Thiamine Suppresses Thermal Hyperalgesia, Inhibits Hyperexcitability, and Lessens Alterations of Sodium Currents in Injured, Dorsal Root Ganglion Neurons in Rats

Background B vitamins can effectively attenuate inflammatory and neuropathic pain in experimental animals, while their efficacy in treating clinical pain syndromes remains unclear. To understand possible mechanisms underlying B vitamin-induced analgesia and provide further evidence that may support the clinical utility of B vitamins in chronic pain treatment, this study investigated effects of thiamine (B1) on the excitability and Na currents of dorsal root ganglion (DRG) neurons that have been altered by nerve injury. Methods Nerve injury was mimicked by chronic compression of DRG in rats. Neuropathic pain was evidenced by the presence of thermal hyperalgesia. Intracellular and patch-clamp recordings were made in vitro from intact and dissociated DRG neurons, respectively. Results (1) In vivo intraperitoneal administration of B1 (66 mg/kg/day, 10-14 doses) significantly inhibited DRG compression-induced neural hyperexcitability, in addition to suppressing thermal hyperalgesia. (2) In vitro perfusion of B1 (0.1, 1 and 10 mM) resulted in a dose-dependent inhibition of DRG neuron hyperexcitability. In addition, the DRG neurons exhibited size-dependent sensitivity to B1 treatment, i.e., the small and the medium-sized neurons, compared to the large neurons, were significantly more sensitive. (3) Both in vitro (1 mM) and in vivo application of B1 significantly reversed DRG compression-induced down-regulation of tetrodotoxin-resistant but not tetrodotoxin-sensitive Na current density in the small neurons. B1 at 1 mM also reversed the compression-induced hyperpolarizing shift of the inactivation curve of the tetrodotoxin-resistant currents and the upregulated ramp currents in small DRG neurons. Conclusion Thiamine can reduce hyperexcitability and lessen alterations of Na currents in injured DRG neurons, in addition to suppressing thermal hyperalgesia.

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Similar Electrophysiological Changes in Axotomized and Neighboring Intact Dorsal Root Ganglion Neurons

Journal of Neurophysiology ◽

10.1152/jn.00855.2002 ◽

2003 ◽

Vol 89 (3) ◽

pp. 1588-1602 ◽

Cited By ~ 168

Author(s):

Chao Ma ◽

Yousheng Shu ◽

Zheng Zheng ◽

Yong Chen ◽

Hang Yao ◽

...

Keyword(s):

Dorsal Root Ganglion ◽

Dorsal Root ◽

Receptive Fields ◽

Input Resistance ◽

Spinal Nerve ◽

Dorsal Root Ganglion Neurons ◽

Drg Neurons ◽

Ganglion Neurons

We investigated electrophysiological changes in chronically axotomized and neighboring intact dorsal root ganglion (DRG) neurons in rats after either a peripheral axotomy consisting of an L5 spinal nerve ligation (SNL) or a central axotomy produced by an L5 partial rhizotomy (PR). SNL produced lasting hyperalgesia to punctate indentation and tactile allodynia to innocuous stroking of the foot ipsilateral to the injury. PR produced ipsilateral hyperalgesia without allodynia with recovery by day 10. Intracellular recordings were obtained in vivo from the cell bodies (somata) of axotomized and intact DRG neurons, some with functionally identified peripheral receptive fields. PR produced only minor electrophysiological changes in both axotomized and intact somata in L5 DRG. In contrast, extensive changes were observed after SNL in large- and medium-sized, but not small-sized, somata of intact (L4) as well as axotomized (L5) DRG neurons. These changes included (in relation to sham values) higher input resistance, lower current and voltage thresholds, and action potentials with longer durations and slower rising and falling rates. The incidence of spontaneous activity, recorded extracellularly from dorsal root fibers in vitro, was significantly higher (in relation to sham) after SNL but not after PR, and occurred in myelinated but not unmyelinated fibers from both L4 (9.1%) and L5 (16.7%) DRGs. We hypothesize that the changes in the electrophysiological properties of axotomized and intact DRG neurons after SNL are produced by a mechanism associated with Wallerian degeneration and that the hyperexcitability of intact neurons may contribute to SNL-induced hyperalgesia and allodynia.

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Estrogen Elicits Dorsal Root Ganglion Axon Sprouting via a Renin-Angiotensin System

Endocrinology ◽

10.1210/en.2008-0061 ◽

2008 ◽

Vol 149 (7) ◽

pp. 3452-3460 ◽

Cited By ~ 49

Author(s):

Anuradha Chakrabarty ◽

Audrey Blacklock ◽

Stanislav Svojanovsky ◽

Peter G. Smith

Keyword(s):

Estrogen Receptor ◽

Dorsal Root Ganglion ◽

Dorsal Root ◽

Drg Neurons ◽

Predisposing Factor ◽

Adult Rats ◽

Axon Sprouting ◽

Pain Syndromes

Many painful conditions occur more frequently in women, and estrogen is a predisposing factor. Estrogen may contribute to some pain syndromes by enhancing axon outgrowth by sensory dorsal root ganglion (DRG) neurons. The objective of the present study was to define mechanisms by which estrogen elicits axon sprouting. The estrogen receptor-α agonist propyl pyrazole triol induced neurite outgrowth from cultured neonatal DRG neurons, whereas the estrogen receptor-β agonist diarylpropionitrile was ineffective. 17β-Estradiol (E2) elicited sprouting from peripherin-positive unmyelinated neurons, but not larger NF200-positive myelinated neurons. Microarray analysis showed that E2 up-regulates angiotensin II (ANGII) receptor type 2 (AT2) mRNA in vitro, and studies in adult rats confirmed increased DRG mRNA and protein in vivo. AT2 plays a central role in E2-induced axon sprouting because AT2 blockade by PD123,319 eliminated estrogen-mediated sprouting in vitro. We assessed whether AT2 may be responding to locally synthesized ANGII. DRG from adult rats expressed mRNA for renin, angiotensinogen, and angiotensin converting enzyme (ACE), and protein products were present and occasionally colocalized within neurons and other DRG cells. We determined if locally synthesized ANGII plays a role in estrogen-mediated sprouting by blocking its formation using the ACE inhibitor enalapril. ACE inhibition prevented estrogen-induced neuritogenesis. These findings support the hypothesis that estrogen promotes DRG nociceptor axon sprouting by up-regulating the AT2 receptor, and that locally synthesized ANGII can induce axon formation. Therefore, estrogen may contribute to some pain syndromes by enhancing the pro-neuritogenic effects of AT2 activation by ANGII.

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Intra-articular AAV-PHP.S mediated chemogenetic targeting of knee-innervating dorsal root ganglion neurons alleviates inflammatory pain in mice

10.1101/2020.02.08.939066 ◽

2020 ◽

Author(s):

Sampurna Chakrabarti ◽

Luke A. Pattison ◽

Balint Doleschall ◽

Rebecca H. Rickman ◽

Helen Blake ◽

...

Keyword(s):

Dorsal Root Ganglion ◽

Knee Joint ◽

Joint Pain ◽

Dorsal Root ◽

Dorsal Root Ganglion Neurons ◽

Drg Neurons ◽

Neuron Excitability ◽

Ganglion Neurons

AbstractObjectiveJoint pain is the major clinical symptom of arthritis that affects millions of people. Controlling the excitability of knee-innervating dorsal root ganglion (DRG) neurons (knee neurons) could potentially provide pain relief. Therefore, our objective was to evaluate whether the newly engineered adeno-associated virus (AAV) serotype, AAV-PHP.S, can deliver functional artificial receptors to control knee neuron excitability following intra-articular knee injection.MethodsAAV-PHP.S virus packaged with dTomato fluorescent protein and either excitatory (Gq) or inhibitory (Gi) designer receptors activated by designer drugs (DREADDs) was injected into the knee joint of adult mice. Labelling of DRG neurons by AAV-PHP.S from the knee was evaluated using immunohistochemistry. Functionality of Gq- and Gi-DREADDs was evaluated using whole-cell patch clamp electrophysiology on acutely cultured DRG neurons. Pain behavior in mice was assessed using a digging assay, dynamic weight bearing and rotarod, before and after intra-peritoneal administration of the DREADD activator, Compound 21.ResultsWe show that AAV-PHP.S can deliver functional genes into the DRG neurons when injected into the knee joint in a similar manner to the well-established retrograde tracer, fast blue. Short-term activation of AAV-PHP.S delivered Gq-DREADD increases excitability of knee neurons in vitro, without inducing overt pain in mice when activated in vivo. By contrast, in vivo Gi-DREADD activation alleviated complete Freund’s adjuvant mediated knee inflammation-induced deficits in digging behavior, with a concomitant decrease in knee neuron excitability observed in vitro.ConclusionsWe describe an AAV-mediated chemogenetic approach to specifically control joint pain, which may be utilized in translational arthritic pain research.

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Cdk5 Phosphorylation of STAT3 in Dorsal Root Ganglion Neurons Is Involved in Promoting Axonal Regeneration After Peripheral Nerve Injury

International Neurourology Journal ◽

10.5213/inj.2040158.080 ◽

2020 ◽

Vol 24 (Suppl 1) ◽

pp. S19-27

Author(s):

Jinyeon Hwang ◽

Uk Namgung

Keyword(s):

Sciatic Nerve ◽

Dorsal Root Ganglion ◽

Peripheral Nerve ◽

Nerve Injury ◽

Axonal Regeneration ◽

Dorsal Root ◽

Immunofluorescence Staining ◽

Drg Neurons ◽

Ganglion Neurons

Purpose: The goal of this study is to investigate the role of cyclin-dependent kinase 5 (Cdk5) in axonal regeneration in dorsal root ganglion (DRG) neurons after peripheral nerve injury.Methods: Crush injury was given on the sciatic nerve in rats. The DRG tissues were prepared 1, 3, and 7 days after injury and used for western blotting and immunofluorescence staining experiments. Primary DRG neurons were prepared and treated with Cdk5 inhibitor roscovitine or used for transfections with plasmid constructs. After immunofluorescence staining, neurite length of DRG neurons was analyzed and compared among experimental groups. In addition, roscovitine was injected into the DRG <i>in vivo</i>, and the sciatic nerve after injury was prepared and used for immunofluorescence staining to analyze axonal regeneration in nerve sections.Results: Levels of Cdk5 and p25 were increased in DRG neurons after sciatic nerve injury (SNI). Levels of S727-p-STAT3, but not Y705-p-STAT3, were increased in the DRG. Immunofluorescence staining revealed that Cdk5 and STAT3 proteins were mostly colocalized in DRG neurons and Y705-p-STAT3 signals were localized within the nucleus area of DRG neurons. A blockade of Cdk5 activity by roscovitine or by transfection with dominant negative Cdk5 (dn-Cdk5) and nonphosphorylatable forms of STAT3 (S727A or Y705F) resulted in significant reductions of the neurite outgrowth of cultured DRG neurons. <i>In vivo</i> administration of roscovitine into the DRG markedly attenuated distal elongation of regenerating axons in the sciatic nerve after injury.Conclusions: Our study demonstrated that Cdk5 activity induced from DRG neurons after SNI increased phosphorylation of STAT3. The activation of Cdk5-STAT3 pathway may be involved in promoting axonal regeneration in the peripheral nerve after injury.

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Inhibition of Hyperpolarization-Activated Cation Current in Medium-Sized DRG Neurons Contributed to the Antiallodynic Effect of Methylcobalamin in the Rat of a Chronic Compression of the DRG

Neural Plasticity ◽

10.1155/2015/197392 ◽

2015 ◽

Vol 2015 ◽

pp. 1-13 ◽

Cited By ~ 4

Author(s):

Ming Zhang ◽

Wenjuan Han ◽

Jianyong Zheng ◽

Fancheng Meng ◽

Xiying Jiao ◽

...

Keyword(s):

Dorsal Root Ganglion ◽

Sensory Neurons ◽

Analgesic Effect ◽

Dorsal Root ◽

High Dose ◽

Dependent Manner ◽

Drg Neurons ◽

Continuous Application

Recently several lines of evidence demonstrated that methylcobalamin (MeCbl) might have potential analgesic effect in experimental and clinical studies. However, it was reported that MeCbl had no effect on treating lumbar spinal stenosis induced pain. Thus, the effects of short-term and long-term administration of MeCbl were examined in the chronic compression of dorsal root ganglion (CCD) model. We found that mechanical allodynia was significantly inhibited by a continuous application of high dose and a single treatment of a super high dose of MeCbl. Little is known about mechanisms underlying the analgesia of MeCbl. We examined the effect of MeCbl on the spontaneous activity (SA), the excitability, and hyperpolarization-activated nonselective cation ion current in compressed medium-sized dorsal root ganglion (DRG) neurons using extracellular single fiber recordingin vivoand whole-cell patch clampin vitro. We found that MeCbl significantly inhibited the SA of A-type sensory neurons in a dose-dependent manner and inhibited the excitability of medium-sized DRG neurons. In addition, MeCbl also decreasedIhcurrent density in injured medium-sized DRG neurons. Our results proved that MeCbl might exert an analgesic effect through the inhibitionIhcurrent and then might inhibit the hyperexcitability of primary sensory neurons under neuropathic pain state.

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Hyperalgesia and Neural Excitability Following Injuries to Central and Peripheral Branches of Axons and Somata of Dorsal Root Ganglion Neurons

Journal of Neurophysiology ◽

10.1152/jn.00802.2002 ◽

2003 ◽

Vol 89 (4) ◽

pp. 2185-2193 ◽

Cited By ~ 58

Author(s):

Xue-Jun Song ◽

Carlos Vizcarra ◽

Dong-Sheng Xu ◽

Ronald L. Rupert ◽

Zheng-Nan Wong

Keyword(s):

Nmda Receptor ◽

Dorsal Root Ganglion ◽

Dorsal Root ◽

Thermal Hyperalgesia ◽

Dorsal Root Ganglion Neurons ◽

Membrane Excitability ◽

Drg Neurons ◽

Mk 801 ◽

Ganglion Neurons

We examined thermal hyperalgesia, excitability of dorsal root ganglion (DRG) neurons, and antinociceptive effects of N-methyl-d-aspartate (NMDA) receptor antagonists in rats with injury to different regions of DRG neurons. The central or peripheral branches of axons of DRG neurons were injured by partial dorsal rhizotomy (PDR) and chronic constriction injury of sciatic nerve (CCI), respectively, or the somata injured by chronic compression of DRG (CCD). Thermal hyperalgesia was evidenced by significantly shortened latencies of foot withdrawal to radiant heat stimulation of the plantar surface. Intracellular recordings were obtained in vitro from L4 and/or L5 ganglia. There are four principle findings: 1) PDR as well as CCD and CCI induced thermal hyperalgesia; 2) PDR produced significantly less severe and shorter duration hyperalgesia than CCD and CCI; 3) intrathecal administration of NMDA receptor antagonistsd-2-amino-5-phosphonovaleric acid (APV) and dizocilpine maleate (MK-801) inhibited thermal hyperalgesia in PDR, CCD, and CCI rats. Pretreatment of APV and MK-801 delayed the emergence of hyperalgesia for 48–72 h, while posttreatment inhibited hyperalgesia for 24–36 h; and 4) CCD and CCI increased excitability of DRG neurons as judged by the significantly lowered threshold currents and action potential voltage thresholds and increased incidence of repetitive discharges. However, PDR did not alter the excitability of DRG neurons. These findings indicate that injury to the dorsal root, compared with injury to the peripheral nerve or DRG somata has different effects on the development of hyperalgesia. These contributions involve different changes in DRG membrane excitability, but each involves pathways (presumably in the spinal cord) that depend on NMDA receptors.

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Upregulation of EMMPRIN (OX47) in Rat Dorsal Root Ganglion Contributes to the Development of Mechanical Allodynia after Nerve Injury

Neural Plasticity ◽

10.1155/2015/249756 ◽

2015 ◽

Vol 2015 ◽

pp. 1-9 ◽

Cited By ~ 3

Author(s):

Qun Wang ◽

Yanyuan Sun ◽

Yingna Ren ◽

Yandong Gao ◽

Li Tian ◽

...

Keyword(s):

Extracellular Matrix ◽

Neuropathic Pain ◽

Dorsal Root Ganglion ◽

Nerve Injury ◽

Messenger Rna ◽

Dorsal Root ◽

Mechanical Allodynia ◽

Cellular Localization ◽

Altered Expression

Matrix metalloproteinases (MMPs) are widely implicated in inflammation and tissue remodeling associated with various neurodegenerative diseases and play an important role in nociception and allodynia. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) plays a key regulatory role for MMP activities. However, the role of EMMPRIN in the development of neuropathic pain is not clear. Western blotting, real-time quantitative RT-PCR (qRT-PCR), and immunofluorescence were performed to determine the changes of messenger RNA and protein of EMMPRIN/OX47 and their cellular localization in the rat dorsal root ganglion (DRG) after nerve injury. Paw withdrawal threshold test was examined to evaluate the pain behavior in spinal nerve ligation (SNL) model. The lentivirus containing OX47 shRNA was injected into the DRG one day before SNL. The expression level of both mRNA and protein of OX47 was markedly upregulated in ipsilateral DRG after SNL. OX47 was mainly expressed in the extracellular matrix of DRG. Administration of shRNA targeted against OX47 in vivo remarkably attenuated mechanical allodynia induced by SNL. In conclusion, peripheral nerve injury induced upregulation of OX47 in the extracellular matrix of DRG. RNA interference against OX47 significantly suppressed the expression of OX47 mRNA and the development of mechanical allodynia. The altered expression of OX47 may contribute to the development of neuropathic pain after nerve injury.

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An integrated cell isolation and purification method for rat dorsal root ganglion neurons

Journal of International Medical Research ◽

10.1177/0300060519855585 ◽

2019 ◽

Vol 47 (7) ◽

pp. 3253-3260

Author(s):

Huaishuang Shen ◽

Minfeng Gan ◽

Huilin Yang ◽

Jun Zou

Keyword(s):

Neuropathic Pain ◽

Dorsal Root Ganglion ◽

Primary Culture ◽

Dorsal Root ◽

Cell Isolation ◽

Dorsal Root Ganglion Neurons ◽

Drg Neurons ◽

Purification Method ◽

Isolation And Purification ◽

Ganglion Neurons

Objective Neurobiology studies are increasingly focused on the dorsal root ganglion (DRG), which plays an important role in neuropathic pain. Existing DRG neuron primary culture methods have considerable limitations, including challenging cell isolation and poor cell yield, which cause difficulty in signaling pathway studies. The present study aimed to establish an integrated primary culture method for DRG neurons. Methods DRGs were obtained from fetal rats by microdissection, and then dissociated with trypsin. The dissociated neurons were treated with 5-fluorouracil to promote growth of neurons from the isolated cells. Then, reverse transcription polymerase chain reaction and immunofluorescence assays were used to identify and purify DRG neurons. Results Isolated DRGs were successfully dissociated and showed robust growth as individual DRG neurons in neurobasal medium. Both mRNA and protein assays confirmed that DRG neurons expressed neurofilament-200 and neuron-specific enolase. Conclusions Highly purified, stable DRG neurons could be easily harvested and grown for extended periods by using this integrated cell isolation and purification method, which may help to elucidate the mechanisms underlying neuropathic pain.

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Microglial BDNF, PI3K, and p-ERK in the Spinal Cord Are Suppressed by Pulsed Radiofrequency on Dorsal Root Ganglion to Ease SNI-Induced Neuropathic Pain in Rats

Pain Research and Management ◽

10.1155/2019/5948686 ◽

2019 ◽

Vol 2019 ◽

pp. 1-15 ◽

Cited By ~ 3

Author(s):

Xueru Xu ◽

Shaoxiong Fu ◽

Xiaomei Shi ◽

Rongguo Liu

Keyword(s):

Spinal Cord ◽

Neuropathic Pain ◽

Dorsal Root Ganglion ◽

Nerve Injury ◽

Calcium Binding ◽

Dorsal Root ◽

Intrathecal Injection ◽

Pulsed Radiofrequency ◽

Erk Phosphorylation ◽

Extracellular Signal

Background. Pulsed radiofrequency (PRF) on the dorsal root ganglion (DRG) has been applied to alleviate neuropathic pain effectively, yet the mechanisms underlying pain reduction owing to this treatment are not clarified completely. The activated microglia, brain-derived neurotrophic factor (BDNF), phosphatidylinositol 3-kinase (PI3K), and phosphorylated extracellular signal-regulated kinase (p-ERK) in the spinal cord were demonstrated to be involved in developing neuropathic pain. Also, it has been just known that PRF on DRG inhibits the microglial activation in nerve injury rats. Here, we aim to investigate whether PRF treatment could regulate the levels of BDNF, PI3K, and p-ERK in the spinal cord of rats with spared nerve injury (SNI) via suppressing the spinal microglia activation to ease neuropathic pain. Methods. The rats with SNI were intrathecally treated with minocycline (specific microglia inhibitor) or same volume of dimethyl sulfoxide once daily, beginning from 1 h before nerve transection to 7 days. PRF was applied adjacent to the L4-L5 DRG of rats with SNI at 45 V for 6 min on the seventh postoperative day, whereas the free-PRF rats were treated without PRF. The withdrawal thresholds were studied, and the spinal levels of ionized calcium-binding adapter molecule 1 (Iba1), BDNF, PI3K, and p-ERK were calculated by western blot analysis, reverse transcription-polymerase chain reaction, and immunofluorescence. Results. The paw withdrawal mechanical threshold and paw withdrawal thermal latency decreased in the ipsilateral hind paws after SNI, and the spinal levels of Iba1, BDNF, PI3K, and p-ERK increased on day 21 after SNI compared with baseline (P<0.01). An intrathecal injection of minocycline led to the reversal of SNI-induced allodynia and increase in levels of Iba1, BDNF, PI3K, and p-ERK. Withdrawal thresholds recovered partially after a single PRF treatment for 14 days, and SNI-induced microglia hyperactivity, BDNF upregulation, and PI3K and ERK phosphorylation in the spinal cord reduced on D14 due to the PRF procedure. Conclusion. Microglial BDNF, PI3K, and p-ERK in the spinal cord are suppressed by the therapy of PRF on DRG to ease SNI-induced neuropathic pain in rats.

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