tetrodotoxin and Demyelinating-Diseases

Neurological sequelae of mild traumatic brain injury are associated with the damage to white matter myelinated axons. In vitro models of axonal injury suggest that the progression to pathological ruin is initiated by the mechanical damage to tetrodotoxin-sensitive voltage-gated sodium channels that breaches the ion balance through alteration in kinetic properties of these channels. In myelinated axons, sodium channels are concentrated at nodes of Ranvier, making these sites vulnerable to mechanical injury. Nodal damage can also be inflicted by injury-induced partial demyelination of paranode/juxtaparanode compartments that flank the nodes and contain high density of voltage-gated potassium channels. Demyelination-induced potassium deregulation can further aggravate axonal damage; however, the role of paranode/juxtaparanode demyelination in immediate impairment of axonal function, and its contribution to the development of axonal depolarization remain elusive. A biophysically realistic computational model of myelinated axon that incorporates ion exchange mechanisms and nodal/paranodal/juxtaparanodal organization was developed and used to study the impact of injury-induced demyelination on axonal signal transmission. Injured axons showed alterations in signal propagation that were consistent with the experimental findings and with the notion of reduced axonal excitability immediately post trauma. Injury-induced demyelination strongly modulated the rate of axonal depolarization, suggesting that trauma-induced damage to paranode myelin can affect axonal transition to degradation. Results of these studies clarify the contribution of paranode demyelination to immediate post trauma alterations in axonal function and suggest that partial paranode demyelination should be considered as another "injury parameter" that is likely to determine the stability of axonal function.

Topics: Action Potentials; Animals; Axons; Demyelinating Diseases; Humans; Ion Channels; Models, Neurological; Myelin Sheath; Ranvier's Nodes; Sodium Channel Blockers; Tetrodotoxin; Time Factors

Excessive nitric oxide formation may contribute to the pathology occurring in diseases affecting central white matter, such as multiple sclerosis. The rat isolated optic nerve preparation was used to investigate the potential toxicity of the molecule towards such tissue. The nerves were exposed to a range of concentrations of different classes of nitric oxide donor for up to 23 h, with or without a subsequent period of recovery, and the damage assessed by quantitative histological methods. Degeneration of axons and macroglia occurred in a time- and concentration-dependent manner, the order of susceptibility being: axons>oligodendrocytes>astrocytes. Use of NONOate donors differing in half-life indicated that nitric oxide delivered in an enduring manner at relatively low concentration was more toxic than the same amount supplied rapidly at high concentration. The mechanism by which nitric oxide affects axons was studied using a donor [3-(n-propylamino)propylamine/NO adduct, PAPA/NO] with an intermediate half-life that produced selective axonopathy after a 2-h exposure (plus 2 h recovery). Axon damage was abolished if, during the exposure, Na(+) or Ca(2+) was removed from the bathing medium or the sodium channel inhibitors tetrodotoxin or BW619C89 (sipatrigine) were added. In electrophysiological experiments, the donor elicited a biphasic depolarisation. The second, larger component (occurring after 7-10 min) was associated with a block of nerve conduction and could be inhibited by tetrodotoxin. Coincident with the secondary depolarisation was a reduction in ATP levels by about 50%, an effect that was also inhibited by tetrodotoxin. It is concluded that nitric oxide, in submicromolar concentrations, can kill axons and macroglia in white matter. The findings lend support to the hypothesis that nitric oxide may be of importance to white matter pathologies, particularly those in which inducible nitric oxide synthase is expressed. The axonopathy, at least when elicited over relatively short time intervals, is likely to be caused by metabolic inhibition. As in anoxia and anoxia/aglycaemia, nitric oxide-induced destruction of axons is likely to be caused by the Ca(2+) overload that follows a reduction in ATP levels in the face of continued influx of Na(+) through voltage-dependent channels.

Topics: Adenosine Triphosphate; Animals; Axons; Calcium; Central Nervous System; Demyelinating Diseases; Disease Models, Animal; Dose-Response Relationship, Drug; Membrane Potentials; Nerve Degeneration; Nerve Fibers, Myelinated; Neuroglia; Neurotoxins; Nitric Oxide; Nitric Oxide Donors; Optic Nerve; Organ Culture Techniques; Rats; Rats, Wistar; Sodium; Tetrodotoxin

Contactin (also known as F3, F11) is a surface glycoprotein that has significant homology with the beta2 subunit of voltage-gated Na(+) channels. Contactin and Na(+) channels can be reciprocally coimmunoprecipitated from brain homogenates, indicating association within a complex. Cells cotransfected with Na(+) channel Na(v)1.2alpha and beta1 subunits and contactin have threefold to fourfold higher peak Na(+) currents than cells with Na(v)1.2alpha alone, Na(v)1.2/beta1, Na(v)1.2/contactin, or Na(v)1.2/beta1/beta2. These cells also have a correspondingly higher saxitoxin binding, suggesting an increased Na(+) channel surface membrane density. Coimmunoprecipitation of different subunits from cell lines shows that contactin interacts specifically with the beta1 subunit. In the PNS, immunocytochemical studies show a transient colocalization of contactin and Na(+) channels at new nodes of Ranvier forming during remyelination. In the CNS, there is a particularly high level of colocalization of Na(+) channels and contactin at nodes both during development and in the adult. Contactin may thus significantly influence the functional expression and distribution of Na(+) channels in neurons.

Topics: Animals; Axons; Binding, Competitive; Brain Chemistry; Cell Adhesion Molecules, Neuronal; Cell Line; Cell Membrane; CHO Cells; Contactins; Cricetinae; Demyelinating Diseases; Female; Gene Expression; Lysophosphatidylcholines; NAV1.2 Voltage-Gated Sodium Channel; Nerve Tissue Proteins; Patch-Clamp Techniques; Precipitin Tests; Protein Subunits; Ranvier's Nodes; Rats; Saxitoxin; Sciatic Nerve; Sodium; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin; Transfection

Ectopic action potentials can arise at regions of axonal demyelination, and are believed to contribute to a range of symptoms in patients with demyelinating conditions such as multiple sclerosis. The mechanism(s) by which the ectopic impulses are generated are uncertain. We have previously shown that such ectopic activity can result from inward potassium currents. Paradoxically, the potassium channel blocking agent 4-aminopyridine (4AP) can sometimes cause ectopic activity in demyelinating lesions. To study this phenomenon we have made intra-axonal recordings of ectopic activity in demyelinated axons, both in the presence and absence of 5 mM 4AP. 4AP promoted a pattern of firing which was observed, albeit less frequently, in demyelinated axons in the absence of this drug, namely trains of single impulses, or trains of short, high-frequency bursts of impulses. When recorded close to the demyelinated lesion, the impulses were generated by an underlying, almost sinusoidal oscillation of the membrane potential. This oscillation was abolished by the sodium channel blocking agent tetrodotoxin (0.1-1 microM). We conclude that the ectopic spiking induced by 4AP is generated by membrane potential oscillations associated with the site of demyelination. The sodium-dependent current underlying these oscillations, together with the prolonged inward potassium currents which we have previously described, may contribute to the generation of ectopic discharges in a range of disorders of myelinated axons.

Topics: 4-Aminopyridine; Action Potentials; Animals; Axons; Demyelinating Diseases; Electrophysiology; Male; Potassium Channel Blockers; Rats; Rats, Sprague-Dawley; Rats, Wistar; Reaction Time; Sciatic Nerve; Sodium Channels; Spinal Cord; Tetrodotoxin

To investigate whether local anesthetic neurotoxicity results from sodium channel blockade, we compared the effects of intrathecally administered lidocaine, bupivacaine, and tetrodotoxin (TTX), the latter a highly selective sodium channel blocker, on sensory function and spinal cord morphology in a rat model. First, to determine relative anesthetic potency, 25 rats implanted with intrathecal catheters were subjected to infusions of lidocaine (n = 8), bupivacaine (n = 8), or TTX (n = 9). The three drugs produced parallel dose-effect curves that differed significantly from one another: the EC50 values for lidocaine, bupivacaine, and TTX were 28.2 mM (0.66%), 6.6 mM (0.19%), and 462 nM, respectively. Twenty-five additional rats were then given intrathecal lidocaine (n = 8), bupivacaine (n = 8), or TTX (n = 9) at concentrations 10 times the calculated EC50 for sensory block. Lidocaine and bupivacaine induced persistent sensory impairment, whereas TTX did not. Finally, 28 rats were given either intrathecal bupivacaine (n = 10) or TTX (n = 9) at 10 times the EC50, or normal saline (n = 9). Significant sensory impairment again occurred after infusion of bupivacaine, but not after infusion of TTX or saline. Neuropathologic evaluation revealed moderate to severe nerve root injury in bupivacaine-treated animals; histologic changes in TTX- and saline-treated animals were minimal, similar, and restricted to the area adjacent to the catheter. These results indicate that local anesthetic neurotoxicity does not result from blockade of the sodium channel, and suggest that development of a safer anesthetic is a realistic goal.

Topics: Animals; Bupivacaine; Demyelinating Diseases; Dose-Response Relationship, Drug; Injections, Spinal; Ion Channel Gating; Lidocaine; Male; Nerve Block; Nerve Degeneration; Rats; Rats, Sprague-Dawley; Reaction Time; Sensation Disorders; Sodium Channel Blockers; Sodium Channels; Spinal Cord; Spinal Nerve Roots; 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

The distribution of ionic channels is thought to play an important role in the recovery of function following demyelination. Rat sciatic nerves were demyelinated by lysolecithin and single fibers were examined with patch clamp techniques. Voltage-dependent sodium currents were measured in both internodal and nodal regions. The results suggested that while there is a sharp gradient in channel density at the node of Ranvier, the total number of internodal channels far exceeds the total number of nodal channels. The nodal channels do not diffuse freely following demyelination and the internodal channels may thus serve important functions as conduction is restored.

Topics: Animals; Demyelinating Diseases; Membrane Potentials; Nerve Fibers, Myelinated; Ranvier's Nodes; Rats; Rats, Inbred Lew; Sodium Channels; Tetrodotoxin

Topics: Action Potentials; Adsorption; Animals; Binding Sites; Demyelinating Diseases; Diffusion; Electric Stimulation; In Vitro Techniques; Kinetics; Models, Biological; Rabbits; Receptors, Drug; Tetrodotoxin; Vagus Nerve