Effect of Alkali Metal Cations on Slow Inactivation of Cardiac Na+ Channels

Human heart Na⁺ channels were expressed transiently in both mammalian cells and Xenopus oocytes, and Na⁺ currents measured using 150 mM intracellular Na⁺. The kinetics of decaying outward Na⁺ current in response to 1-s depolarizations in the F1485Q mutant depends on the predominant cation in the extracellular solution, suggesting an effect on slow inactivation. The decay rate is lower for the alkali metal cations Li⁺, Na⁺, K⁺, Rb⁺, and Cs⁺ than for the organic cations Tris, tetramethylammonium, N-methylglucamine, and choline. In whole cell recordings, raising [Na⁺]_o from 10 to 150 mM increases the rate of recovery from slow inactivation at −140 mV, decreases the rate of slow inactivation at relatively depolarized voltages, and shifts steady-state slow inactivation in a depolarized direction. Single channel recordings of F1485Q show a decrease in the number of blank (i.e., null) records when [Na⁺]_o is increased. Significant clustering of blank records when depolarizing at a frequency of 0.5 Hz suggests that periods of inactivity represent the sojourn of a channel in a slow-inactivated state. Examination of the single channel kinetics at +60 mV during 90-ms depolarizations shows that neither open time, closed time, nor first latency is significantly affected by [Na⁺]_o. However raising [Na⁺]_o decreases the duration of the last closed interval terminated by the end of the depolarization, leading to an increased number of openings at the depolarized voltage. Analysis of single channel data indicates that at a depolarized voltage a single rate constant for entry into a slow-inactivated state is reduced in high [Na⁺]_o, suggesting that the binding of an alkali metal cation, perhaps in the ion-conducting pore, inhibits the closing of the slow inactivation gate.     

Na+ Influx Inhibits Neuronal Ca2+ Channels

Experiments on isolated rat brain neurons with an elevated intracellular sodium concentration (due to tetanic stimulation) demonstrated the existence of earlier unknown negative modulation of calcium channels by intracellular sodium.     

Functional Interaction between the Scaffold Protein Kidins220/ARMS and Neuronal Voltage-Gated Na+ Channels

Kidins220 (kinase D-interacting substrate of 220 kDa)/ankyrin repeat-rich membrane spanning (ARMS) acts as a signaling platform at the plasma membrane and is implicated in a multitude of neuronal functions, including the control of neuronal activity. Here, we used the Kidins220^−/− mouse model to study the effects of Kidins220 ablation on neuronal excitability. Multielectrode array recordings showed reduced evoked spiking activity in Kidins220^−/− hippocampal networks, which was compatible with the increased excitability of GABAergic neurons determined by current-clamp recordings. Spike waveform analysis further indicated an increased sodium conductance in this neuronal subpopulation. Kidins220 association with brain voltage-gated sodium channels was shown by co-immunoprecipitation experiments and Na⁺ current recordings in transfected HEK293 cells, which revealed dramatic alterations of kinetics and voltage dependence. Finally, an in silico interneuronal model incorporating the Kidins220-induced Na⁺ current alterations reproduced the firing phenotype observed in Kidins220^−/− neurons. These results identify Kidins220 as a novel modulator of Na_v channel activity, broadening our understanding of the molecular mechanisms regulating network excitability.     

Block of Brain Sodium Channels by Peptide Mimetics of the Isoleucine,Phenylalanine, and Methionine (IFM) Motif from the Inactivation Gate

Inactivation of sodium channels is thought to be mediated by an inactivation gate formed by the intracellular loop connecting domains III and IV. A hydrophobic motif containing the amino acid sequence isoleucine, phenylalanine, and methionine (IFM) is required for the inactivation process. Peptides containing the IFM motif, when applied to the cytoplasmic side of these channels, produce two types of block: fast block, which resembles the inactivation process, and slow, use-dependent block stimulated by strong depolarizing pulses. Fast block by the peptide ac-KIFMK-NH₂, measured on sodium channels whose inactivation was slowed by the α-scorpion toxin from Leiurus quinquestriatus (LqTx), was reversed with a time constant of 0.9 ms upon repolarization. In contrast, control and LqTx-modified sodium channels were slower to recover from use-dependent block. For fast block, linear peptides of three to six amino acid residues containing the IFM motif and two positive charges were more effective than peptides with one positive charge, whereas uncharged IFM peptides were ineffective. Substitution of the IFM residues in the peptide ac-KIFMK-NH₂ with smaller, less hydrophobic residues prevented fast block. The positively charged tripeptide IFM-NH₂ did not cause appreciable fast block, but the divalent cation IFM-NH(CH₂)₂NH₂ was as effective as the pentapeptide ac-KIFMK-NH₂. The constrained peptide cyclic KIFMK containing two positive charges did not cause fast block. These results indicate that the position of the positive charges is unimportant, but flexibility or conformation of the IFM-containing peptide is important to allow fast block. Slow, use-dependent block was observed with IFM-containing peptides of three to six residues having one or two positive charges, but not with dipeptides or phenylalanine-amide. In contrast to its lack of fast block, cyclic KIFMK was an effective use-dependent blocker. Substitutions of amino acid residues in the tripeptide IFM-NH₂ showed that large hydrophobic residues are preferred in all three positions for slow, use-dependent block. However, substitution of the large hydrophobic residue diphenylalanine or the constrained residues phenylglycine or tetrahydroisoquinoline for phe decreased potency, suggesting that this phe residue must be able to enter a restricted hydrophobic pocket during the binding of IFM peptides. Together, the results on fast block and slow, use-dependent block indicate that IFM peptides form two distinct complexes of different stability and structural specificity with receptor site(s) on the sodium channel. It is proposed that fast block represents binding of these peptides to the inactivation gate receptor, while slow, use-dependent block represents deeper binding of the IFM peptides in the pore.     

Identification of the Intracellular Na+ Sensor in Slo2.1 Potassium Channels

Slo2 potassium channels have a very low open probability under normal physiological conditions, but are readily activated in response to an elevated [Na⁺]_i (e.g. during ischemia). An intracellular Na⁺ coordination motif (DX(R/K)XXH) was previously identified in Kir3.2, Kir3.4, Kir5.1, and Slo2.2 channel subunits. Based loosely on this sequence, we identified five potential Na⁺ coordination motifs in the C terminus of the Slo2.1 subunit. The Asp residue in each sequence was substituted with Arg, and single mutant channels were heterologously expressed in Xenopus oocytes. The Na⁺ sensitivity of each of the mutant channels was assessed by voltage clamp of oocytes using micropipettes filled with 2 m NaCl. Wild-type channels and four of the mutant Slo2.1 channels were rapidly activated by leakage of NaCl solution into the cytoplasm. D757R Slo2.1 channels were not activated by NaCl, but were activated by the fenamate niflumic acid, confirming their functional expression. In whole cell voltage clamp recordings of HEK293 cells, wild-type but not D757R Slo2.1 channels were activated by a [NaCl]_i of 70 mm. Thus, a single Asp residue can account for the sensitivity of Slo2.1 channels to intracellular Na⁺. In excised inside-out macropatches of HEK293 cells, activation of wild-type Slo2.1 currents by 3 mm niflumic acid was 14-fold greater than activation achieved by increasing [NaCl]_i from 3 to 100 mm. Thus, relative to fenamates, intracellular Na⁺ is a poor activator of Slo2.1.     

Idiosyncratic Gating of HERG-like K+ Channels in Microglia

A simple kinetic model is presented to explain the gating of a HERG-like voltage-gated K⁺ conductance described in the accompanying paper (Zhou, W., F.S. Cayabyab, P.S. Pennefather, L.C. Schlichter, and T.E. DeCoursey. 1998. J. Gen. Physiol. 111:781–794). The model proposes two kinetically distinct closing pathways, a rapid one favored by depolarization (deactivation) and a slow one favored by hyperpolarization (inactivation). The overlap of these two processes leads to a window current between −50 and +20 mV with a peak at −36 mV of ∼12% maximal conductance. The near absence of depolarization-activated outward current in microglia, compared with HERG channels expressed in oocytes or cardiac myocytes, can be explained if activation is shifted negatively in microglia. As seen with experimental data, availability predicted by the model was more steeply voltage dependent, and the midpoint more positive when determined by making the holding potential progressively more positive at intervals of 20 s (starting at −120 mV), rather than progressively more negative (starting at 40 mV). In the model, this hysteresis was generated by postulating slow and ultra-slow components of inactivation. The ultra-slow component takes minutes to equilibrate at −40 mV but is steeply voltage dependent, leading to protocol-dependent modulation of the HERG-like current. The data suggest that “deactivation” and “inactivation” are coupled through the open state. This is particularly evident in isotonic Cs⁺, where a delayed and transient outward current develops on depolarization with a decay time constant more voltage dependent and slower than the deactivation process observed at the same potential after a brief hyperpolarization.     

Macroscopic Na+ Currents in the “Nonconducting” Shaker Potassium Channel Mutant W434F

C-type inactivation in Shaker potassium channels inhibits K⁺ permeation. The associated structural changes appear to involve the outer region of the pore. Recently, we have shown that C-type inactivation involves a change in the selectivity of the Shaker channel, such that C-type inactivated channels show maintained voltage-sensitive activation and deactivation of Na⁺ and Li⁺ currents in K⁺-free solutions, although they show no measurable ionic currents in physiological solutions. In addition, it appears that the effective block of ion conduction produced by the mutation W434F in the pore region may be associated with permanent C-type inactivation of W434F channels. These conclusions predict that permanently C-type inactivated W434F channels would also show Na⁺ and Li⁺ currents (in K⁺-free solutions) with kinetics similar to those seen in C-type-inactivated Shaker channels. This paper confirms that prediction and demonstrates that activation and deactivation parameters for this mutant can be obtained from macroscopic ionic current measurements. We also show that the prolonged Na⁺ tail currents typical of C-type inactivated channels involve an equivalent prolongation of the return of gating charge, thus demonstrating that the kinetics of gating charge return in W434F channels can be markedly altered by changes in ionic conditions.     

钠离子通道研究进展

细胞电活动是生命现象的基本特征之一,而离子通道是其结构和功能的基础。因此,研究离子通道作用机制具有重大的理论和现实意义。许多神经系统和心血管疾病,如多发性硬化症、癫痫、脑溢血、神经痛、Brugada综合征(BrS)、进行性心脏传导缺陷(PCCD)和原发性心室纤颤(IVF)等,都与钠离子通道氨基酸序列和结构发生的改变有关。该文对钠离子通道的研究进展进行综述。     

Effects of Permeant Cations on K+ Channel Gating in Nerve Axons Revisited

An increase in extracellular potassium ion concentration, K _o , significantly slows the potassium channel deactivation rate in squid giant axons, as previously shown. Surprisingly, the effect does not occur in all preparations which, coupled with the voltage independence of this result in preparations in which it does occur, suggests that it is mediated at a site outside of the electric field of the channel, and that this site is accessible to potassium ions in some preparations, but not in others. In other words, the effect does not appear to be related to occupancy of the channel by potassium ions. This conclusion is supported by a four-barrier, three-binding site model of single file diffusion through the channel in which one site, at most, is unoccupied by a potassium ion (single-vacancy model). The model is consistent with current-voltage relations with various levels of K _o , and, by definition, with multiple occupancy by K⁺. The model predicts that occupancy of any given site is essentially independent of K _o (or K _i ). The effects of extracellular Rb⁺ and Cs⁺ on gating are strongly voltage dependent, and they were observed in all preparations investigated. Consequently, the mechanism underlying these results would appear to be different from that which underlies the effect of K⁺ on gating. In particular, the effect of Rb⁺ on gating is reduced by strong hyperpolarization, which in the context of the occupancy hypothesis, is consistent with the voltage dependence of the current-voltage relation in the presence of Rb⁺. The primary, novel, finding in this study is that the effects of Cs⁺ are counterintuitive in this regard. Specifically, the slowing of channel deactivation rate by Cs⁺ is also reduced by hyperpolarization, similar to the Rb⁺ results, whereas blockade is enhanced, which is seemingly inconsistent with the concept that occupancy of the channel by Cs⁺ underlies the effect of this ion on gating. This result is further elucidated by barrier modeling of the current-voltage relation in the presence of Cs⁺. Received: 19 December 1995/Revised: 10 June 1996     

A Conserved Residue Cluster That Governs Kinetics of ATP-dependent Gating of Kir6.2 Potassium Channels

ATP-sensitive potassium (K_ATP) channels are heteromultimeric complexes of an inwardly rectifying Kir channel (Kir6.x) and sulfonylurea receptors. Their regulation by intracellular ATP and ADP generates electrical signals in response to changes in cellular metabolism. We investigated channel elements that control the kinetics of ATP-dependent regulation of K_ATP (Kir6.2 + SUR1) channels using rapid concentration jumps. WT Kir6.2 channels re-open after rapid washout of ATP with a time constant of ∼60 ms. Extending similar kinetic measurements to numerous mutants revealed fairly modest effects on gating kinetics despite significant changes in ATP sensitivity and open probability. However, we identified a pair of highly conserved neighboring amino acids (Trp-68 and Lys-170) that control the rate of channel opening and inhibition in response to ATP. Paradoxically, mutations of Trp-68 or Lys-170 markedly slow the kinetics of channel opening (500 and 700 ms for W68L and K170N, respectively), while increasing channel open probability. Examining the functional effects of these residues using φ value analysis revealed a steep negative slope. This finding implies that these residues play a role in lowering the transition state energy barrier between open and closed channel states. Using unnatural amino acid incorporation, we demonstrate the requirement for a planar amino acid at Kir6.2 position 68 for normal channel gating, which is potentially necessary to localize the ϵ-amine of Lys-170 in the phosphatidylinositol 4,5-bisphosphate-binding site. Overall, our findings identify a discrete pair of highly conserved residues with an essential role for controlling gating kinetics of Kir channels.     

多不饱和脂肪酸对钾通道的调控作用及机理

多不饱和脂肪酸具有包括离子通道在内的众多作用靶点,通过作用于这些靶点,可以有效保护免疫系统、神经系统和心血管系统的功能,在一定程度上保护人体健康。电压门控钾离子通道家族K_V7通道和大电导钙离子激活的钾离子通道(BK_Ca)广泛表达于机体的各类组织中,具有重要的生理或病理功能。本综述围绕K_V7和BK_Ca通道,根据对已有报道的汇总,多不饱和脂肪酸可以增大K_V7和BK_Ca通道的电流幅值,其中对K_V7通道电流的影响主要是改变其电压依赖特性和最大电导值,而对BK_Ca通道电流的影响主要是改变其孔道区域关闭态的构象。此外,多不饱和脂肪酸对K_V7和BK_Ca通道功能的调节也会受到共表达的辅助亚基影响,但相关机制有待进一步阐明。深入理解多不饱和脂肪酸对K_V7和BK_Ca通道调节作用效果和分子机制,有助于全面理解K_V7和BK     

Separation of Gating Properties from Permeation and Block in mslo Large Conductance Ca-activated K+ Channels

In this and the following paper we have examined the kinetic and steady-state properties of macroscopic mslo Ca-activated K⁺ currents in order to interpret these currents in terms of the gating behavior of the mslo channel. To do so, however, it was necessary to first find conditions by which we could separate the effects that changes in Ca²⁺ concentration or membrane voltage have on channel permeation from the effects these stimuli have on channel gating. In this study we investigate three phenomena which are unrelated to gating but are manifest in macroscopic current records: a saturation of single channel current at high voltage, a rapid voltage-dependent Ca²⁺ block, and a slow voltage-dependent Ba²⁺ block. Where possible methods are described by which these phenomena can be separated from the effects that changes in Ca²⁺ concentration and membrane voltage have on channel gating. Where this is not possible, some assessment of the impact these effects have on gating parameters determined from macroscopic current measurements is provided. We have also found that without considering the effects of Ca²⁺ and voltage on channel permeation and block, macroscopic current measurements suggest that mslo channels do not reach the same maximum open probability at all Ca²⁺ concentrations. Taking into account permeation and blocking effects, however, we find that this is not the case. The maximum open probability of the mslo channel is the same or very similar over a Ca²⁺ concentration range spanning three orders of magnitude indicating that over this range the internal Ca²⁺ concentration does not limit the ability of the channel to be activated by voltage.     

Electrophysiological Properties of Ion Channels in Ascaris suum Tissue Incorporated into Planar Lipid Bilayers

Ion channels are important targets of anthelmintic agents. In this study, we identified 3 types of ion channels in Ascaris suum tissue incorporated into planar lipid bilayers using an electrophysiological technique. The most frequent channel was a large-conductance cation channel (209 pS), which accounted for 64.5% of channels incorporated (n=60). Its open-state probability (P_o) was ~0.3 in the voltage range of −60~+60 mV. A substate was observed at 55% of the main-state. The permeability ratio of Cl⁻ to K⁺ (P_Cl/P_K) was ~0.5 and P_Na/P_K was 0.81 in both states. Another type of cation channel was recorded in 7.5% of channels incorporated (n=7) and discriminated from the large-conductance cation channel by its smaller conductance (55.3 pS). Its Po was low at all voltages tested (~0.1). The third type was an anion channel recorded in 27.9% of channels incorporated (n=26). Its conductance was 39.0 pS and P_Cl/P_K was 8.6±0.8. P_o was ~1.0 at all tested potentials. In summary, we identified 2 types of cation and 1 type of anion channels in Ascaris suum. Gating of these channels did not much vary with voltage and their ionic selectivity is rather low. Their molecular nature, functions, and potentials as anthelmintic drug targets remain to be studied further.     

Ion Regulation of the Kinetics of Potential-Dependent Potassium Channels

We apply a theoretical approach developed earlier. The interaction ofions that permeate a channel with slowly relaxing charged channel-forminggroups (ion-conformational interaction – ICI) is addressed by thisapproach. One can describe the ion concentration influence (ion regulation)on channel functioning in this manner. A patch-clamp method in awhole-cell configuration is used to study the ICI. For this purpose theinfluence of an external concentration of potassium ions on thepotential-dependent potassium current (I_A) in the externalmembrane of GH₃ cells was studied. The increase of[K⁺ _out] from 5 mM to 100 mM causes anon-monotonous shift of current-voltage dependencies. The dependence of bothan activation time constant tgr_n and a steady-state activation(n) on [K⁺]_out have a minimum andmaximum respectively. The analysis of the results suggests that the observedeffects are caused by ICI. A physical model is developed to describe thedependence of the potassium channel kinetics on the external concentrationof the ions and the membrane potential. The deformation of the closedstate of the gate and the corresponding energy shifts cause the observednon-monotonous dependencies due to ICI. Thus, the general theoreticalapproach has an experimental confirmation and is applied to concreteexamples. Formulas for concentrational dependencies of the channel kineticsare given for practical uses.     

G-protein activation modulates pseudo-periodic oscillation of Na channel

We have shown before that the duration and amplitude of both prolonged (1-160 s) and short (100-1000 ms) depolarizing prepulse altered all the steady-state and kinetic parameters of rNav1.2a voltage-gated sodium channel in a pseudo-oscillatory fashion with variable time period and amplitude, often superimposed on a linear trend. In this study, we have examined the effect of G-protein activation on pseudo-oscillatory properties of the rNav1.2a sodium channel alpha subunit, heterologously expressed in Chinese hamster ovary cells. G-protein modification caused insignificant changes in the slow pseudo-periodic oscillation of the activation properties of sodium channel; only the time period of the oscillation was altered from approximately 30 to 21s. In contrast, G-protein activation abolished the faster component of pseudo-periodic oscillation in steady-state inactivation properties of sodium channel; the conditioning duration dependence of steady-state inactivation becomes monotonic in nature.     

Block of N-type Calcium Channels in Chick Sensory Neurons by External Sodium

L-type Ca²⁺ channels select for Ca²⁺ over sodium Na⁺ by an affinity-based mechanism. The prevailing model of Ca²⁺ channel permeation describes a multi-ion pore that requires pore occupancy by at least two Ca²⁺ ions to generate a Ca²⁺ current. At [Ca²⁺] < 1 μM, Ca²⁺ channels conduct Na⁺. Due to the high affinity of the intrapore binding sites for Ca²⁺ relative to Na⁺, addition of μM concentrations of Ca²⁺ block Na⁺ conductance through the channel. There is little information, however, about the potential for interaction between Na⁺ and Ca²⁺ for the second binding site in a Ca²⁺ channel already occupied by one Ca²⁺. The two simplest possibilities, (a) that Na⁺ and Ca²⁺ compete for the second binding site or (b) that full time occupancy by one Ca²⁺ excludes Na⁺ from the pore altogether, would imply considerably different mechanisms of channel permeation. We are studying permeation mechanisms in N-type Ca²⁺ channels. Similar to L-type Ca²⁺ channels, N-type channels conduct Na⁺ well in the absence of external Ca²⁺. Addition of 10 μM Ca²⁺ inhibited Na⁺ conductance by 95%, and addition of 1 mM Mg²⁺ inhibited Na⁺ conductance by 80%. At divalent ion concentrations of 2 mM, 120 mM Na⁺ blocked both Ca²⁺ and Ba²⁺ currents. With 2 mM Ba²⁺, the IC₅₀ for block of Ba²⁺ currents by Na⁺ was 119 mM. External Li⁺ also blocked Ba²⁺ currents in a concentration-dependent manner, with an IC₅₀ of 97 mM. Na⁺ block of Ba²⁺ currents was dependent on [Ba²⁺]; increasing [Ba²⁺] progressively reduced block with an IC₅₀ of 2 mM. External Na⁺ had no effect on voltage-dependent activation or inactivation of the channel. These data suggest that at physiological concentrations, Na⁺ and Ca²⁺ compete for occupancy in a pore already occupied by a single Ca²⁺. Occupancy of the pore by Na⁺ reduced Ca²⁺ channel conductance, such that in physiological solutions, Ca²⁺ channel currents are between 50 and 70% of maximal.     

Voltage Gating of Shaker K+ Channels : The Effect of Temperature on Ionic and Gating Currents

Ionic (I_i) and gating currents (I_g) from noninactivating Shaker H4 K⁺ channels were recorded with the cut-open oocyte voltage clamp and macropatch techniques. Steady state and kinetic properties were studied in the temperature range 2–22°C. The time course of I_i elicited by large depolarizations consists of an initial delay followed by an exponential rise with two kinetic components. The main I_i component is highly temperature dependent (Q₁₀ > 4) and mildly voltage dependent, having a valence times the fraction of electric field (z) of 0.2–0.3 e_o. The I_g On response obtained between −60 and 20 mV consists of a rising phase followed by a decay with fast and slow kinetic components. The main I_g component of decay is highly temperature dependent (Q₁₀ > 4) and has a z between 1.6 and 2.8 e_o in the voltage range from −60 to −10 mV, and ∼0.45 e_o at more depolarized potentials. After a pulse to 0 mV, a variable recovery period at −50 mV reactivates the gating charge with a high temperature dependence (Q₁₀ > 4). In contrast, the reactivation occurring between −90 and −50 mV has a Q₁₀ = 1.2. Fluctuation analysis of ionic currents reveals that the open probability decreases 20% between 18 and 8°C and the unitary conductance has a low temperature dependence with a Q₁₀ of 1.44. Plots of conductance and gating charge displacement are displaced to the left along the voltage axis when the temperature is decreased. The temperature data suggests that activation consists of a series of early steps with low enthalpic and negative entropic changes, followed by at least one step with high enthalpic and positive entropic changes, leading to final transition to the open state, which has a negative entropic change.     

Characterization of the Participation of Sodium Channels on the Rise in Na+ Induced by 4-Aminopyridine (4-AP) in Synaptosomes

The participation of voltage-sensitive Na+ channels (VSSC) on the changes on internal (i) Na+, K+, Ca2+, and on DA, Glu, and GABA release caused by different concentrations of 4-AP was investigated in striatum synaptosomes. TTX, which abolished the increase in Na(i) (as determined with SBFI), induced by 0.1 mM 4-AP only inhibited by 30% the rise in Na(i) induced by 1 mM 4-AP. One millimolar 4-AP markedly decreased the fluorescence of the K+ indicator dye PBFI but 0.1 mM 4-AP did not. Like 1 mM 4-AP, ouabain decreased PBFI fluorescence and increased a considerable fraction of Na(i) in a TTX-insensitive manner. In contrast with the different TTX sensitivity of the rise in Na(i) induced by 0.1 and 1 mM 4-AP, the rise in Ca(i) (as determined with fura-2) induced by the two concentrations of 4-AP was markedly inhibited by TTX, as well as by omega-agatoxin in combination with omega-conotoxin GVIA, indicating that only the TTX-sensitive fraction of the rise in Na(i) induced by 4-AP is linked with the activation of presynaptic Ca2+ channels. It is concluded that the TTX-sensitive fraction of neurotransmitter release evoked by 4-AP is released by exocytosis, and the TTX insensitive fraction involves reversal of the neurotransmitters transporters. This contrasts with the exocytosis evoked by high K+ that is unchanged by TTX and with the neurotransmitter-transporter-mediated release evoked by veratridine, which is highly TTX sensitive and does not require activation of Ca2+ channels.     

Inactivation Kinetics and Mechanical Gating of Piezo1 Ion Channels Depend on Subdomains within the Cap

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脊神经损伤后钠电流的变化及其离子通道机制

目的：检测脊神经切断大鼠背根节（DRG）神经元重复放电能力和钠电流的变化,并研究介导其电流变化的钠通道亚型的表达情况。方法：脊神经切断术后2～8d慢性痛大鼠模型背根节急性分离,对中等直径DRG神经元运用全细胞膜片钳技术记录神经元放电和钠电流的变化。对背根节神经元进行RT-PCR检测,分析其钠通道亚型的表达情况。结果：电流钳下,实验组DRG神经元在电流刺激下产生重复放电,而对照组神经元多诱发单个动作电位,电压钳记录发现实验组背根节神经元快钠电流和持续性钠电流幅值均明显大于对照组,PCR结果显示,Nav1.3、Nav1.7和Nav1.8通道亚型mRNA表达显著增高。结论：钠通道介导了脊神经受损模型的DRG神经元兴奋性增高,持续性钠电流可能通过调节阈下膜电位振荡的产生调节神经元兴奋性。