tetrodotoxin and Peripheral-Nervous-System-Diseases

Neuropathic pain, a persistent chronic pain resulting from damage to the central or peripheral nervous system, is a condition that severely affects the quality-of-life of millions of individuals worldwide. The treatment of neuropathic pain is still an unmet medical need; however, recent advances in our understanding of mechanisms underlying the perception and transmission of painful stimuli offer significant potential for improvement of therapies directed to neuropathic pain. Ectopic activity in damaged and dysfunctional sensory afferents is believed to have a role in the generation and maintenance of neuropathic pain. One of the mechanisms underlying this ectopic firing involves abnormal modulation of voltage-gated sodium channels (NaVs) in the soma and axonal membranes of dorsal root ganglion (DRG) sensory neurons. In fact, NaV blockers have been clinically validated as treatments for neuropathic pain. However, current drugs are weak, non-selective inhibitors of NaVs with dose-limiting CNS and cardiovascular side effects that prevent their use in long-term therapy. Selective NaV tetrodotoxin-resistant channels (NaV 1.8 and NaV 1.9) are expressed exclusively in nociceptive neurons in the DRGs where they play a key role in normal and/or pathological pain sensation, providing an opportunity for the development of novel peripheral analgesics with a better safety profile.

Topics: Anesthetics, Local; Ganglia, Spinal; Humans; Ion Channel Gating; Neurons, Afferent; Pain; Peripheral Nervous System Diseases; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin

Physiological and pharmacological evidence both have demonstrated a critical role for voltage-gated sodium channels (VGSCs) in many types of chronic pain syndromes because these channels play a fundamental role in the excitability of neurons in the central and peripheral nervous systems. Alterations in function of these channels appear to be intimately linked to hyperexcitability of neurons. Many types of pain appear to reflect neuronal hyperexcitability, and importantly, use-dependent sodium channel blockers are effective in the treatment of many types of chronic pain. This review focuses on the role of VGSCs in the hyperexcitability of sensory primary afferent neurons and their contribution to the inflammatory or neuropathic pain states. The discrete localization of the tetrodotoxin (TTX)-resistant channels, in particular NaV1.8, in the peripheral nerves may provide a novel opportunity for the development of a drug targeted at these channels to achieve efficacious pain relief with an acceptable safety profile.

Topics: Analgesics; Anesthetics, Local; Animals; Anticonvulsants; Humans; Hyperalgesia; Inflammation; Ion Channel Gating; Neurons, Afferent; Pain; Peripheral Nervous System Diseases; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin

Topics: Animals; Central Nervous System Diseases; Diagnosis, Differential; Fishes; Humans; Peripheral Nervous System Diseases; Prognosis; Respiration, Artificial; Respiratory Paralysis; Tetrodotoxin

Neuropathic pain is a significant public health challenge, yet the underlying mechanisms remain poorly understood. Painful small fiber neuropathy (SFN) may be caused by gain-of-function mutations in Nav1.8, a sodium channel subtype predominantly expressed in peripheral nociceptive neurons. However, it is not clear how Nav1.8 disease mutations induce sensory neuron hyperexcitability. Here we studied two mutations in Nav1.8 associated with hypersensitive sensory neurons: G1662S reported in painful SFN; and T790A, which underlies increased pain behaviors in the

Topics: Action Potentials; Animals; Disease Models, Animal; Gain of Function Mutation; Humans; Ion Channel Gating; Ion Transport; Male; Mice; Mice, Neurologic Mutants; Mice, Transgenic; Mutation, Missense; NAV1.8 Voltage-Gated Sodium Channel; Neuralgia; Nociception; Patch-Clamp Techniques; Peripheral Nervous System Diseases; Point Mutation; Rats; Rats, Sprague-Dawley; Recombinant Proteins; RNA Interference; Sensory Receptor Cells; Sodium; Tetrodotoxin

The over-expression of voltage-gated sodium channels (VGSCs) in dorsal root ganglion (DRG) neurons following peripheral nerve injury contributes to neuropathic pain by generation of the ectopic discharges of action potentials. However, mechanisms underlying the change in VGSCs' expression are poorly understood. Our previous work has demonstrated that the pro-inflammatory cytokine TNF-α up-regulates VGSCs. In the present work we tested if anti-inflammatory cytokine IL-10, which had been proven to be effective for treating neuropathic pain, had the opposite effect. Western blot and immunofluorescence results showed that IL-10 receptor was localized in DRG neurons. Recombinant rat IL-10 (200 pg/ml) not only reduced the densities of TTX-sensitive and Nav1.8 currents in control DRG neurons, but also reversed the increase of the sodium currents induced by rat recombinant TNF-α (100 pg/ml), as revealed by patch-clamp recordings. Consistent with the electrophysiological results, real-time PCR and western blot revealed that IL-10 (200 pg/ml) down-regulated VGSCs in both mRNA and protein levels and reversed the up-regulation of VGSCs by TNF-α. Moreover, repetitive intrathecal administration of rrIL-10 for 3 days (4 times per day) attenuated mechanical allodynia in L5 spinal nerve ligation model and profoundly inhibited the excitability of DRG neurons. These results suggested that the down-regulation of the sodium channels in DRG neurons might contribute to the therapeutic effect of IL-10 on neuropathic pain.

Topics: Animals; Cells, Cultured; Disease Models, Animal; Down-Regulation; Ganglia, Spinal; Interleukin-10; Ligation; Male; Membrane Potentials; Nerve Tissue Proteins; Neurons; Pain Measurement; Peripheral Nervous System Diseases; Rats; Rats, Sprague-Dawley; Receptors, Interleukin-10; Sodium Channel Blockers; Tetrodotoxin; Tumor Necrosis Factor-alpha; Voltage-Gated Sodium Channels

Peripheral nerve hyperexcitability (PNH) is one of the distal peripheral neuropathy phenotypes often present in patients affected by type 2 diabetes mellitus (T2DM). Through in vivo and ex vivo electrophysiological recordings in db/db mice, a model of T2DM, we observed that, in addition to reduced nerve conduction velocity, db/db mice also develop PNH. By using pharmacological inhibitors, we demonstrated that the PNH is mediated by the decreased activity of K(v)1-channels. In agreement with these data, we observed that the diabetic condition led to a reduced presence of the K(v)1.2-subunits in juxtaparanodal regions of peripheral nerves in db/db mice and in nerve biopsies from T2DM patients. Together, these observations indicate that the T2DM condition leads to potassium channel-mediated PNH, thus identifying them as a potential drug target to treat some of the DPN related symptoms.

Topics: Action Potentials; Age Factors; Animals; Blood Glucose; Body Weight; Diabetes Mellitus, Type 2; Disease Models, Animal; Electric Stimulation; Humans; Kv1.2 Potassium Channel; Male; Mice; Mice, Mutant Strains; Mutation; Neural Conduction; Peripheral Nerves; Peripheral Nervous System Diseases; Potassium Channel Blockers; Protein Subunits; Ranvier's Nodes; Receptors, Leptin; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin

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.. 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.. (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.. Thiamine can reduce hyperexcitability and lessen alterations of Na currents in injured DRG neurons, in addition to suppressing thermal hyperalgesia.

Topics: Animals; Behavior, Animal; Cell Size; Electrophysiology; Ganglia, Spinal; Hot Temperature; Hyperalgesia; Male; Nerve Compression Syndromes; Neurons; Patch-Clamp Techniques; Peripheral Nervous System Diseases; Rats; Rats, Sprague-Dawley; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin; Thiamine; Vitamins

Satellite glial cells in the dorsal root ganglion (DRG), like the better-studied glia cells in the spinal cord, react to peripheral nerve injury or inflammation by activation, proliferation, and release of messengers that contribute importantly to pathological pain. It is not known how information about nerve injury or peripheral inflammation is conveyed to the satellite glial cells. Abnormal spontaneous activity of sensory neurons, observed in the very early phase of many pain models, is one plausible mechanism by which injured sensory neurons could activate neighboring satellite glial cells. We tested effects of locally inhibiting sensory neuron activity with sodium channel blockers on satellite glial cell activation in a rat spinal nerve ligation (SNL) model. SNL caused extensive satellite glial cell activation (as defined by glial fibrillary acidic protein [GFAP] immunoreactivity) which peaked on day 1 and was still observed on day 10. Perfusion of the axotomized DRG with the Na channel blocker tetrodotoxin (TTX) significantly reduced this activation at all time points. Similar findings were made with a more distal injury (spared nerve injury model), using a different sodium channel blocker (bupivacaine depot) at the injury site. Local DRG perfusion with TTX also reduced levels of nerve growth factor (NGF) in the SNL model on day 3 (when activated glia are an important source of NGF), without affecting the initial drop of NGF on day 1 (which has been attributed to loss of transport from target tissues). Local perfusion in the SNL model also significantly reduced microglia activation (OX-42 immunoreactivity) on day 3 and astrocyte activation (GFAP immunoreactivity) on day 10 in the corresponding dorsal spinal cord. The results indicate that early spontaneous activity in injured sensory neurons may play important roles in glia activation and pathological pain.

Topics: Animals; Biomarkers; Bupivacaine; CD11b Antigen; Disease Models, Animal; Ganglia, Spinal; Glial Fibrillary Acidic Protein; Gliosis; Ligation; Male; Microglia; Nerve Growth Factor; Neuralgia; Peripheral Nerve Injuries; Peripheral Nerves; Peripheral Nervous System Diseases; Rats; Rats, Sprague-Dawley; Satellite Cells, Perineuronal; Sensory Receptor Cells; Sodium Channel Blockers; Tetrodotoxin; Time Factors

Some chronic pain conditions are maintained or enhanced by sympathetic activity. In animal models of pathological pain, abnormal sprouting of sympathetic fibers around large- and medium-sized sensory neurons is observed in dorsal root ganglia (DRGs). Large- and medium-sized cells are also more likely to be spontaneously active, suggesting that sprouting may be related to neuron activity. We previously showed that sprouting could be reduced by systemic or locally applied lidocaine. In the complete sciatic nerve transection model in rats, spontaneous activity initially originates in the injury site; later, the DRGs become the major source of spontaneous activity. In this study, spontaneous activity reaching the DRG soma was reduced by early nerve blockade (local perfusion of the transected nerve with TTX for the 1st 7 days after injury). This significantly reduced sympathetic sprouting. Conversely, increasing spontaneous activity by local nerve perfusion with K(+) channel blockers increased sprouting. The hyperexcitability and spontaneous activity of DRG neurons observed in this model were also significantly reduced by early nerve blockade. These effects of early nerve blockade on sprouting, excitability, and spontaneous activity were all observed 4-5 wk after the end of early nerve blockade, indicating that the early period of spontaneous activity in the injured nerve is critical for establishing the more long-lasting pathologies observed in the DRG. Individual spontaneously active neurons, labeled with fluorescent dye, were five to six times more likely than quiescent cells to be co-localized with sympathetic fibers, suggesting a highly localized correlation of activity and sprouting.

Topics: Action Potentials; Animals; Autonomic Nerve Block; Axotomy; Disease Models, Animal; Female; Fluorescent Dyes; Ganglia, Spinal; Growth Cones; Nerve Regeneration; Neural Conduction; Neuralgia; Neurons, Afferent; Organ Culture Techniques; Peripheral Nerve Injuries; Peripheral Nerves; Peripheral Nervous System Diseases; Potassium Channel Blockers; Rats; Rats, Sprague-Dawley; Sciatic Neuropathy; Sodium Channel Blockers; Sympathetic Fibers, Postganglionic; Tetrodotoxin; Time Factors

In this work we hypothesized that denervation in flexor digitorum brevis (FDB) muscle from ageing mice is more extensive than predicted by standard functional and structural assays used in the past. In addition, we asked whether denervation is a fully or partially developed process. Despite the reported alteration in skeletal muscle innervation, the quantification of the extension and magnitude of denervation in ageing rodents has remained elusive. To address these two questions we utilized a combination of electrophysiological and immunohistochemical assays directed to detecting the expression of tetrodotoxin (TTX)-resistant sodium channels (Na(v)1.5) in FDB muscles from young-adult and senescent mice. Sodium current density measured with the macropatch cell-attached technique did not show significant differences between FDB fibres from young and old mice. The TTX dose-response curve, using the whole cell voltage-clamp technique, showed three populations of fibres in senescent mice, one similar to fibres from young mice (TTX sensitive), another one similar to fibres from experimentally denervated muscle (TTX resistant), and a third group intermediate between these two. Partially and fully denervated fibres added up to approximately 50% of the total number of fibres tested, a number that concurs with the percentage of fibres positive for the Na(v)1.5 channel by specific immunostaining.

Topics: Aging; Anesthetics, Local; Animals; Membrane Potentials; Mice; Mice, Inbred Strains; Muscle Fibers, Skeletal; Muscle Proteins; Muscle, Skeletal; NAV1.5 Voltage-Gated Sodium Channel; Patch-Clamp Techniques; Peripheral Nervous System Diseases; Sodium Channels; Tetrodotoxin

Neuropathic pain is a debilitating chronic syndrome that often arises from injuries to peripheral nerves. Such pain has been hypothesized to be the result of an aberrant expression and function of sodium channels at the site of injury. Here, we show that intrathecal administration of specific antisense oligodeoxynucleotides (ODN) to the peripheral tetrodotoxin (TTX)-resistant sodium channel, NaV1.8, resulted in a time-dependent uptake of the ODN by dorsal root ganglion (DRG) neurons, a selective "knock-down" of the expression of NaV1.8, and a reduction in the slow-inactivating, TTX-resistant sodium current in the DRG cells. The ODN treatment also reversed neuropathic pain induced by spinal nerve injury, without affecting non-noxious sensation or response to acute pain. These data provide direct evidence linking NaV1.8 to neuropathic pain. As NaV1.8 expression is restricted to sensory neurons, this channel offers a highly specific and effective molecular target for the treatment of neuropathic pain.

Topics: Anesthetics, Local; Animals; Ganglia, Spinal; Male; Oligodeoxyribonucleotides, Antisense; Pain; Pain Threshold; Peripheral Nervous System Diseases; Rats; Rats, Sprague-Dawley; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin

Voltage-dependent Na-currents were studied, using whole cell voltage clamp, in acutely dissociated, large (mostly Abeta-fiber type) cutaneous afferent dorsal root ganglia neurons (L(4) and L(5)) from the adult rat. Cells were dissociated 14-17 days after axotomy. Control and axotomized neurons were identified via the retrograde marker hydroxy-stilbamide (fluorogold) which was injected into the lateral and plantar region of the skin of the foot and were studied using whole cell patch clamp techniques within 12-20 h of dissociation and plating. Cells were dissociated 14-17 days after injury. Both control and axotomized neurons displayed complex Na-currents composed of components with distinct kinetic and pharmacological properties. The large (48-50 microm diameter) control cutaneous afferent neurons, many of which likely give rise to myelinated Abeta-fibers, exhibited Na-currents with both slow and fast inactivating kinetics. The fast inactivating current in large cutaneous afferent dorsal root ganglion neurons was tetrodotoxin-sensitive and recovered from inactivation approximately four-fold faster at -60 mV (P<0.001) and approximately six-fold faster at -70 mV (P<0.001) than the tetrodotoxin-sensitive current in small (<30 microm diameter) neurons. Further, while the tetrodotoxin-sensitive currents in smaller dorsal root ganglion neurons (mainly C-fiber type) reprime approximately four-fold faster following peripheral axotomy, repriming of the fast inactivating current in larger cutaneous afferent neurons was not significantly altered following axotomy. However, while 77% of control large neurons were observed to express the slower inactivating, tetrodotoxin-resistant current, only 45% of these large neurons did after axotomy. These results indicate that large adult cutaneous afferent dorsal root ganglion neurons (Abeta-type) express tetrodotoxin-sensitive Na-currents, which have much faster repriming than Na-currents in small (C-type) neurons, both before, and after axotomy. Like small neurons, the majority of large neurons downregulate the tetrodotoxin-resistant current following sciatic nerve section.

Topics: Animals; Axotomy; Cell Size; Down-Regulation; Female; Fluorescent Dyes; Ganglia, Spinal; Membrane Potentials; Nerve Fibers; Nerve Fibers, Myelinated; Neural Conduction; Neuralgia; Neurons, Afferent; Patch-Clamp Techniques; Peripheral Nervous System Diseases; Rats; Rats, Wistar; Recovery of Function; Sciatic Nerve; Sodium Channels; Stilbamidines; Tetrodotoxin

The tetrodotoxin-resistant voltage-gated sodium channel Nav 1.8 is expressed only in nociceptive sensory neurons. This channel has been proposed to contribute significantly to the sensitization of primary sensory neurons after injury. We have studied the nociceptive behaviours of mice carrying a null mutation in the Nav 1.8 gene (Nav 1.8 -/-) in models of peripheral inflammation as well as a model of neuropathic pain. The results from the present studies reveal that Nav 1.8 is a necessary mediator of NGF-induced thermal hyperalgesia but is not essential for PGE2-evoked hypersensitivity. Neuropathic pain behaviours were unchanged in Nav 1.8 -/- mice indicating that this channel is not involved in the alteration of sensory thresholds following peripheral nerve injury.

Topics: Animals; Dinoprostone; Disease Models, Animal; Female; Ganglia, Spinal; Hyperalgesia; Ligation; Male; Mice; Mice, Knockout; Nerve Growth Factor; Neuralgia; Neurons, Afferent; Nociceptors; Peripheral Nervous System Diseases; Sciatic Nerve; Sodium Channels; Tetrodotoxin

A neuromuscular blocking factor has been described in the serum of patients with Miller-Fisher syndrome (MFS). We here examined the effect of immunoglobulins (Ig) on neuromuscular transmission in mice recording quantal endplate currents by means of a perfused macro-patch-clamp electrode. Ig and IgM- and IgG-fractions from an anti-GQ1b-positive patient with typical MFS were highly purified. After application of MFS-IgG, quantal release decreased 1000-fold within 2 min. Returning to control solution the average release came back to the baseline level within 4 min. In contrast, control-IgG and MFS-IgM did not cause any blocking effect. The very fast and fully reversible presynaptic blockade of release caused by the highly purified IgG-fraction may be one factor producing muscle weakness in MFS.

Topics: Animals; Demyelinating Diseases; Diaphragm; Humans; Immunoglobulin G; Immunoglobulin M; In Vitro Techniques; Mice; Mice, Inbred BALB C; Motor Endplate; Neuromuscular Junction; Peripheral Nervous System Diseases; Syndrome; Tetrodotoxin