment of postherpetic neuralgia suggests angiotensin signaling is involved in neuropathic pain. However, transcriptome analysis indicates a lack of AT2R gene (Agtr2) expression in human and rodent sensory ganglia, raising questions regarding the tissue/cell target underlying the analgesic effect of AT2R antagonism. We show that selective antagonism of AT2R attenuates neuropathic but not inflammatory mechanical and cold pain hypersensitivity behaviors in mice. Agtr2-expressing macrophages (MΦs) constitute the predominant immune cells that infiltrate the site of nerve injury. Interestingly, neuropathic mechanical and cold pain hypersensitivity can be attenuated by chemogenetic depletion of peripheral MΦs and AT2R-null hematopoietic cell transplantation. Our study identifies AT2R on peripheral MΦs as a critical trigger for pain sensitization at the site of nerve injury, and therefore proposes a translatable peripheral mechanism underlying chronic neuropathic pain.

Green Pharma: A New Strategy for Drug Discovery in Academia by Targeting Glial Cells and ATP Receptors

Neuropathic pain associated with cancer, diabetic neuropathy, and postherpetic neuralgia is a type of intractable chronic pain characterized by mechanical allodynia and abnormal pain hypersensitivity evoked by innocuous stimuli. However, this disorder has no specific treatment. We previously showed that the purinergic receptor P2X4 (P2X4R), a subtype of ATP-gated nonselective cation channels, is highly upregulated in spinal microglia after peripheral nerve injury, and blocking the function of P2X4R reverses mechanical allodynia. In the present study, we screened a chemical library of 1979 clinically approved compounds (a gift from the Drug Discovery Initiative at the University of Tokyo) aimed at achieving "Eco-Pharma," which refers to seeking new effects of existing drugs. We demonstrated that duloxetine, a serotonin and noradrenaline reuptake inhibitor, has an inhibitory effect on rat and human P2X4R. In rat primary cultured microglial cells, duloxetine also inhibited P2X4R-mediated responses. Moreover, intrathecal administration of duloxetine in a model of neuropathic pain reversed nerve injury-induced mechanical allodynia. Based on those results, we suggest that the inhibition of P2X4R expressed in microglial cells may be involved in the antiallodynic effect of duloxetine in neuropathic pain. Furthermore, in this review, we discuss a new strategy for drug discovery called "Green Pharma" (a merger of "Eco-Pharma" and "Green chemistry" and referring to the development of eco-friendly pharmaceuticals).

A bivalent compound targeting CCR5 and the mu opioid receptor treats inflammatory arthritis pain in mice without inducing pharmacologic tolerance

BackgroundPain accompanies rheumatoid arthritis and other chronic inflammatory conditions and is difficult to manage. Although opioids provide potent analgesia, chronic opioid use can cause tolerance and addiction. Recent studies have demonstrated functional interactions between chemokine and opioid receptor signaling pathways. Reported heterodimerization of chemokine and opioid receptors led our group to develop bivalent compounds that bind both types of receptors, with the goal of targeting opioids to sites of inflammation. MCC22 is a novel bivalent compound containing a CCR5 antagonist and mu opioid receptor (MOR) agonist pharmacophores linked through a 22-atom spacer. We evaluated the efficacy of MCC22 in the K/B.g7 T-cell receptor transgenic mouse model of spontaneous inflammatory arthritis.MethodsMCC22 or morphine was administered intraperitoneally at varying doses to arthritic K/B.g7 mice or nonarthritic control mice. Mechanical pain hypersensitivity was measured each day before and after drug administration, using the electronic von Frey test. The potency of MCC22 relative to that of morphine was calculated. Functional readouts of pain included grip strength and nesting behavior. A separate dosing regimen was used to determine whether the drugs induced pharmacologic tolerance.ResultsMCC22 provided ~ 3000-fold more potent analgesia than morphine in this model. Daily treatment with MCC22 also led to a cumulative analgesic effect, reducing the daily baseline pain level. MCC22 produced no observable analgesic effect in nonarthritic control mice. Importantly, repeated administration of MCC22 did not induce pharmacologic tolerance, whereas a similar regimen of morphine did. Both grip strength and nesting behaviors improved among arthritic mice treated with MCC22. Ankle thickness and arthritis scores were not affected by MCC22. The analgesic effect of MCC22 was abolished in K/B.g7 mice genetically lacking CCR5, demonstrating the receptor specificity of the antagonist pharmacophore.ConclusionsMCC22 is a novel bivalent ligand that targets CCR5 and MOR. Our findings demonstrate that MCC22 provides highly potent analgesia and improved functional outcomes in a model of inflammatory arthritis, without inducing typical opioid tolerance. These findings suggest that MCC22 or similar compounds could be used to treat the pain associated with inflammatory arthritis and related conditions, while minimizing the risks typically associated with chronic opioid use.

Fast Green FCF Alleviates Pain Hypersensitivity and Down-Regulates the Levels of Spinal P2X4 Expression and Pro-inflammatory Cytokines in a Rodent Inflammatory Pain Model.

Fast Green FCF (FGF), a biocompatible dye, recently drew attention as a potential drug to treat amyloid-deposit diseases due to its effects against amyloid fibrillogenesis in vitro and a high degree of safety. However, its role in inflammatory pain is unknown. Our study aimed to investigate the effect of FGF in the inflammatory pain model induced by complete Freund’s adjuvant (CFA) and to identify the associated mechanisms. We found that systemic administration of FGF reversed mechanical and thermal pain hypersensitivity evoked by CFA in a dose-dependent manner. FGF treatment decreased purinergic spinal P2X4 expression in the spinal cord of CFA-inflamed mice. FGF also down-regulated spinal and peripheral pro-inflammatory cytokines [tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6)], but did not alter the spinal level of nerve growth factor (NGF) or brain-derived neurotrophic factor (BDNF). In conclusion, our results suggest the potential of FGF for controlling the progress of inflammatory pain.

Mice lacking Kcns1 in peripheral neurons show increased basal and neuropathic pain sensitivity.

Voltage-gated potassium (Kv) channels are increasingly recognised as key regulators of nociceptive excitability. Kcns1 is one of the first potassium channels to be associated with neuronal hyperexcitability and mechanical sensitivity in the rat, as well as pain intensity and risk of developing chronic pain in humans. Here, we show that in mice, Kcns1 is predominantly expressed in the cell body and axons of myelinated sensory neurons positive for neurofilament-200, including Aδ-fiber nociceptors and low-threshold Aβ mechanoreceptors. In the spinal cord, Kcns1 was detected in laminae III to V of the dorsal horn where most sensory A fibers terminate, as well as large motoneurons of the ventral horn. To investigate Kcns1 function specifically in the periphery, we generated transgenic mice in which the gene is deleted in all sensory neurons but retained in the central nervous system. Kcns1 ablation resulted in a modest increase in basal mechanical pain, with no change in thermal pain processing. After neuropathic injury, Kcns1 KO mice exhibited exaggerated mechanical pain responses and hypersensitivity to both noxious and innocuous cold, consistent with increased A-fiber activity. Interestingly, Kcns1 deletion also improved locomotor performance in the rotarod test, indicative of augmented proprioceptive signalling. Our results suggest that restoring Kcns1 function in the periphery may be of some use in ameliorating mechanical and cold pain in chronic states.

Control of mechanical pain hypersensitivity in mice through ligand-targeted photoablation of TrkB-positive sensory neurons

Mechanical allodynia is a major symptom of neuropathic pain whereby innocuous touch evokes severe pain. Here we identify a population of peripheral sensory neurons expressing TrkB that are both necessary and sufficient for producing pain from light touch after nerve injury in mice. Mice in which TrkB-Cre-expressing neurons are ablated are less sensitive to the lightest touch under basal conditions, and fail to develop mechanical allodynia in a model of neuropathic pain. Moreover, selective optogenetic activation of these neurons after nerve injury evokes marked nociceptive behavior. Using a phototherapeutic approach based upon BDNF, the ligand for TrkB, we perform molecule-guided laser ablation of these neurons and achieve long-term retraction of TrkB-positive neurons from the skin and pronounced reversal of mechanical allodynia across multiple types of neuropathic pain. Thus we identify the peripheral neurons which transmit pain from light touch and uncover a novel pharmacological strategy for its treatment.

Parathyroid Hormone-Related Peptide Elicits Peripheral TRPV1-dependent Mechanical Hypersensitivity.

Bone metastasis in breast, prostate and lung cancers often leads to chronic pain, which is poorly managed by existing analgesics. The neurobiological mechanisms that underlie chronic pain associated with bone-metastasized cancers are not well understood, but sensitization of peripheral nociceptors by tumor microenvironment factors has been demonstrated to be important. Parathyroid hormone-related peptide (PTHrP) is highly expressed in bone-metastasized breast and prostate cancers, and is critical to growth and proliferation of these tumors in the bone tumor microenvironment. Previous studies have suggested that PTHrP could sensitize nociceptive sensory neurons, resulting in peripheral pain hypersensitivity. In this study, we found that PTHrP induces both heat and mechanical hypersensitivity, that are dependent on the pain-transducing transient receptor potential channel family vanilloid, member-1 (TRPV1), but not the mechano-transducing TRPV4 and TRPA1 ion channels. Functional ratiometric Ca2+ imaging and voltage-clamp electrophysiological analysis of cultured mouse DRG neurons show significant potentiation of TRPV1, but not TRPA1 or TRPV4 channel activation by PTHrP. Interestingly, PTHrP exposure led to the slow and sustained activation of TRPV1, in the absence of any exogenous channel agonist, and is dependent on the expression of the type-1 parathyroid hormone receptor (PTH1), as well as on downstream phosphorylation of the channel by protein kinase C (PKC). Accordingly, local administration of specific small-molecule antagonists of TRPV1 to mouse hindpaws after the development of PTHrP-induced mechanical hypersensitivity led to its significant attenuation. Collectively, our findings suggest that PTHrP/PTH1-mediated flow activation of TRPV1 channel contributes at least in part to the development and maintenance of peripheral mechanical pain hypersensitivity, and could therefore constitute a mechanism for nociceptor sensitization in the context of metastatic bone cancer pain.

Blocking Mammalian Target of Rapamycin (mTOR) Alleviates Neuropathic Pain Induced by Chemotherapeutic Bortezomib

Bortezomib (BTZ) is largely used as a chemotherapeutic agent for the treatment of cancer. However, one of the significant limiting complications of BTZ is painful peripheral neuropathy during BTZ therapy. Drugs preventing and/or treating the painful symptoms induced by BTZ are lacking since the underlying mechanisms leading to neuropathic pain remain largely unclear. The purposes of this study were to examine 1) the effects of blocking mammalian target of rapamycin (mTOR) on mechanical pain and cold hypersensitivity evoked by BTZ and 2) the underlying mechanisms responsible for the role of mTOR in regulating BTZ-induced neuropathic pain. Behavioral test was performed to determine mechanical pain and cold sensitivity in a rat model. Western blot analysis and ELISA were used to examine expression of mTOR and phosphatidylinositide 3-kinase (p-PI3K) signals, and the levels of substance P and calcitonin gene-related peptide (CGRP). Systemic injection of BTZ significantly increased mechanical pain and cold sensitivity as compared with control animals (P< 0.05 vs. control rats). The expression of p-mTOR, mTOR-mediated phosphorylation of p70 ribosomal S6 protein kinase 1 (p-S6K1), 4E-binding protein 4 (p-4E-BP1) as well as p-PI3K was amplified in the dorsal horn of spinal cord of BTZ rats as compared with control rats. Blocking mTOR by intrathecal infusion of rapamycin attenuated mechanical pain and cold hypersensitivity. Blocking PI3K signal also attenuated activities of mTOR, which was accompanied with decreasing neuropathic pain. Inhibition of either mTOR or PI3K blunted enhancement of the spinal substance P and CGRP in BTZ rats. The data for the first time revealed specific signaling pathways leading to BTZ-induced peripheral neuropathic pain, including the activation of mTOR and PI3K. Inhibition of these signal pathways alleviates pain. Targeting one or more of these signaling molecules may present new opportunities for treatment and management of peripheral painful neuropathy observed during chemotherapeutic application of BTZ.

Anxiety- and activity-related effects of paracetamol on healthy and neuropathic rats.

Paracetamol has recently been suggested to affect emotion processing in addition to alleviating pain in humans. We investigated in adult male Hannover–Wistar rats whether acute intraperitoneally administrated paracetamol affects behavior in tests measuring anxiety, mood, motor activity, and memory. Unoperated rats received saline or a low (50 mg/kg) or high (300 mg/kg) dose of paracetamol, while rats with a spared nerve injury (SNI) model of neuropathy and sham‐operated rats received saline or the low dose of paracetamol. Rats were tested on open‐field (OFT), elevated plus‐maze (EPM), light‐dark box (LDB), novel‐object recognition (NOR), sucrose preference, rotarod, and monofilament tests. In unoperated rats, both the low and high dose of paracetamol reduced line crossings, and grooming time in the OFT, and novel preference in NOR. The high dose of paracetamol increased the time spent in the closed arm in EPM, reduced the number of rearings and leanings in OFT, the time spent in the light box in LDB, and sucrose preference. Paracetamol had no significant effect on the rotarod test measuring motor activity. The low dose of paracetamol suppressed mechanical pain hypersensitivity in SNI rats, without influencing pain behavior in sham‐operated rats. Saline‐ but not paracetamol‐treated SNI rats spent more time than sham‐operated rats in the closed arm in the EPM test. Together the results suggest that a high dose of paracetamol increases anxiety‐like and anhedonic behavior, and impairs recognition memory in unoperated controls, while in neuropathy, a low dose of paracetamol reduces nerve injury‐associated anxiety probably by reducing neuropathic pain.

Social interaction with a cagemate in pain increases allogrooming and induces pain hypersensitivity in the observer rats.

Empathy, which is a highly cognitive and emotional process, is the ability to share the emotional states of others. Empathy has also been observed in rodents. The empathic sharing of the distressful experience of a conspecific can even motivate altruistic behaviors, which are critical for survival. However, previous studies investigating empathy or prosocial behaviors in rodents mainly employed fearful or other stressful stimuli to elicit emotional changes; whether pain empathy can also motivate prosocial behaviors has yet to be investigated. By using the writhing test, the present study found that cagemate observer (CO) rats, compared with non-cagemate observer (NCO) rats, increased partner-directed grooming (allogrooming) toward conspecifics that had received an intraperitoneal injection of acetic acid during a dyadic social interaction. Following a dyadic social interaction with a demonstrator that received an intraperitoneal injection of acetic acid, the CO rats, compared with NCO rats, exhibited bilateral mechanical pain hypersensitivity and an enhanced acetic acid-induced writhing response. Our results here provided further evidence of pain empathy in rats, suggesting that empathy for pain may motivate prosocial behaviors in rats.

Attenuation of mechanical pain hypersensitivity by treatment with Peptide5, a connexin-43 mimetic peptide, involves inhibition of NLRP3 inflammasome in nerve-injured mice

Connexin43 (Cx43) hemichannels in spinal cord astrocytes are implicated in the maintenance of neuropathic pain following peripheral nerve injury. Peptide5 is a Cx43 mimetic peptide that blocks hemichannels. In this study, we investigated the effects of spinal delivery of Peptide5 on mechanical pain hypersensitivity in two mouse models of neuropathic pain, peripheral nerve injury and chemotherapy-induced peripheral neuropathy (CIPN). We demonstrated that 10days following a chronic constriction injury (CCI) of the sciatic nerve, Cx43 expression, co-localised predominantly with astrocytes, was increased in the ipsilateral L3–L5 lumbar spinal cord. An intrathecal injection of Peptide5 into nerve-injured mice, on day 10 when pain was well-established, caused significant improvement in mechanical pain hypersensitivity 8h after injection. Peptide5 treatment resulted in significantly reduced Cx43, and microglial and astrocyte activity in the dorsal horn of the spinal cord, as compared to control saline-treated CCI mice. Further in vitro investigations on primary astrocyte cultures showed that 1h pre-treatment with Peptide5 significantly reduced adenosine triphosphate (ATP) release in response to extracellular calcium depletion. Since ATP is a known activator of the NOD-like receptor protein 3 (NLRP3) inflammasome complex, a key mediator of neuroinflammation, we examined the effects of Peptide5 treatment on NLRP3 inflammasome expression. We found that NLRP3, its adaptor apoptosis-associated spec-like protein (ASC) and caspase-1 protein were increased in the ipsilateral spinal cord of CCI mice and reduced to naïve levels following Peptide5 treatment. In the models of oxaliplatin- and paclitaxel-induced peripheral neuropathy, treatment with Peptide5 had no effect on mechanical pain hypersensitivity. Interestingly, in these CIPN models, although spinal Cx43 expression was significantly increased at day 13 following chemotherapy, NLRP3 expression was not altered. These results suggest that the analgesic effect of Peptide5 is specifically achieved by reducing NLRP3 expression. Together, our findings demonstrate that blocking Cx43 hemichannels with Peptide5 after nerve injury attenuates mechanical pain hypersensitivity by specifically targeting the NLRP3 inflammasome in the spinal cord.

The Locus Coeruleus-Norepinephrine System Mediates Empathy for Pain through Selective Up-Regulation of P2X3 Receptor in Dorsal Root Ganglia in Rats.

Empathy for pain (vicariously felt pain), an ability to feel, recognize, understand and share the painful emotions of others, has been gradually accepted to be a common identity in both humans and rodents, however, the underlying neural and molecular mechanisms are largely unknown. Recently, we have developed a rat model of empathy for pain in which pain can be transferred from a cagemate demonstrator (CD) in pain to a naïve cagemate observer (CO) after 30 min dyadic priming social interaction. The naïve CO rats display both mechanical pain hypersensitivity (hyperalgesia) and enhanced spinal nociception. Chemical lesions of bilateral medial prefrontal cortex (mPFC) abolish the empathic pain response completely, suggesting existence of a top-down facilitation system in production of empathy for pain. However, the social transfer of pain was not observed in non-cagemate observer (NCO) after dyadic social interaction with a non-cagemate demonstrator (NCD) in pain. Here we showed that dyadic social interaction with a painful CD resulted in elevation of circulating norepinephrine (NE) and increased neuronal activity in the locus coeruleus (LC) in the CO rats. Meanwhile, CO rats also had over-expression of P2X3, but not TRPV1, in the dorsal root ganglia (DRG). Chemical lesion of the LC-NE neurons by systemic DSP-4 and pharmacological inhibition of central synaptic release of NE by clonidine completely abolished increase in circulating NE and P2X3 receptor expression, as well as the sympathetically-maintained development of empathic mechanical hyperalgesia. However, in the NCO rats, neither the LC-NE neuronal activity nor the P2X3 receptor expression was altered after dyadic social interaction with a painful NCD although the circulating corticosterone and NE were elevated. Finally, in the periphery, both P2X3 receptor and α1 adrenergic receptor were found to be involved in the development of empathic mechanical hyperalgesia. Taken together with our previous results, empathy for pain observed in the CO rats is likely to be mediated by activation of the top-down mPFC-LC/NE-sympathoadrenomedullary (SAM) system that further up-regulates P2X3 receptors in the periphery, however, social stress observed in the NCO rats is mediated by activation of both hypothalamic-pituitary-adrenocortical axis and SAM axis.

The transcription factor C/EBPβ in the dorsal root ganglion contributes to peripheral nerve trauma-induced nociceptive hypersensitivity.

Changes in gene transcription in the dorsal root ganglion (DRG) after nerve trauma contribute to the genesis of neuropathic pain. We report that peripheral nerve trauma caused by chronic constriction injury (CCI) increased the abundance of the transcription factor C/EBPβ (CCAAT/enhancer binding protein β) in the DRG. Blocking this increase mitigated the development and maintenance of CCI-induced mechanical, thermal, and cold pain hypersensitivities without affecting basal responses to acute pain and locomotor activity. Conversely, mimicking this increase produced hypersensitivity to mechanical, thermal, or cold pain. In the ipsilateral DRG, C/EBPβ promoted a decrease in the abundance of the voltage-gated potassium channel subunit Kv1.2 and μ opioid receptor (MOR) at the mRNA and protein levels, which would be predicted to increase excitability in the ipsilateral DRG neurons and reduce the efficacy of morphine analgesia. These effects required C/EPBβ-mediated transcriptional activation of Ehmt2 (euchromatic histone-lysine N-methyltransferase 2), which encodes G9a, an epigenetic silencer of the genes encoding Kv1.2 and MOR. Blocking the increase in C/EBPβ in the DRG improved morphine analgesia after CCI. These results suggest that C/EBPβ is an endogenous initiator of neuropathic pain and could be a potential target for the prevention and treatment of this disorder.

Early life vincristine exposure evokes mechanical pain hypersensitivity in the developing rat.

Vincristine (VNC) is commonly used to treat pediatric cancers, including the most prevalent childhood malignancy, acute lymphoblastic leukemia. Although clinical evidence suggests that VNC causes peripheral neuropathy in children, the degree to which pediatric chemotherapeutic regimens influence pain sensitivity throughout life remains unclear, in part because of the lack of an established animal model of chemotherapy-induced neuropathic pain during early life. Therefore, this study investigated the effects of VNC exposure between postnatal days (P) 11 and 21 on mechanical and thermal pain sensitivity in the developing rat. Low doses of VNC (15 or 30 μg/kg) failed to alter nociceptive withdrawal reflexes at any age examined compared with vehicle-injected littermate controls. Meanwhile, high dose VNC (60 μg/kg) evoked mechanical hypersensitivity in both sexes beginning at P26 that persisted until adulthood and included both static and dynamic mechanical allodynia. Hind paw withdrawal latencies to noxious heat and cold were unaffected by high doses of VNC, suggesting a selective effect of neonatal VNC on mechanical pain sensitivity. Gross and fine motor function appeared normal after VNC treatment, although a small decrease in weight gain was observed. The VNC regimen also produced a significant decrease in intraepidermal nerve fiber density in the hind paw skin by P33. Overall, the present results demonstrate that high-dose administration of VNC during the early postnatal period selectively evokes a mechanical hypersensitivity that is slow to emerge during adolescence, providing further evidence that aberrant sensory input during early life can have prolonged consequences for pain processing.

1-(2,4-Dibromophenyl)-3,6,6-trimethyl-1,5,6,7-tetrahydro-4H-indazol-4-one: A Novel Opioid Receptor Agonist with Less Accompanying Gastrointestinal Dysfunction than Morphine.

The authors investigated the pharmacology and signaling pathways of the opioid receptors modulated by compound 1, 1-(2,4-dibromophenyl)-3,6,6-trimethyl-1,5,6,7-tetrahydro-4H-indazol-4-one. In vitro studies of compound 1 were assessed by using a radioligand-binding assay (n = 3), a cyclic adenosine monophosphate assay (n = 3), a β-arrestin assay (n = 3), an internalization assay (n = 3), and an immunohistochemistry (n = 8). In vivo studies of compound 1 were characterized using a tail-flick test (n = 5 to 6), tail-clip test (n = 7), von Frey hair test (n = 5), and charcoal meal test (n = 5). Compound 1 elicited robust effects in μ-opioid (mean ± SD; binding affinity: 15 ± 2 nM; cyclic adenosine monophosphate assay: 24 ± 6 nM), δ-opioid (82 ± 7 nM; 1.9 ± 0.1 μM), and κ-opioid (76 ± 9 nM; 1.4 ± 0.5 μM) receptor-expressing cells. Compound 1 acts as a full agonist of β-arrestin-2 recruitment in μ-opioid (1.1 ± 0.3 μM) and δ-opioid (9.7 ± 1.9 μM) receptor-expressing cells. Compound 1 caused less gastrointestinal dysfunction (charcoal meal test: morphine: 82 ± 5%; compound 1: 42 ± 5%) as well as better antinociception in mechanical pain hypersensitivity (tail-clip test: morphine: 10 ± 3 s; compound 1: 19 ± 1 s) and in cancer-induced pain (von Frey hair test: morphine: 0.1 ± 0.1 g; compound 1: 0.3 ± 0.1 g) than morphine at equi-antinociceptive doses. Compound 1 produced antinociception with less gastrointestinal dysfunction than morphine.