scholarly journals Evidence for a role of Nav1.6 in facilitating increases in neuronal hyperexcitability during epileptogenesis

2013 ◽  
Vol 110 (5) ◽  
pp. 1144-1157 ◽  
Author(s):  
Nicholas J. Hargus ◽  
Aradhya Nigam ◽  
Edward H. Bertram ◽  
Manoj K. Patel
Keyword(s):  
Entorhinal Cortex ◽  
Neuronal Excitability ◽  
Epileptic Seizures ◽  
Na Channel ◽  
Na Current ◽  
Neuronal Hyperexcitability ◽  
Cellular Events ◽  
Na Currents ◽  
Electrical Induction ◽  
Resurgent Currents

During epileptogenesis a series of molecular and cellular events occur, culminating in an increase in neuronal excitability, leading to seizure initiation. The entorhinal cortex has been implicated in the generation of epileptic seizures in both humans and animal models of temporal lobe epilepsy. This hyperexcitability is due, in part, to proexcitatory changes in ion channel activity. Sodium channels play an important role in controlling neuronal excitability, and alterations in their activity could facilitate seizure initiation. We sought to investigate whether medial entorhinal cortex (mEC) layer II neurons become hyperexcitable and display proexcitatory behavior of Na channels during epileptogenesis. Experiments were conducted 7 days after electrical induction of status epilepticus (SE), a time point during the latent period of epileptogenesis and before the onset of seizures. mEC layer II stellate neurons from post-SE animals were hyperexcitable, eliciting action potentials at higher frequencies compared with control neurons. Na channel currents recorded from post-SE neurons revealed increases in Na current amplitudes, particularly persistent and resurgent currents, as well as depolarized shifts in inactivation parameters. Immunocytochemical studies revealed increases in voltage-gated Na (Nav) 1.6 isoform levels. The toxin 4,9-anhydro-tetrodotoxin, which has greater selectivity for Nav1.6 over other Na channel isoforms, suppressed neuronal hyperexcitability, reduced macroscopic Na currents, persistent and resurgent Na current densities, and abolished depolarized shifts in inactivation parameters in post-SE neurons. These studies support a potential role for Nav1.6 in facilitating the hyperexcitability of mEC layer II neurons during epileptogenesis.

scholarly journals Kinetic analysis of the action of Leiurus scorpion alpha-toxin on ionic currents in myelinated nerve.

1985 ◽  
Vol 86 (5) ◽  
pp. 739-762 ◽  
Author(s):  
G K Wang ◽  
G Strichartz
Keyword(s):  
Steady State ◽  
Ionic Currents ◽  
Na Channel ◽  
Dependent Manner ◽  
Na Current ◽  
Toxin Concentration ◽  
Toxin Molecule ◽  
Channel Inactivation ◽  
Na Currents ◽  
Voltage Dependent

The effects of a neurotoxin, purified from the venom of the scorpion Leiurus quinquestriatus, on the ionic currents of toad single myelinated fibers were studied under voltage-clamp conditions. Unlike previous investigations using crude scorpion venom, purified Leiurus toxin II alpha at high concentrations (200-400 nM) did not affect the K currents, nor did it reduce the peak Na current in the early stages of treatment. The activation of the Na channel was unaffected by the toxin, the activation time course remained unchanged, and the peak Na current vs. voltage relationship was not altered. In contrast, Na channel inactivation was considerably slowed and became incomplete. As a result, a steady state Na current was maintained during prolonged depolarizations of several seconds. These steady state Na currents had a different voltage dependence from peak Na currents and appeared to result from the opening of previously inactivated Na channels. The opening kinetics of the steady state current were exponential and had rates approximately 100-fold slower than the normal activation processes described for transitions from the resting state to the open state. In addition, the dependence of the peak Na current on the potential of preceding conditioning pulses was also dramatically altered by toxin treatment; this parameter reached a minimal value near a membrane potential of -50 mV and then increased continuously to a "plateau" value at potentials greater than +50 mV. The amplitude of this plateau was dependent on toxin concentration, reaching a maximum value equal to approximately 50% of the peak current; voltage-dependent reversal of the toxin's action limits the amplitude of the plateauing effect. The measured plateau effect was half-maximum at a toxin concentration of 12 nM, a value quite similar to the concentration producing half of the maximum slowing of Na channel inactivation. The results of Hill plots for these actions suggest that one toxin molecule binds to one Na channel. Thus, the binding of a single toxin molecule probably both produces the steady state currents and slows the Na channel inactivation. We propose that Leiurus toxin inhibits the conversion of the open state to inactivated states in a voltage-dependent manner, and thereby permits a fraction of the total Na permeability to remain at membrane potentials where inactivation is normally complete.

Relationship between Na+ current expression and cell-cell coupling in astrocytes cultured from rat hippocampus

1991 ◽  
Vol 65 (4) ◽  
pp. 989-1002 ◽  
Author(s):  
H. Sontheimer ◽  
S. G. Waxman ◽  
B. R. Ransom
Keyword(s):  
Lucifer Yellow ◽  
Na Channel ◽  
Dye Coupling ◽  
Cell Coupling ◽  
Na Current ◽  
Na Currents ◽  
Coupled Cells ◽  
Hippocampal Astrocytes ◽  
Cell Cell

1. Cell-cell coupling between hippocampal astrocytes in culture was studied by following the intracellular spread of the low molecular weight fluorescent dye Lucifer yellow (LY). Dye coupling appeared as early as 24 h after plating, at which time approximately 20% of all astrocytes that physically contacted neighboring cells showed dye coupling. 2. The percentage of coupled cells increased with time in culture and peaked after 10 days in vitro (DIV) when approximately 50% of astrocytes showed coupling. Further time in culture, up to 20 DIV, did not increase the percentage of coupled cells. Thus, coupled and noncoupled astrocytes coexist in hippocampal cultures in approximately equal numbers. 3. Na+ currents were expressed in a subpopulation of hippocampal astrocytes and changed characteristics during in vitro development. A "neuronal type" of Na+ current, so called because of an h alpha curve that had a midpoint near -60 mV, was observed within the first 5 days post-plating. A "glial type" of Na+ current, characterized by a -25 mV shift in its h alpha curve, was only expressed after 6 days in culture. 4. Na+ current expression was restricted to hippocampal astrocytes that did not exhibit dye coupling; astrocytes that exhibited dye coupling (n = 39) did not show measurable Na+ currents. 5. The failure to see Na+ currents in coupled astrocytes cannot be explained by insufficient space-clamp since astrocytes acutely uncoupled with octanol (10 microM) did not reveal Na+ current expression. Control experiments showed that low concentrations of octanol (i.e., 10-100 microM) did not block Na+ currents; blockage of Na+ currents by octanol was only observed at high concentrations (e.g., 50-fold the concentration used for uncoupling). These observations support the idea that Na(+)-channel expression was restricted to noncoupled astrocytes. 6. The time courses for the development of cell coupling and Na+ current expression appeared to be inversely correlated and suggested a gradual increase in cell coupling in concert with a loss in Na+ current expression with time in culture.

Resurgent Na Currents in Four Classes of Neurons of the Cerebellum

2004 ◽  
Vol 92 (5) ◽  
pp. 2831-2843 ◽  
Author(s):  
Fatemeh S. Afshari ◽  
Krzysztof Ptak ◽  
Zayd M. Khaliq ◽  
Tina M. Grieco ◽  
N. Traverse Slater ◽  
...  
Keyword(s):  
Open Channel ◽  
Granule Cells ◽  
Na Channel ◽  
Na Channels ◽  
Brush Cells ◽  
Cerebellar Neurons ◽  
Na Currents ◽  
Unipolar Brush Cells ◽  
Resurgent Current ◽  
Resurgent Currents

Action potential firing rates are generally limited by the refractory period, which depends on the recovery from inactivation of voltage-gated Na channels. In cerebellar Purkinje neurons, the kinetics of Na channels appear specialized for rapid firing. Upon depolarization, an endogenous open-channel blocker rapidly terminates current flow but prevents binding of the “fast” inactivation gate. Upon repolarization, unbinding of the blocker produces “resurgent” Na current while allowing channels to recover rapidly. Because other cerebellar neurons, including granule cells, unipolar brush cells, and neurons of the cerebellar nuclei, also fire rapidly, we tested whether these cells might also express Na channels with resurgent kinetics. Neurons were acutely isolated from mice and rats, and TTX-sensitive Na currents were recorded under voltage clamp. Unlike Purkinje cells, the other cerebellar neurons produced only tiny resurgent currents in solutions optimized for voltage-clamping Na currents (50 mM Na+; Co2+ substitution for Ca2+). Under more physiological ionic conditions (155 mM Na+; 2 mM Ca2+ with 0.03 mM Cd2+), however, granule cells, unipolar brush cells, and cerebellar nuclear cells all produced robust resurgent currents. The increase in resurgent current, which was greater than predicted by the Goldman-Hodgkin-Katz equation, appeared to result from a combination of knock-off of open-channel blockers by permeating ions as well as relief of divalent block at negative potentials. These results indicate that resurgent current is typical of many cerebellar neurons and suggest that rapid open-channel block and unblock may be a widespread mechanism for restoration of Na channel availability in rapidly firing neurons.

scholarly journals Na channel distribution in vertebrate skeletal muscle.

1986 ◽  
Vol 87 (6) ◽  
pp. 907-932 ◽  
Author(s):  
J H Caldwell ◽  
D T Campbell ◽  
K G Beam
Keyword(s):  
Skeletal Muscle ◽  
Current Density ◽  
Sharp Decrease ◽  
Na Channel ◽  
Na Current ◽  
Na Currents ◽  
Current Densities ◽  
Kinetics Of ◽  
A Current ◽  
Vertebrate Skeletal Muscle

The loose patch voltage clamp has been used to map Na current density along the length of snake and rat skeletal muscle fibers. Na currents have been recorded from (a) endplate membrane exposed by removal of the nerve terminal, (b) membrane near the endplate, (c) extrajunctional membrane far from both the endplate and the tendon, and (d) membrane near the tendon. Na current densities recorded directly on the endplate were extremely high, exceeding 400 mA/cm2 in some patches. The membrane adjacent to the endplate has a current density about fivefold lower than that of the endplate, but about fivefold higher than the membrane 100-200 micron from the endplate. Small local variations in Na current density are recorded in extrajunctional membrane. A sharp decrease in Na current density occurs over the last few hundred micrometers from the tendon. We tested the ability of tetrodotoxin to block Na current in regions close to and far from the endplate and found no evidence for toxin-resistant channels in either region. There was also no obvious difference in the kinetics of Na current in the two regions. On the basis of the Na current densities measured with the loose patch clamp, we conclude that Na channels are abundant in the endplate and near-endplate membrane and are sparse close to the tendon. The current density at the endplate is two to three orders of magnitude higher than at the tendon.

Na+ Channel Expression and Neuronal Function in the Na+/H+ Exchanger 1 Null Mutant Mouse

2003 ◽  
Vol 89 (1) ◽  
pp. 229-236 ◽  
Author(s):  
Ying Xia ◽  
Peng Zhao ◽  
Jin Xue ◽  
Xiang Q. Gu ◽  
Xiaolu Sun ◽  
...  
Keyword(s):  
Current Density ◽  
Cortical Neurons ◽  
Neuronal Excitability ◽  
Epileptic Seizures ◽  
Mutant Mice ◽  
Na Channel ◽  
Membrane Excitability ◽  
Altered Expression ◽  
Ca1 Neurons ◽  
Channel Expression

Mice lacking Na+/H+ exchanger 1 (NHE1) suffer from recurrent seizures and die early postnatally. Although the mechanisms for seizures are not well established, our previous electrophysiological work has shown that neuronal excitability and Na+ current density are increased in hippocampal CA1 neurons of these mutant mice. However, it is unknown whether this increased density is related to altered expression or functional regulation of Na+ channels. In this work, we asked three questions: is the increased excitability limited to CA1 neurons, is the increased Na+ current density related to an increased Na+ channel expression, and, if so, which Na+channel subtype(s) is upregulated? Using neurophysiological, autoradiographic, and immunoblotting techniques, we showed that both CA1 and cortical neurons have an increase in membrane excitability and Na+ current density; Na+ channel density is selectively upregulated in the hippocampus and cortex ( P < 0.05); and Na+ channel subtype I is significantly increased in the hippocampus and Na+channel subtype II is increased in the cortex. Our results demonstrate that mice lacking NHE1 upregulate their Na+ channel expression in the hippocampal and cortical regions selectively; this leads to an increase in Na+ current density and membrane excitability. We speculate that neuronal overexcitability due to Na+ channel upregulation in the hippocampus and cortex forms the basis of epileptic seizures in NHE1 mutant mice.

scholarly journals Altered stereoselectivity of cocaine and bupivacaine isomers in normal and batrachotoxin-modified Na+ channels.

1992 ◽  
Vol 100 (6) ◽  
pp. 1003-1020 ◽  
Author(s):  
G K Wang ◽  
S Y Wang
Keyword(s):  
Gh3 Cells ◽  
Na Channel ◽  
Optical Isomers ◽  
Na Channels ◽  
Na Current ◽  
Response Curves ◽  
Whole Cell Patch Clamp ◽  
Na Currents ◽  
Steady State Inactivation ◽  
Order Of Magnitude

The inhibitory effects of local anesthetics (LAs) of cocaine and bupivacaine optical isomers on Na+ currents were studied in clonal GH3 cells under whole-cell patch clamp conditions. At holding potential of -100 mV, all four isomers inhibited peak Na+ currents when the cell was stimulated infrequently. The dose-response curves of this tonic block of peak Na+ currents by (-)/(+) cocaine and (-)/(+) bupivacaine were well fitted by the Langmuir isotherm, suggesting that one LA isomer blocked one Na+ channel. Each pair of isomers showed no greater than a twofold difference in stereoselectivity toward Na+ channels. Additional block of Na+ currents occurred when the cell was stimulated at 2 Hz. This use-dependent block was also observed in all four isomers, which again displayed little stereoselectivity. The voltage dependence of the use-dependent block produced by cocaine isomers did not overlap with the activation of Na+ channels but did overlap with the steady-state inactivation (h infinity), indicating that cocaine can bind directly to the inactivated state of Na+ channels before channel opening. In comparison, the peak batrachotoxin (BTX)-modified Na+ currents were little inhibited by cocaine and bupivacaine isomers. However, the maintained BTX-modified Na+ currents were highly sensitive toward the (-) form of cocaine and bupivacaine isomers during a prolonged depolarization. As a result, a profound time-dependent block of BTX-modified Na+ currents was evident in the presence of these LA isomers. The estimated values of the equilibrium dissociation constant (KD in micromolar) at +50 mV were 35.8, 661, 7.0, and 222 for (-)/(+) cocaine and (-)/(+) bupivacaine, respectively. Although chloramine-T (CT) also modified the fast inactivation of Na+ channels and gave rise to a maintained Na+ current during a prolonged depolarization, LA isomers showed no greater stereoselectivity in blocking this maintained current than in blocking the normal transient Na+ current. We conclude that (a) cocaine and bupivacaine isomers exhibit only weak stereoselectivity toward the LA receptor in normal and CT-treated Na+ channels, (b) BTX drastically modifies the configuration of the LA binding site so that the LA stereoselectivity of the open Na+ channels is altered by an order of magnitude, and (c) the (-) forms of cocaine and bupivacaine interact strongly with the open state of BTX-modified Na+ channels but only weakly, if at all, with the closed state. The last finding may explain why most LA drugs were reported to be less effective toward BTX-modified Na+ channels.(ABSTRACT TRUNCATED AT 400 WORDS)

Pharmacological properties of voltage-gated Na+ currents in motor neurones from a hydrozoan jellyfish Polyorchis penicillatus

1996 ◽  
Vol 199 (4) ◽  
pp. 941-948 ◽  
Author(s):  
J Spafford ◽  
N Grigoriev ◽  
A Spencer
Keyword(s):  
Local Anaesthetics ◽  
Bay K 8644 ◽  
Na Channel ◽  
Pharmacological Properties ◽  
Dependent Manner ◽  
Na Current ◽  
Polyorchis Penicillatus ◽  
Na Currents ◽  
Motor Neurones ◽  
High Concentrations

The Na+ current of 'swimming motor neurones' in the hydromedusan Polyorchis penicillatus was tetrodotoxin-insensitive. The local anaesthetics lidocaine and procainamide caused partial, non use-dependent blockade of the Na+ channel. Veratridine produced partial blockade of the Na+ channel without affecting inactivation. An order of blocking potency of di- and trivalent cations was established as: La3+ = Zn2+ = Cd2+ &gt; Ni2+ &gt; Mn2+ = Co2+ &gt; Ca2+ &gt; Ba2+ &gt; Mg2+. All these cations, except Ba2+, produced depolarizing shifts in the conductance-voltage curves. Even at relatively high concentrations, the dihydropyridines nicardipine, nitrendipine and (+)Bay K 8644 produced only weak blockade of the Na+ current; while nimodipine, nifedipine and (-)Bay K 8644 were ineffective. Diltiazem and verapamil weakly blocked the Na+ current in a dose-dependent manner with no evidence of use-dependence. The calmodulin inhibitors W7 and calmidazolium were ineffective blockers of Na+ currents. Crude Conus venoms and the Conus peptides, &micro;-conotoxin GIIA, &micro;O-conotoxin MrVIA, omega-conotoxin GVIA and omega-conotoxin MVIIC, were without effect. Capsaicin produced rapid, reversable blockade of Na+ current. It has been suggested that 'primitive' Na+ channels could be expected to have pharmacological properties that are intermediate between those of Na+ and Ca2+ channels. If such channels exist, the Na+ channel described here is clearly not one of them.

Cn11, the first example of a scorpion toxin that is a true blocker of Na+ currents in crayfish neurons

2002 ◽  
Vol 205 (6) ◽  
pp. 869-876
Author(s):  
Martha E. Ramirez-Dominguez ◽  
Timoteo Olamendi-Portugal ◽  
Ubaldo Garcia ◽  
Consuelo Garcia ◽  
Hugo Arechiga ◽  
...  
Keyword(s):  
Procambarus Clarkii ◽  
Maximum Amplitude ◽  
Na Channel ◽  
Amino Acid Residues ◽  
Scorpion Toxins ◽  
Na Current ◽  
Gating Mechanism ◽  
Na Currents ◽  
Gland System ◽  
K Currents

SUMMARY A novel crustacean toxin (Cn11) was isolated and characterized from the venom of the Mexican scorpion Centruroides noxius Hoffmann. It contains 63 amino acid residues and is stabilized by four disulphide bridges. It is lethal to crustaceans (Cambarellus montezumae), less toxic to insects (crickets) and non-toxic to mammals (mice) at the doses assayed. In neurons isolated from the X organ–sinus gland system of the crayfish Procambarus clarkii, it blocks the Na+ currents with an estimated Km of 320 nmol l–1, without affecting the Ca2+ and K+ currents. The voltage-gated tetrodotoxin-sensitive Na+ current was recorded from X organ neurons in culture 24 h after plating using the whole-cell clamp configuration. The Na+ current was isolated by blocking Ca2+ currents with Cd2+ and Cs+ and K+ currents with tetraethylammonium and 4-aminopyridine. Under control conditions, the Na+ currents were activated at –40 mV with a maximum amplitude at 0 mV. In the presence of 1 μmol l–1 Cn11, the Na+ current amplitude was reduced by 75 % without apparent modifications to the gating mechanism. These findings suggest that Cn11 selectively blocks a Na+ channel. It is the first representative of a new group of scorpion toxins specific for this molecular target.

scholarly journals Evidence for a population of sleepy sodium channels in squid axon at low temperature.

1982 ◽  
Vol 79 (5) ◽  
pp. 739-758 ◽  
Author(s):  
D R Matteson ◽  
C M Armstrong
Keyword(s):  
Steady State ◽  
Single Channel ◽  
Reversal Potential ◽  
Current Amplitude ◽  
Na Channel ◽  
Na Current ◽  
Current Increase ◽  
Temperature Changes ◽  
Na Currents ◽  
Steady State Current

We have studied the effects of temperature changes on Na currents in squid giant axons. Decreases in temperature in the 15-1 degrees C range decrease peak Na current with a Q10 of 2.2. Steady state currents, which are tetrodotoxin sensitive and have the same reversal potential as peak currents, are almost unaffected by temperature changes. After removal of inactivation by pronase treatment, steady state current amplitude has a Q10 of 2.3. Na currents generated at large positive voltages sometimes exhibit a biphasic activation pattern. The first phase activates rapidly and partially inactivates and is followed by a secondary slow current increase that lasts several milliseconds. Peak Na current amplitude can be increased by delivering large positive prepulses, an effect that is more pronounced at low temperatures. The slow activation phase is eliminated after a positive prepulse. The results are consistent with the hypothesis that there are two forms of the Na channel: (a) rapidly activating channels that completely inactivate, and (b) slowly activating "sleepy" channels that inactivate slowly if at all. Some fast channels are assumed to be converted to sleepy channels by cooling, possibly because of a phase transition in the membrane. The existence of sleepy channels complicates the determination of the Q10 of gating parameters and single-channel conductance.

scholarly journals Protein arginine methylation facilitates KCNQ channel-PIP2 interaction leading to seizure suppression

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Hyun-Ji Kim ◽  
Myong-Ho Jeong ◽  
Kyung-Ran Kim ◽  
Chang-Yun Jung ◽  
Seul-Yi Lee ◽  
...  
Keyword(s):  
Neuronal Excitability ◽  
Epileptic Seizures ◽  
Arginine Methylation ◽  
Neuronal Hyperexcitability ◽  
Channel Modulation ◽  
Kcnq Channels ◽  
Kinase C ◽  
Arginine Residues ◽  
Novel Target ◽  
Protein Arginine Methyltransferase 1

KCNQ channels are critical determinants of neuronal excitability, thus emerging as a novel target of anti-epileptic drugs. To date, the mechanisms of KCNQ channel modulation have been mostly characterized to be inhibitory via Gq-coupled receptors, Ca2+/CaM, and protein kinase C. Here we demonstrate that methylation of KCNQ by protein arginine methyltransferase 1 (Prmt1) positively regulates KCNQ channel activity, thereby preventing neuronal hyperexcitability. Prmt1+/- mice exhibit epileptic seizures. Methylation of KCNQ2 channels at 4 arginine residues by Prmt1 enhances PIP2 binding, and Prmt1 depletion lowers PIP2 affinity of KCNQ2 channels and thereby the channel activities. Consistently, exogenous PIP2 addition to Prmt1+/- neurons restores KCNQ currents and neuronal excitability to the WT level. Collectively, we propose that Prmt1-dependent facilitation of KCNQ-PIP2 interaction underlies the positive regulation of KCNQ activity by arginine methylation, which may serve as a key target for prevention of neuronal hyperexcitability and seizures.

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