Slow inactivation in voltage-gated sodium channels

Slow inactivation in voltage-gated sodium channels is a biophysical process that governs the availability of sodium channels over extended periods of time. Slow inactivation, therefore, plays an important role in controlling membrane excitability, firing properties, and spike frequency adaptation. Defective slow inactivation is associated with several diseases of cell excitability, such as hyperkalemic periodic paralysis, myotonia, idiopathic ventricular fibrillation and long-QT syndrome. These associations underscore the physiological importance of this phenomenon. Nevertheless, our understanding of the molecular substrates for slow inactivation is still fragmentary. This review covers the current state of knowledge concerning the molecular underpinnings of slow inactivation, and its relationship with other biophysical processes of voltage-gated sodium channels.     

Excitability constraints on voltage-gated sodium channels

We study how functional constraints bound and shape evolution through an analysis of mammalian voltage-gated sodium channels. The primary function of sodium channels is to allow the propagation of action potentials. Since Hodgkin and Huxley, mathematical models have suggested that sodium channel properties need to be tightly constrained for an action potential to propagate. There are nine mammalian genes encoding voltage-gated sodium channels, many of which are more than approximately 90% identical by sequence. This sequence similarity presumably corresponds to similarity of function, consistent with the idea that these properties must be tightly constrained. However, the multiplicity of genes encoding sodium channels raises the question: why are there so many? We demonstrate that the simplest theoretical constraints bounding sodium channel diversity--the requirements of membrane excitability and the uniqueness of the resting potential--act directly on constraining sodium channel properties. We compare the predicted constraints with functional data on mammalian sodium channel properties collected from the literature, including 172 different sets of measurements from 40 publications, wild-type and mutant, under a variety of conditions. The data from all channel types, including mutants, obeys the excitability constraint; on the other hand, channels expressed in muscle tend to obey the constraint of a unique resting potential, while channels expressed in neuronal tissue do not. The excitability properties alone distinguish the nine sodium channels into four different groups that are consistent with phylogenetic analysis. Our calculations suggest interpretations for the functional differences between these groups.     

Conotoxin modulation of voltage-gated sodium channels

The rising phase of the action potential in excitable cells is mediated by voltage-gated sodium channels (VGSCs), of which there are nine mammalian subtypes with distinct tissue distribution and biophysical properties. The involvement of certain VGSC subtypes in disease states such as pain and epilepsy highlights the need for agents that modulate VGSCs in a subtype-specific manner. Conotoxins from marine snails of the Conus genus constitute a promising source of such modulators, since these peptide toxins have evolved to become selective for various membrane receptors, ion channels and transporters in excitable cells. This review covers the structure and function of three classes of conopeptides that modulate VGSCs: the pore-blocking mu-conotoxins, the delta-conotoxins which delay or inhibit VGSC inactivation, and the muO-conotoxins which inhibit VGSC Na(+) conductance independent of the tetrodotoxin binding site. Some of these toxins have potential therapeutic and research applications, in particular the muO-conotoxins, which may develop into potential drug leads for the treatment of pain states.     

Growth of normal and neoplastic rat pleural mesothelial cells in the presence of conditioned medium from neoplastic mesothelial cells

Several transformed cells have been demonstrated to secrete growth factors. We studied the effect of conditioned medium from neoplastic rat pleural mesothelial cells on normal and neoplastic mesothelial cell growth. The results showed that the concentrated conditioned medium stimulated neoplastic mesothelial cell growth but inhibited reversibly normal mesothelial cell growth.     

A superfamily of voltage-gated sodium channels in bacteria

NaChBac, a six-alpha-helical transmembrane-spanning protein cloned from Bacillus halodurans, is the first functionally characterized bacterial voltage-gated Na(+)-selective channel. As a highly expressing ion channel protein, NaChBac is an ideal candidate for high resolution structural determination and structure-function studies. The biological role of NaChBac, however, is still unknown. In this report, another 11 structurally related bacterial proteins are described. Two of these functionally expressed as voltage-dependent Na(+) channels (Na(V)PZ from Paracoccus zeaxanthinifaciens and Na(V)SP from Silicibacter pomeroyi). Na(V)PZ and Na(V)SP share approximately 40% amino acid sequence identity with NaChBac. When expressed in mammalian cell lines, both Na(V)PZ and Na(V)SP were Na(+)-selective and voltage-dependent. However, their kinetics and voltage dependence differ significantly. These single six-alpha-helical transmembrane-spanning subunits constitute a widely distributed superfamily (Na(V)Bac) of channels in bacteria, implying a fundamental prokaryotic function. The degree of sequence homology (22-54%) is optimal for future comparisons of Na(V)Bac structure and function of similarity and dissimilarity among Na(V)Bac proteins. Thus, the Na(V)Bac superfamily is fertile ground for crystallographic, electrophysiological, and microbiological studies.     

Dynamic compartmentalization of the voltage-gated sodium channels in axons

One of the major physiological roles of the neuronal voltage-gated sodium channel is to generate action potentials at the axon hillock/initial segment and to ensure propagation along myelinated or unmyelinated fibers to nerve terminal. These processes require a precise distribution of sodium channels accumulated at high density in discrete subdomains of the nerve membrane. In neurons, information relevant to ion channel trafficking and compartmentalization into sub-domains of the plasma membrane is far from being elucidated. Besides, whereas information on dendritic targeting is beginning to emerge, less is known about the mechanisms leading to the polarized distribution of proteins in axon. To obtain a better understanding of how neurons selectively target sodium channels to discrete subdomains of the nerve, we addressed the question as to whether any of the large intracellular regions of Nav1.2 contain axonal sorting and/or clustering signals. We first obtained evidence showing that addition of the cytoplasmic carboxy-terminal region of Nav1.2 restricted the distribution of a dendritic-axonal reporter protein to axons of hippocampal neurons. The analysis of mutants revealed that a di-leucine-based motif mediates chimera compartmentalization in axons and its elimination in soma and dendrites by endocytosis. The analysis of the others generated chimeras showed that the determinant conferring sodium channel clustering at the axonal initial segment is contained within the cytoplasmic loop connecting domains II-III of Nav1.2. Expression of a soluble Nav1.2 II-III linker protein led to the disorganization of endogenous sodium channels. The motif was sufficient to redirect a somatodendritic potassium channel to the axonal initial segment, a process involving association with ankyrin G. Thus, it is conceivable that concerted action of the two determinants is required for sodium channel compartmentalization in axons.     

Trafficking and cellular distribution of voltage-gated sodium channels

Electrical excitability in cells such as neurons and myocytes depends not only upon the expression of voltage-gated sodium channels but also on their correct targeting within the plasma membrane. Placing sodium channels within a broader cell biological context is beginning to shed new light on a variety of important questions such as the integration of neuronal signaling. Mutations that affect sodium channel trafficking have been shown to underlie several life-threatening conditions including cardiac arrhythmias, revealing an important clinical context to these studies.     

电压门控钠通道在神经病理性痛中的作用

电压门控钠通道(VGSC)在神经病理性痛的发生和维持中起重要作用。非特异性的通道阻断剂是神经病理性痛的一种治疗手段,但由于可能产生严重的副作用而限制了其使用。最近研究揭示了几种主要在外周感觉神经系统中表达的VGSC的亚型与神经病理性痛密切相关,发展特异性的通道亚型阻断剂将成为治疗神经病理性痛的重要研究方向。     

The role of voltage-gated sodium channels in neuropathic pain

Use-dependent inhibitors of voltage-gated sodium channels (VGSC) are important therapeutic tools for chronic pain management, but are limited by possible severe side effects. Recent studies have provided much new information on the function of several voltage-gated sodium channels that are predominantly expressed in peripheral sensory neurons, and on their possible link to pathological pain states arising from injuries to the sensory nerve. The use of antisense oligonucleotides to target specific channel subtypes shows that the functional localization of the channel subtype Na(V)1.8 after nerve injury is essential for persistent pain states. The putative roles of Na(V)1.3 and Na(V)1.9 in neuropathic pain are also discussed. These studies may form a basis for developing inhibitors to target specific channel subtype(s) for use in chronic pain treatment.     

Theoretical and simulation studies on voltage-gated sodium channels

Voltage-gated sodium (Na_v) channels are indispensable membrane elements for the generation and propagation of electric signals in excitable cells. The successes in the crystallographic studies on prokaryotic Na_v channels in recent years greatly promote the mechanistic investigation of these proteins and their eukaryotic counterparts. In this paper, we mainly review the progress in computational studies, especially the simulation studies, on these proteins in the past years.     

Molecular interaction of delta-conotoxins with voltage-gated sodium channels

Various neurotoxic peptides modulate voltage-gated sodium (Na(V)) channels and thereby affect cellular excitability. Delta-conotoxins from predatory cone snails slow down inactivation of Na(V) channels, but their interaction site and mechanism of channel modulation are unknown. Here, we show that delta-conotoxin SVIE from Conus striatus interacts with a conserved hydrophobic triad (YFV) in the domain-4 voltage sensor of Na(V) channels. This site overlaps with that of the scorpion alpha-toxin Lqh-2, but not with the alpha-like toxin Lqh-3 site. Delta-SVIE functionally competes with Lqh-2, but exhibits strong cooperativity with Lqh-3, presumably by synergistically trapping the voltage sensor in its "on" position.     

电压门控钠离子通道与恶性肿瘤的转移

电压门控钠离子通道(VGSC)是可兴奋组织中动作电位的关键离子通道,具有重要的生理功能.近年来国内外研究发现,VGSC在转移的前列腺癌、乳腺癌、卵巢癌、宫颈癌等细胞中表达,其增加了癌细胞的运动和侵袭,促使了癌症的转移,其还将被作为治疗靶点而进行药物开发和临床应用.     

Molecular mechanisms of neurotoxin action on voltage-gated sodium channels

Voltage-gated sodium channels are the molecular targets for a broad range of neurotoxins that act at six or more distinct receptor sites on the channel protein. These toxins fall into three groups. Both hydrophilic low molecular mass toxins and larger polypeptide toxins physically block the pore and prevent sodium conductance. Alkaloid toxins and related lipid-soluble toxins alter voltage-dependent gating of sodium channels via an allosteric mechanism through binding to intramembranous receptor sites. In contrast, polypeptide toxins alter channel gating by voltage sensor trapping through binding to extracellular receptor sites. The results of recent studies that define the receptor sites and mechanisms of action of these diverse toxins are reviewed here.     

Adult alveolar epithelial cells express multiple subtypes of voltage-gated K+ channels that are located in apical membrane

Whole cell perforated patch-clampexperiments were performed with adult rat alveolar epithelial cells.The holding potential was 60 mV, and depolarizing voltage stepsactivated voltage-gated K⁺ (Kv) channels. Thevoltage-activated currents exhibited a mean reversal potential of 32mV. Complete activation was achieved at 10 mV. The currents exhibitedslow inactivation, with significant variability in the time coursebetween cells. Tail current analysis revealed cell-to-cell variabilityin K⁺ selectivity, suggesting contributions of multiple Kv-subunits to the whole cell current. The Kv channels also displayedsteady-state inactivation when the membrane potential was held atdepolarized voltages with a window current between 30 and 5 mV.Analysis of RNA isolated from these cells by RT-PCR revealed thepresence of eight Kv -subunits (Kv1.1, Kv1.3, Kv1.4, Kv2.2, Kv4.1,Kv4.2, Kv4.3, and Kv9.3), three -subunits (Kv1.1, Kv2.1, andKv3.1), and two K⁺ channel interacting protein (KChIP)isoforms (KChIP2 and KChIP3). Western blot analysis with available Kv-subunit antibodies (Kv1.1, Kv1.3, Kv1.4, Kv4.2, and Kv4.3) showedlabeling of 50-kDa proteins from alveolar epithelial cells grown inmonolayer culture. Immunocytochemical analysis of cells from monolayersshowed that Kv1.1, Kv1.3, Kv1.4, Kv4.2, and Kv4.3 were localized to theapical membrane. We conclude that expression of multiple Kv -, -,and KChIP subunits explains the variability in inactivation gating andK⁺ selectivity observed between cells and that Kv channelsin the apical membrane may contribute to basal K⁺ secretionacross the alveolar epithelium.

     

Sensory neuron voltage-gated sodium channels as analgesic drug targets

Voltage-gated sodium channels are crucial determinants of neuronal excitability and signalling; some specific channel subtypes have been implicated in a number of chronic pain conditions. Human genetic studies show gain-of-function or loss-of-function mutations in Na(V)1.7 lead to an enhancement or lack of pain, respectively, whilst transgenic mouse and knockdown studies have implicated Na(V)1.3, Na(V)1.8 and Na(V)1.9 in peripheral pain pathways. The development of subtype-specific sodium channel blockers, though clearly desirable, has been technically challenging. Recent advances exploiting both natural products and small molecule selective channel blockers have demonstrated that this approach to pain control is feasible. These observations provide a rationale for the development of new analgesics without the side effect profile of broad spectrum sodium channel blockers.     

BACE1 regulates voltage-gated sodium channels and neuronal activity

BACE1 activity is significantly increased in the brains of Alzheimer's disease patients, potentially contributing to neurodegeneration. The voltage-gated sodium channel (Na(v)1) beta2-subunit (beta2), a type I membrane protein that covalently binds to Na(v)1 alpha-subunits, is a substrate for BACE1 and gamma-secretase. Here, we find that BACE1-gamma-secretase cleavages release the intracellular domain of beta2, which increases mRNA and protein levels of the pore-forming Na(v)1.1 alpha-subunit in neuroblastoma cells. Similarly, endogenous beta2 processing and Na(v)1.1 protein levels are elevated in brains of BACE1-transgenic mice and Alzheimer's disease patients with high BACE1 levels. However, Na(v)1.1 is retained inside the cells and cell surface expression of the Na(v)1 alpha-subunits and sodium current densities are markedly reduced in both neuroblastoma cells and adult hippocampal neurons from BACE1-transgenic mice. BACE1, by cleaving beta2, thus regulates Na(v)1 alpha-subunit levels and controls cell-surface sodium current densities. BACE1 inhibitors may normalize membrane excitability in Alzheimer's disease patients with elevated BACE1 activity.     

Taste receptor cells express voltage-dependent potassium channels in a cell age-specific manner

Two voltage-dependent potassium channels, KCNQ1 and KCNH2, are expressed in the taste buds and were identified as strong candidates involved in the repolarization of taste receptor cells expressing phospholipase C-beta2 and TRPM5 (beta2/M5-TRCs). In cell type-specific expression, KCNQ1 was expressed in most taste bud cells, including beta2/M5-TRCs, whereas KCNH2 was expressed in a subset of beta2/M5-TRCs with no correlation with their taste modality, such as sweet or bitter taste reception. Expression of KCNH2 was restricted to young beta2/M5-TRCs. These results suggest that taste bud cells other than beta2/M5-TRCs are depolarized by some stimuli and also that beta2/M5-TRCs have cell age-dependent molecular mechanisms of repolarization.     

Distribution and function of voltage-gated sodium channels in the nervous system

Voltage-gated sodium channels (VGSCs) are the basic ion channels for neuronal excitability, which are crucial for the resting potential and the generation and propagation of action potentials in neurons. To date, at least nine distinct sodium channel isoforms have been detected in the nervous system. Recent studies have identified that voltage-gated sodium channels not only play an essential role in the normal electrophysiological activities of neurons but also have a close relationship with neurological diseases. In this study, the latest research findings regarding the structure, type, distribution, and function of VGSCs in the nervous system and their relationship to neurological diseases, such as epilepsy, neuropathic pain, brain tumors, neural trauma, and multiple sclerosis, are reviewed in detail.