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Bone cancer pain (BCP) represents a prevalent symptom among cancer patients with bone metastases, yet its underlying mechanisms remain elusive. This study investigated the transcriptional regulation mechanism of Kv7(KCNQ)/M potassium channels in DRG neurons and its involvement in the development of BCP in rats. We show that HDAC2-mediated transcriptional repression of kcnq2/kcnq3 genes, which encode Kv7(KCNQ)/M potassium channels in dorsal root ganglion (DRG), contributes to the sensitization of DRG neurons and the pathogenesis of BCP in rats. Also, HDAC2 requires the formation of a corepressor complex with MeCP2 and Sin3A to execute transcriptional regulation of kcnq2/kcnq3 genes. Moreover, EREG is identified as an upstream signal molecule for HDAC2-mediated kcnq2/kcnq3 genes transcription repression. Activation of EREG/EGFR-ERK-Runx1 signaling, followed by the induction of HDAC2-mediated transcriptional repression of kcnq2/kcnq3 genes in DRG neurons, leads to neuronal hyperexcitability and pain hypersensitivity in tumor-bearing rats. Consequently, the activation of EREG/EGFR-ERK-Runx1 signaling, along with the subsequent transcriptional repression of kcnq2/kcnq3 genes by HDAC2 in DRG neurons, underlies the sensitization of DRG neurons and the pathogenesis of BCP in rats. These findings uncover a potentially targetable mechanism contributing to bone metastasis-associated pain in cancer patients.
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Neoplasias Ósseas , Dor do Câncer , Receptores ErbB , Gânglios Espinais , Histona Desacetilase 2 , Canal de Potássio KCNQ2 , Animais , Histona Desacetilase 2/metabolismo , Histona Desacetilase 2/genética , Canal de Potássio KCNQ2/genética , Canal de Potássio KCNQ2/metabolismo , Gânglios Espinais/metabolismo , Gânglios Espinais/patologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/secundário , Neoplasias Ósseas/patologia , Ratos , Dor do Câncer/genética , Dor do Câncer/metabolismo , Dor do Câncer/patologia , Receptores ErbB/metabolismo , Receptores ErbB/genética , Canal de Potássio KCNQ3/genética , Canal de Potássio KCNQ3/metabolismo , Transcrição Gênica , Proteína 2 de Ligação a Metil-CpG/genética , Proteína 2 de Ligação a Metil-CpG/metabolismo , Complexo Correpressor Histona Desacetilase e Sin3/genética , Transdução de Sinais/genética , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Humanos , Feminino , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Ratos Sprague-Dawley , Sistema de Sinalização das MAP Quinases/genéticaRESUMO
Cisplatin-based chemotherapy is a common treatment for paediatric cancer. Unfortunately, cisplatin treatment causes neuropathic pain, a highly prevalent adverse health related complication in adult childhood cancer survivors. Due to minimal understanding of this condition, there are currently no condition tailored analgesics available. Here we investigated an alteration in nociceptor maturation that results in neuronal sensitisation and manifestation of cisplatin induced survivorship pain in a TrkA dependent manner. Cisplatin was administered (i.p. 0.1â¯mg/kg Postnatal day 14 and 16) to neonatal male and female Wistar rats and nociceptive behavioural assays were performed. In vitro studies utilised isolated neonatal dorsal root ganglia sensory neurons treated with cisplatin (5⯵g/ml) to elucidate impact upon nociceptor activation and neurite growth, in combination with TrkA inhibition (GW441756 10â¯nM and 100â¯nM). Cisplatin treated male and female neonatal Wistar rats developed a delayed but lasting mechanical and heat hypersensitivity. Cisplatin administration led to increased TrkA expression in dorsal root ganglia sensory neurons. Nerve growth factor (NGF) induced TrkA activation led to sensory neuritogenesis and nociceptor sensitisation, which could be prevented through pharmacological TrkA inhibition (GW441756 either s.c. 100â¯nM or i.p. 2â¯mg/kg). Administration of TrkA antagonist suppressed cisplatin induced TRPV1 mediated nociceptor sensitisation and prevented cisplatin induced neuropathic pain. These studies provide greater understanding of the underlying mechanisms that cause cisplatin induced childhood cancer survivorship pain and allowing identification of potential therapeutic targets.
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BACKGROUND: Delivering optogenetic genes to the peripheral sensory nervous system provides an efficient approach to study and treat neurological disorders and offers the potential to reintroduce sensory feedback to prostheses users and those who have incurred other neuropathies. Adeno-associated viral (AAV) vectors are a common method of gene delivery due to efficiency of gene transfer and minimal toxicity. AAVs are capable of being designed to target specific tissues, with transduction efficacy determined through the combination of serotype and genetic promoter selection, as well as location of vector administration. The dorsal root ganglia (DRGs) are collections of cell bodies of sensory neurons which project from the periphery to the central nervous system (CNS). The anatomical make-up of DRGs make them an ideal injection location to target the somatosensory neurons in the peripheral nervous system (PNS). COMPARISON TO EXISTING METHODS: Previous studies have detailed methods of direct DRG injection in rats and dorsal horn injection in mice, however, due to the size and anatomical differences between rats and strains of mice, there is only one other published method for AAV injection into murine DRGs for transduction of peripheral sensory neurons using a different methodology. NEW METHOD/RESULTS: Here, we detail the necessary materials and methods required to inject AAVs into the L3 and L4 DRGs of mice, as well as how to harvest the sciatic nerve and L3/L4 DRGs for analysis. This methodology results in optogenetic expression in both the L3/L4 DRGs and sciatic nerve and can be adapted to inject any DRG.
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Over half of spinal cord injury (SCI) patients develop opioid-resistant chronic neuropathic pain. Safer alternatives to opioids for treatment of neuropathic pain are gabapentinoids (e.g., pregabalin and gabapentin). Clinically, gabapentinoids appear to amplify opioid effects, increasing analgesia and overdose-related adverse outcomes, but in vitro proof of this amplification and its mechanism are lacking. We previously showed that after SCI, sensitivity to opioids is reduced by fourfold to sixfold in rat sensory neurons. Here, we demonstrate that after injury, gabapentinoids restore normal sensitivity of opioid inhibition of cyclic AMP (cAMP) generation, while reducing nociceptor hyperexcitability by inhibiting voltage-gated calcium channels (VGCCs). Increasing intracellular Ca2+ or activation of L-type VGCCs (L-VGCCs) suffices to mimic SCI effects on opioid sensitivity, in a manner dependent on the activity of the Raf1 proto-oncogene, serine/threonine-protein kinase C-Raf, but independent of neuronal depolarization. Together, our results provide a mechanism for potentiation of opioid effects by gabapentinoids after injury, via reduction of calcium influx through L-VGCCs, and suggest that other inhibitors targeting these channels may similarly enhance opioid treatment of neuropathic pain.
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Analgésicos Opioides , AMP Cíclico , Gabapentina , Neuralgia , Transdução de Sinais , Traumatismos da Medula Espinal , Animais , Neuralgia/tratamento farmacológico , Neuralgia/metabolismo , AMP Cíclico/metabolismo , Ratos , Traumatismos da Medula Espinal/tratamento farmacológico , Traumatismos da Medula Espinal/metabolismo , Analgésicos Opioides/farmacologia , Gabapentina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Ratos Sprague-Dawley , Masculino , Canais de Cálcio Tipo L/metabolismo , Cálcio/metabolismo , Pregabalina/farmacologia , Pregabalina/uso terapêutico , Sinergismo Farmacológico , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/efeitos dos fármacosRESUMO
Lysophosphatidic acids (LPAs) evoke nociception and itch in mice and humans. In this study, we assessed the signaling paths. Hydroxychloroquine was injected intradermally to evoke itch in mice, which evoked an increase of LPAs in the skin and in the thalamus, suggesting that peripheral and central LPA receptors (LPARs) were involved in HCQ-evoked pruriception. To unravel the signaling paths, we assessed the localization of candidate genes and itching behavior in knockout models addressing LPAR5, LPAR2, autotaxin/ENPP2 and the lysophospholipid phosphatases, as well as the plasticity-related genes Prg1/LPPR4 and Prg2/LPPR3. LacZ reporter studies and RNAscope revealed LPAR5 in neurons of the dorsal root ganglia (DRGs) and in skin keratinocytes, LPAR2 in cortical and thalamic neurons, and Prg1 in neuronal structures of the dorsal horn, thalamus and SSC. HCQ-evoked scratching behavior was reduced in sensory neuron-specific Advillin-LPAR5-/- mice (peripheral) but increased in LPAR2-/- and Prg1-/- mice (central), and it was not affected by deficiency of glial autotaxin (GFAP-ENPP2-/-) or Prg2 (PRG2-/-). Heat and mechanical nociception were not affected by any of the genotypes. The behavior suggested that HCQ-mediated itch involves the activation of peripheral LPAR5, which was supported by reduced itch upon treatment with an LPAR5 antagonist and autotaxin inhibitor. Further, HCQ-evoked calcium fluxes were reduced in primary sensory neurons of Advillin-LPAR5-/- mice. The results suggest that LPA-mediated itch is primarily mediated via peripheral LPAR5, suggesting that a topical LPAR5 blocker might suppress "non-histaminergic" itch.
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Hidroxicloroquina , Camundongos Knockout , Prurido , Receptores de Ácidos Lisofosfatídicos , Animais , Receptores de Ácidos Lisofosfatídicos/metabolismo , Receptores de Ácidos Lisofosfatídicos/genética , Prurido/induzido quimicamente , Prurido/metabolismo , Prurido/genética , Prurido/tratamento farmacológico , Camundongos , Hidroxicloroquina/farmacologia , Gânglios Espinais/metabolismo , Gânglios Espinais/efeitos dos fármacos , Masculino , Diester Fosfórico Hidrolases/metabolismo , Diester Fosfórico Hidrolases/genética , Lisofosfolipídeos/metabolismo , Camundongos Endogâmicos C57BL , Transdução de Sinais/efeitos dos fármacosRESUMO
Neurological symptoms associated with COVID-19, acute and long term, suggest SARS-CoV-2 affects both the peripheral and central nervous systems (PNS/CNS). Although studies have shown olfactory and hematogenous invasion into the CNS, coinciding with neuroinflammation, little attention has been paid to susceptibility of the PNS to infection or to its contribution to CNS invasion. Here we show that sensory and autonomic neurons in the PNS are susceptible to productive infection with SARS-CoV-2 and outline physiological and molecular mechanisms mediating neuroinvasion. Our infection of K18-hACE2 mice, wild-type mice, and golden Syrian hamsters, as well as primary peripheral sensory and autonomic neuronal cultures, show viral RNA, proteins, and infectious virus in PNS neurons, satellite glial cells, and functionally connected CNS tissues. Additionally, we demonstrate, in vitro, that neuropilin-1 facilitates SARS-CoV-2 neuronal entry. SARS-CoV-2 rapidly invades the PNS prior to viremia, establishes a productive infection in peripheral neurons, and results in sensory symptoms often reported by COVID-19 patients.
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COVID-19 , Neuropilina-1 , SARS-CoV-2 , Animais , SARS-CoV-2/fisiologia , SARS-CoV-2/patogenicidade , COVID-19/virologia , COVID-19/patologia , COVID-19/metabolismo , Camundongos , Neuropilina-1/metabolismo , Neuropilina-1/genética , Viremia/virologia , Sistema Nervoso Central/virologia , Sistema Nervoso Central/patologia , Sistema Nervoso Central/metabolismo , Células Receptoras Sensoriais/virologia , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/patologia , Mesocricetus , Humanos , Enzima de Conversão de Angiotensina 2/metabolismo , Camundongos Endogâmicos C57BL , Internalização do Vírus , MasculinoRESUMO
The cell intrinsic mechanisms directing peripheral nerve regeneration have remained largely understudied, thus limiting our understanding of these processes and constraining the advancement of novel clinical therapeutics. The use of primary adult rat dorsal root ganglion (DRG) neurons cultured in vitro is well established. Despite this, these cells can be challenging to culture and have so far not been amenable to robust transfection or live-cell imaging. The ability to transfect these cells with fluorescent plasmid constructs to label subcellular structures, combined with high resolution time-lapse imaging has the potential to provide invaluable insight into how peripheral neurons coordinate their regenerative response, and which specific cellular structures are involved in this process. Here we describe a protocol that facilitates transfection and subsequent live-imaging of adult rat DRG neurons.
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Gânglios Espinais , Regeneração Nervosa , Neurônios , Animais , Gânglios Espinais/citologia , Regeneração Nervosa/fisiologia , Ratos , Neurônios/citologia , Neurônios/fisiologia , Neurônios/metabolismo , Células Cultivadas , Transfecção/métodos , Imagem com Lapso de Tempo/métodosRESUMO
Studies indicate that the lysine-specific demethylase 4A (KDM4A), acts as a key player in neuropathic pain, driving the process through its involvement in promoting neuroinflammation. Emerging evidence reveals that C-C Motif Chemokine Ligand 2 (CCL2) participates in neuroinflammation, which plays an important role in the development and maintenance of neuropathic pain. However, it remains unclear if KDM4A plays a role in regulating CCL2 in neuropathic pain. This study found that following spinal nerve transection (SNT) of the lumbar 5 nerve root in rats, the expression of KDM4A and CCL2 increased in the ipsilateral L4/5 dorsal root ganglia (DRG). Injecting KDM4A siRNA into the DRGs of rats post-SNT resulted in a higher paw withdrawal threshold (PWT) and paw-withdrawal latency (PWL) compared to the KDM4A scRNA group. In addition, prior microinjection of AAV-EGFP-KDM4A shRNA also alleviates the decrease in PWT and PWL caused by SNT. Correspondingly, microinjection of AAV-EGFP-KDM4A shRNA subsequent to SNT reduced the established mechanical and thermal hyperalgesia. Furthermore, AAV-EGFP-KDM4A shRNA injection decreased the expression of CCL2 in DRGs. ChIP-PCR analysis revealed that increased binding of p-STAT1 with the CCL2 promoter induced by SNT was inhibited by AAV-EGFP-KDM4A shRNA treatment. These findings suggest that KDM4A potentially influences neuropathic pain by regulating CCL2 expression in DRGs.
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Prostaglandin E2 (PGE2) is a major contributor to inflammatory pain hyperalgesia, however, the extent to which it modulates the activity of nociceptive axons is incompletely understood. We developed and characterized a microfluidic cell culture model to investigate sensitisation of the axons of dorsal root ganglia neurons. We show that application of PGE2 to fluidically isolated axons leads to sensitisation of their responses to depolarising stimuli. Interestingly the application of PGE2 to the DRG axons elicited a direct and persistent spiking activity propagated to the soma. Both the persistent activity and the membrane depolarisation in the axons are abolished by the EP4 receptor inhibitor and a blocker of cAMP synthesis. Further investigated into the mechanisms of the spiking activity showed that the PGE2 evoked depolarisation was inhibited by Nav1.8 sodium channel blockers but was refractory to the application of TTX or zatebradine. Interestingly, the depolarisation of axons was blocked by blocking ANO1 channels with T16Ainh-A01. We further show that PGE2-elicited axonal responses are altered by the changes in chloride gradient within the axons following treatment with bumetanide a Na-K-2Cl cotransporter NKCC1 inhibitor, but not by VU01240551 an inhibitor of potassium-chloride transporter KCC2. Our data demonstrate a novel role for PGE2/EP4/cAMP pathway which culminates in a sustained depolarisation of sensory axons mediated by a chloride current through ANO1 channels. Therefore, using a microfluidic culture model, we provide evidence for a potential dual function of PGE2 in inflammatory pain: it sensitises depolarisation-evoked responses in nociceptive axons and directly triggers action potentials by activating ANO1 and Nav1.8 channels.
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Anoctamina-1 , Axônios , Dinoprostona , Gânglios Espinais , Canal de Sódio Disparado por Voltagem NAV1.8 , Canal de Sódio Disparado por Voltagem NAV1.8/metabolismo , Animais , Dinoprostona/farmacologia , Dinoprostona/metabolismo , Axônios/metabolismo , Axônios/efeitos dos fármacos , Axônios/fisiologia , Gânglios Espinais/metabolismo , Gânglios Espinais/efeitos dos fármacos , Ratos , Anoctamina-1/metabolismo , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/efeitos dos fármacos , Potenciais de Ação/efeitos dos fármacos , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Ratos Sprague-Dawley , Células Cultivadas , Membro 2 da Família 12 de Carreador de Soluto/metabolismo , AMP Cíclico/metabolismoRESUMO
GRT-X, which targets both the mitochondrial translocator protein (TSPO) and the Kv7.2/3 (KCNQ2/3) potassium channels, has been shown to efficiently promote recovery from cervical spine injury. In the present work, we investigate the role of GRT-X and its two targets in the axonal growth of dorsal root ganglion (DRG) neurons. Neurite outgrowth was quantified in DRG explant cultures prepared from wild-type C57BL6/J and TSPO-KO mice. TSPO was pharmacologically targeted with the agonist XBD173 and the Kv7 channels with the activator ICA-27243 and the inhibitor XE991. GRT-X efficiently stimulated DRG axonal growth at 4 and 8 days after its single administration. XBD173 also promoted axonal elongation, but only after 8 days and its repeated administration. In contrast, both ICA27243 and XE991 tended to decrease axonal elongation. In dissociated DRG neuron/Schwann cell co-cultures, GRT-X upregulated the expression of genes associated with axonal growth and myelination. In the TSPO-KO DRG cultures, the stimulatory effect of GRT-X on axonal growth was completely lost. However, GRT-X and XBD173 activated neuronal and Schwann cell gene expression after TSPO knockout, indicating the presence of additional targets warranting further investigation. These findings uncover a key role of the dual mode of action of GRT-X in the axonal elongation of DRG neurons.
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Axônios , Gânglios Espinais , Receptores de GABA , Animais , Gânglios Espinais/metabolismo , Gânglios Espinais/citologia , Camundongos , Axônios/metabolismo , Receptores de GABA/metabolismo , Receptores de GABA/genética , Canal de Potássio KCNQ2/metabolismo , Canal de Potássio KCNQ2/genética , Camundongos Knockout , Camundongos Endogâmicos C57BL , Células Cultivadas , Células de Schwann/metabolismo , Células de Schwann/efeitos dos fármacos , Células de Schwann/citologia , Técnicas de Cocultura , Neurônios/metabolismo , Neurônios/efeitos dos fármacosRESUMO
The somatosensory system detects peripheral stimuli that are translated into behaviors necessary for survival. Fishes and amphibians possess two somatosensory systems in the trunk: the primary somatosensory system, formed by the Rohon-Beard neurons, and the secondary somatosensory system, formed by the neural crest cell-derived neurons of the Dorsal Root Ganglia. Rohon-Beard neurons have been characterized as a transient population that mostly disappears during the first days of life and is functionally replaced by the Dorsal Root Ganglia. Here, I follow Rohon-Beard neurons in vivo and show that the entire repertoire remains present in zebrafish from 1-day post-fertilization until the juvenile stage, 15-days post-fertilization. These data indicate that zebrafish retain two complete somatosensory systems until at least a developmental stage when the animals display complex behavioral repertoires.
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Peixe-Zebra , Animais , Peixe-Zebra/embriologia , Gânglios Espinais/embriologia , Neurônios/fisiologia , Crista Neural/citologia , Crista Neural/embriologia , Crista Neural/fisiologiaRESUMO
Persistent hyperactivity of nociceptors is known to contribute significantly to long-lasting sensitization and ongoing pain in many clinical conditions. It is often assumed that nociceptor hyperactivity is mainly driven by continuing stimulation from inflammatory mediators. We have tested an additional possibility: that persistent increases in excitability promoting hyperactivity can be induced by a prototypical cellular signaling pathway long known to induce late-phase long-term potentiation (LTP) of synapses in brain regions involved in memory formation. This cAMP-PKA-CREB-gene transcription-protein synthesis pathway was tested using whole-cell current clamp methods on small dissociated sensory neurons (primarily nociceptors) from dorsal root ganglia (DRGs) excised from previously uninjured ("naïve") rats. Six-hour treatment with the specific Gαs-coupled 5-HT4 receptor agonist, prucalopride, or with the adenylyl cyclase activator, forskolin, induced long-term hyperexcitability (LTH) in DRG neurons that manifested 12-24 hours later as action potential (AP) discharge (ongoing activity, OA) during artificial depolarization to -45 mV, a membrane potential that is normally subthreshold for AP generation. Prucalopride treatment also induced significant long-lasting depolarization of resting membrane potential (from -69 to -66 mV), enhanced depolarizing spontaneous fluctuations (DSFs) of membrane potential, and indications of reduced AP threshold and rheobase. LTH was prevented by co-treatment of prucalopride with inhibitors of PKA, CREB, gene transcription, and protein synthesis. As in the induction of synaptic memory, many other cellular signals are likely to be involved. However, the discovery that this prototypical memory induction pathway can induce nociceptor LTH, along with reports that cAMP signaling and CREB activity in DRGs can induce hyperalgesic priming, suggest that early, temporary, cAMP-induced transcriptional and translational mechanisms can induce nociceptor LTH that might last for long periods. An interesting possibility is that these mechanisms can also be reactivated by re-exposure to inflammatory mediators such as serotonin during subsequent challenges to bodily integrity, "reconsolidating" the cellular memory and thereby extending the duration of persistent nociceptor hyperexcitability.
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The etiology of interstitial cystitis/bladder pain syndrome (IC/BPS) is unknown but likely multifactorial. IC/BPS symptoms can be exacerbated by psychological stress, but underlying mechanisms remain to be defined. Transient receptor potential vanilloid 1 (TRPV1) channels, expressed on nerve fibers, have been implicated in bladder dysfunction and colonic hypersensitivity with stress in rodents. Histamine/H1R activation of TRPV1+ nerves increases bladder afferent fiber sensitivity to distension. TRPV1 channels are also expressed on mast cells, previously implicated in contributing to IC/BPS etiology and symptoms. We have examined the contribution of TRPV1 and mast cells to bladder dysfunction after repeated variate stress (RVS). RVS increased (P ≤ 0.05) serum and fecal corticosterone expression and induced anxiety-like behavior in wild-type (WT) mice. Intravesical instillation of the selective TRPV1 antagonist capsazepine (CPZ) rescued RVS-induced bladder dysfunction in WT mice. Trpv1 knockout (KO) mice did not increase voiding frequency with RVS and did not exhibit increased serum corticosterone expression despite exhibiting anxiety-like behavior. Mast cell-deficient mice (B6.Cg-Kitw-sh) failed to demonstrate RVS-induced increased voiding frequency or serum corticosterone expression, whereas control (no stress) mast cell-deficient mice had similar functional bladder capacity to WT mice. TRPV1 protein expression was significantly increased in the rostral lumbar (L1-L2) spinal cord and dorsal root ganglia (DRG) in WT mice exposed to RVS, but no changes were observed in lumbosacral (L6-S1) spinal segments or DRG. These studies demonstrated TRPV1 and mast cell involvement in RVS-induced increased voiding frequency and suggest that TRPV1 and mast cells may be useful targets to mitigate stress-induced urinary bladder dysfunction.NEW & NOTEWORTHY Using pharmacological tools and transgenic mice in a repeated variate stress (RVS) model in female mice, we demonstrate that transient receptor potential vanilloid 1 (TRPV1) and mast cells contribute to the increased voiding frequency observed following RVS. TRPV1 and mast cells should continue to be considered as targets to improve bladder function in stress-induced bladder dysfunction.
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Corticosterona , Mastócitos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Estresse Psicológico , Canais de Cátion TRPV , Bexiga Urinária , Animais , Canais de Cátion TRPV/metabolismo , Canais de Cátion TRPV/genética , Mastócitos/metabolismo , Feminino , Bexiga Urinária/metabolismo , Bexiga Urinária/inervação , Estresse Psicológico/complicações , Estresse Psicológico/metabolismo , Corticosterona/sangue , Modelos Animais de Doenças , Cistite Intersticial/metabolismo , Cistite Intersticial/fisiopatologia , Cistite Intersticial/patologia , Cistite Intersticial/genética , Camundongos , Micção , Capsaicina/farmacologia , Capsaicina/análogos & derivados , Comportamento Animal , Ansiedade/metabolismoRESUMO
Peripheral nerve damage often leads to the onset of neuropathic pain (NeuP). This condition afflicts millions of people, significantly burdening healthcare systems and putting strain on families' financial well-being. Here, we will focus on the role of peripheral sensory neurons, specifically the Dorsal Root Ganglia neurons (DRG neurons) in the development of NeuP. After axotomy, DRG neurons activate regenerative signals of axons-soma communication to promote a gene program that activates an axonal branching and elongation processes. The results of a neuronal morphological cytoskeleton change are not always associated with functional recovery. Moreover, any axonal miss-targeting may contribute to NeuP development. In this review, we will explore the epidemiology of NeuP and its molecular causes at the level of the peripheral nervous system and the target organs, with major focus on the neuronal cross-talk between intrinsic and extrinsic factors. Specifically, we will describe how failures in the neuronal regenerative program can exacerbate NeuP.
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Introduction: The distributions of extrinsic neurons innervating the colon show differences in experimental animals from humans, including the vagal and spinal parasympathetic innervation to the distal colon. The neuroanatomical tracing to the mouse proximal colon has not been studied in details. This study aimed to trace the locations of extrinsic neurons projecting to the mouse proximal colon compared to the distal colon using dual retrograde tracing. Methods: The parasympathetic and sensory neurons projecting to colon were assessed using Cholera Toxin subunit B conjugated to Alexa-Fluor 488 or 555 injected in the proximal and distal colon of the same mice. Results: Retrograde tracing from the proximal and distal colon labeled neurons in the dorsal motor nucleus of the vagus (DMV) and the nodose ganglia, while the tracing from the distal colon did not label the parasympathetic neurons in the lumbosacral spinal cord at L6-S1. Neurons in the pelvic ganglia which were cholinergic projected to the distal colon. There were more neurons in the DMV and nodose ganglia projecting to the proximal than distal colon. The right nodose ganglion had a higher number of neurons than the left ganglion innervating the proximal colon. In the dorsal root ganglia (DRG), the highest number of neurons traced from the distal colon were at L6, and those from the proximal colon at T12. DRG neurons projected closely to the cholinergic neurons in the intermediolateral column of L6 spinal cord. Small percentages of neurons with dual projections to both the proximal and distal colon existed in the DMV, nodose ganglia and DRG. We also observed long projecting neurons traced from the caudal distal colon to the transverse and proximal colon, some of which were calbindin immunoreactive, while there were no retrogradely labeled neurons traced from the proximal to distal colon. Discussion: These data demonstrated that the vagal motor and motor and sensory neurons innervate both the proximal and distal colon in mice, and the autonomic neurons in the intermediate zone of the lumbosacral spinal cord do not project directly to the mouse colon, which differs from that in humans.
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BACKGROUND: Cervicogenic headache is characterized by chronic posterior neck pain radiating to one side of the head, resulting from cervical spine bone or soft tissue diseases. Cervical ganglioneuroma (GN), a rare benign neuroblastic tumor, especially in the cervical spine, may cause cervicogenic headache-like symptoms. OBJECTIVE: We report a case of GN which was surgically removed successfully to relieve the symptom. CASE REPORT: A 68-year-old male presented with right posterior neck pain with referred pain to the ipsilateral occipital area in May 2020. Despite administration of medications, physical therapy, and spine interventions, the symptoms intermittently recurred over one year. In July 2021, the patient complained of painful limitation of neck motion, especially on right-sided bending; no motor or sensory deficits, except for subjective numbness of the finger tips, were detected. Plain radiography of the cervical spine showed moderate degenerative changes in the mid-cervical spine. Cervical MRI revealed a cystic mass (1.5 cm × 0.5 cm × 1 cm-in size) around the right C2 dorsal root ganglion adjacent to the C1-C2 facet joint. His symptoms significantly improved after complete tumor excision. CONCLUSION: GN of the upper cervical spine should be considered when persistent cervicogenic headache is refractory to conservative management. In such a case, advanced imaging studies such as MRI should be performed for early diagnosis and appropriate treatment.
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Introduction: Peripheral sensory neurons serve as the initial responders to the external environment. How these neurons react to different sensory stimuli, such as mechanical or thermal forces applied to the skin, remains unclear. Methods: Using in vivo two-photon Ca2+ imaging in the lumbar 4 dorsal root ganglion (DRG) of awake Thy1.2-GCaMP6s mice, we assessed neuronal responses to various mechanical (punctate or dynamic) and thermal forces (heat or cold) sequentially applied to the paw plantar surface. Results: Our data indicate that in normal awake male mice, approximately 14 and 38% of DRG neurons respond to either single or multiple modalities of stimulation. Anesthesia substantially reduces the number of responsive neurons but does not alter the ratio of cells exhibiting single-modal responses versus multi-modal responses. Following peripheral nerve injury, DRG cells exhibit a more than 5.1-fold increase in spontaneous neuronal activity and a 1.5-fold increase in sensory stimulus-evoked activity. As neuropathic pain resulting from nerve injury progresses, the polymodal nature of sensory neurons intensifies. The polymodal population increases from 39.1 to 56.9%, while the modality-specific population decreases from 14.7 to 5.0% within a period of 5 days. Discussion: Our study underscores polymodality as a significant characteristic of primary sensory neurons, which becomes more pronounced during the development of neuropathic pain.
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Currently, the clinical treatment of bone cancer pain (BCP) is mainly related to its pathogenesis. The aim of the present study was to elucidate the potential role of N6-methyladenosine (m6A) in BCP in the spinal cord dorsal root ganglia (DRG) of BCP rats and its specific regulatory mechanism in N-methyl-d-aspartate receptor subunit 2B (NR2B). A rat model of BCP was constructed by tibial injection of Walker256 cells, and ALKBH5 and NR2B expression in the spinal cord DRG was detected. ALKBH5 was silenced or overexpressed in PC12 cells to verify the regulatory effect of ALKBH5 on NR2B. The specific mechanism underlying the interaction between ALKBH5 and NR2B was investigated using methylated RNA immunoprecipitation and dual-luciferase reporter gene assays. The results showed increased expression of m6A, decreased expression of ALKBH5, and increased expression of NR2B in the DRG of the BCP rat model. Overexpression of ALKBH5 inhibited NR2B expression, whereas interference with ALKBH5 caused an increase in NR2B expression. In NR2B, interference with ALKBH5 caused an increase in m6A modification, which caused an increase in NR2B. Taken together, ALKBH5 affected the expression of NR2B by influencing the stability of the m6A modification site of central NR2B, revealing that ALKBH5 is a therapeutic target for BCP.
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Metastatic cancers often lead to distant metastasis, accompanied by debilitating symptoms such as chronic pain, which can be refractory to conventional analgesic modalities. Pulsed radiofrequency ablation (Pulsed RFA) has emerged as a promising intervention for neuropathic pain syndromes, offering long-lasting relief with minimal tissue damage. We present a case of a 36-year-old male with metastatic gastric adenocarcinoma and refractory leg pain due to femoral nerve involvement. Despite aggressive multimodal analgesia, the patient experienced persistent pain, necessitating alternative interventions. Pulsed RFA targeting the right L2-L4 dorsal root ganglia (DRG) provided significant and sustained pain relief, allowing improved functional status and reduced opioid requirements. This case underscores the potential of pulsed RFA as an effective intervention for refractory cancer-related pain, enhancing patients' comfort and quality of life. Further research is warranted to establish its long-term efficacy and safety.
RESUMO
Neuropathic pain (NP) is a common debilitating chronic pain condition with limited effective therapeutics. Further investigating mechanisms underlying NP is therefore of great importance for discovering more promising therapeutic targets. In the current study, we employed high-throughput RNA sequencing to explore transcriptome profiles of mRNAs and microRNAs in the dorsal root ganglia (DRG) following chronic constriction injury (CCI) and also integrated published datasets for comprehensive analysis. First, we established CCI rat model confirmed by behavioral testings, and excavated 467 differentially expressed mRNAs (DEGs) and 16 differentially expressed microRNAs (DEmiRNAs) in the ipsilateral lumbar 4-6 DRG of CCI rats 11 days after surgery. Functional enrichment analysis of 337 upregulated DEGs showed that most of the DEGs were enriched in inflammation- and immune-associated biological processes and signaling pathways. The protein-protein interaction networks were constructed and hub DEGs were screened. Besides hub DEGs, we also identified 113 overlapped DEGs by intersecting our dataset with dataset GSE100122. Subsequently, we predicted potential miRNA-mRNA regulatory pairs using DEmiRNAs and a given set of key DEGs (including hub and overlapped DEGs). By integrative analysis, we found commonly differentially expressed mRNAs and miRNAs following CCI of different time points and different nerve injury types. Highlighted mRNAs include Atf3, Vip, Gal, Npy, Adcyap1, Reg3b, Jun, Cd74, Gadd45a, Tgm1, Csrp3, Sprr1a, Serpina3n, Gap43, Serpinb2 and Vtcn1, while miRNAs include miR-21-5p, miR-34a-5p, miR-200a-3p, miR-130a-5p, miR-216b-5p, miR-217-5p, and miR-541-5p. Additionally, 15 DEGs, including macrophages-specific (Cx3cr1, Arg1, Cd68, Csf1r) and the ones related to macrophages' involvement in NP (Ccl2, Fcgr3a, Bdnf, Ctss, Tyrobp) were verified by qRT-PCR. By functional experiments in future studies, promising therapeutic targets for NP treatment may be identified among these mRNAs and miRNAs.