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Results for “Shal Potassium Channels”

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At least 19 recordsLinked to original sources

PubMed2003

An ER export signal accelerates the surface expression of shal potassium channels in pyloric neurons of the lobster stomatogastric ganglion.

The shal gene encoding the transient potassium current, I(A), plays important roles in shaping the firing properties of neurons in the pyloric network in the stomatogastric ganglion (STG) of the spiny lobster, Panulirus interruptus. However, when we overexpressed the shal protein in pyloric dilator (PD) neurons, the effect of increased I(A )was compensated by a parallel upregulation of the hyperpolarization activated inward current ( I(h)). In an attempt to temporally separate the overexpression of shal from the compensatory up-regulation of I(h) channels, we inserted an endoplasmic reticulum (ER) export signal sequence, FCYENE, into the shal gene. This signal sequence accelerated the surface expression of shal protein in Xenopus oocytes and PD neurons. However, the accelerated expression of shal still did not alter the firing properties of the injected neuron, suggesting that the compensatory upregulation of I(h) occurs simultaneously with the upregulation of I(A).

Amino Acid Sequence↗

PubMed2000

Expression of Shal potassium channel subunits in the adult and developing cochlear nucleus of the mouse.

The pattern of expression of potassium (K(+)) channel subunits is thought to contribute to the establishment of the unique discharge characteristics exhibited by cochlear nucleus (CN) neurons. This study describes the developmental distribution of mRNA for the three Shal channel subunits Kv4.1, Kv4.2 and Kv4.3 within the mouse CN, as assessed with in situ hybridization and RT-PCR techniques. Kv4.1 was not present in CN at any age. Kv4.2 mRNA was detectable as early as postnatal day 2 (P2) in all CN subdivisions, and continued to be constitutively expressed throughout development. Kv4.2 was abundantly expressed in a variety of CN cell types, including all of the major projection neuron classes (i.e., octopus, bushy, stellate, fusiform, and giant cells). In contrast, Kv4.3 was expressed at lower levels and by fewer cell types. Kv4.3-labeled cells were more prevalent in ventral subdivisions than in the dorsal CN. Kv4.3 expression was significantly delayed developmentally in comparison to Kv4.2, as it was detectable only after P14. Although the techniques employed in this study detect mRNA and not protein, it can be inferred from the differential distribution of Kv4 transcripts that CN neurons selectively regulate the expression of Shal K(+) channels among individual neurons throughout development.

Animals↗

PubMed2003

The role of Zn2+ in Shal voltage-gated potassium channel formation.

Voltage-gated potassium channels are formed by the tetramerization of their alpha subunits, in a process that is controlled by their conserved N-terminal T1 domains. The crystal structures of Shaker and Shaw T1 domains reveal interesting differences in structures that are contained within a highly conserved BTB/POZ domain fold. The most surprising difference is that the Shaw T1 domain contains an intersubunit Zn2+ ion that is lacking in the Shaker T1 domain. The Zn2+ coordination motif is conserved in other non-Shaker channels making this the most distinctive difference between these channels and Shaker. In this study we show that Zn2+ is an important co-factor for the tetramerization of isolated Shaw and Shal T1 domains. Addition of Zn2+ increases the amount of tetramer formed, whereas chelation of Zn2+ with phenanthroline blocks tetramerization and causes assembled tetramers to disassemble. Within an intact cell, full-length Shal subunits containing Zn2+ site mutations also fail to form functional channels, with the majority of the protein found to remain monomeric by size exclusion chromatography. Therefore, zinc-mediated tetramerization also is a physiologically important event for full-length functional channel formation.

Animals↗

PubMed2000

Viral gene transfer of dominant-negative Kv4 construct suppresses an O2-sensitive K+ current in chemoreceptor cells.

Hypoxia initiates the neurosecretory response of the carotid body (CB) by inhibiting one or more potassium channels in the chemoreceptor cells. Oxygen-sensitive K(+) channels were first described in rabbit CB chemoreceptor cells, in which a transient outward K(+) current was reported to be reversibly inhibited by hypoxia. Although progress has been made to characterize this current with electrophysiological and pharmacological tools, no attempts have been made to identify which Kv channel proteins are expressed in rabbit CB chemoreceptor cells and to determine their contribution to the native O(2)-sensitive K(+) current. To probe the molecular identity of this current, we have used dominant-negative constructs to block the expression of functional Kv channels of the Shaker (Kv1.xDN) or the Shal (Kv4.xDN) subfamilies, because members of these two subfamilies contribute to the transient outward K(+) currents in other preparations. Delivery of the constructs into chemoreceptor cells has been achieved with adenoviruses that enabled ecdysone-inducible expression of the dominant-negative constructs and reporter genes in polycistronic vectors. In voltage-clamp experiments, we found that, whereas adenoviral infections of chemoreceptor cells with Kv1.xDN did not modify the O(2)-sensitive K(+) current, infections with Kv4.xDN suppressed the transient outward current in a time-dependent manner, significantly depolarized the cells, and abolished the depolarization induced by hypoxia. Our work demonstrate that genes of the Shal K(+) channels underlie the transient outward, O(2)-sensitive, K(+) current of rabbit CB chemoreceptor cells and that this current contributes to the cell depolarization in response to low pO(2).

Adenoviridae↗

PubMed2003

Activity-independent homeostasis in rhythmically active neurons.

The shal gene encodes the transient potassium current (I(A)) in neurons of the lobster stomatogastric ganglion. Overexpression of Shal by RNA injection into neurons produces a large increase in I(A), but surprisingly little change in the neuron's firing properties. Accompanying the increase in I(A) is a dramatic and linearly correlated increase in the hyperpolarization-activated inward current (I(h)). The enhanced I(h) electrophysiologically compensates for the enhanced I(A), since pharmacological blockade of I(h) uncovers the physiological effects of the increased I(A). Expression of a nonfunctional mutant Shal also induces a large increase in I(h), demonstrating a novel activity-independent coupling between the Shal protein and I(h) enhancement. Since I(A) and I(h) influence neuronal activity in opposite directions, our results suggest a selective coregulation of these channels as a mechanism for constraining cell activity within appropriate physiological parameters.

Action Potentials↗

PubMed2005

Shal and shaker differential contribution to the K+ currents in the Drosophila mushroom body neurons.

Shaker, a voltage-dependent K+ channel, is enriched in the mushroom bodies (MBs), the locus of olfactory learning in Drosophila. Mutations in the shaker locus are known to alter excitability, neurotransmitter release, synaptic plasticity, and olfactory learning. However, a direct link of Shaker channels to MB intrinsic neuron (MBN) physiology has not been documented. We found that transcripts for shab, shaw, shaker, and shal, among which only Shaker and Shal have been reported to code for A-type currents, are present in the MBs. The electrophysiological data showed that the absence of functional Shaker channels modifies the distribution of half-inactivation voltages (V(i1/2)) in the MBNs, indicating a segregation of Shaker channels to only a subset (approximately 28%) of their somata. In harmony with this notion, we found that approximately one-fifth of MBNs lacking functional Shaker channels displayed dramatically slowed-down outward current inactivation times and reduced peak-current amplitudes. Furthermore, whereas all MBNs were sensitive to 4-aminopyridine, a nonspecific A-type current blocker, a subset of neurons (approximately 24%) displayed little sensitivity to a Shal-specific toxin. This subset of neurons displaying toxin-insensitive outward currents had more depolarized V(i1/2) values attributable to Shaker channels. Our findings provide the first direct evidence that altered Shaker channel function disrupts MBN physiology in Drosophila. To our surprise, the experimental data also indicate that Shaker channels segregate to a minor fraction of MB neuronal somata (20-30%), and that Shal channels contribute the somatic A-type current in the majority of MBNs.

4-Aminopyridine↗

PubMed1996

Inhibition of the Kv4 (Shal) family of transient K+ currents by arachidonic acid.

We have found that transient A-type currents expressed in Xenopus oocytes from members of the Kv4 family are suppressed by arachidonic acid. Currents from members of the Kv1, Kv2, and Kv3 families showed little or no inhibition by fatty acids in this expression system, although Shaker currents showed a modest increase in peak amplitude. The inhibition of Kv4 channels was not prevented by cyclo-oxygenase, lipoxygenase, or cytochrome P-450 inhibitors and was mimicked by 5,8,11,14-eicosatetraynoic acid, an arachidonic acid analog that is not metabolized by these pathways. Other unsaturated cis fatty acids with more than two double bonds produced a similar effect. In inside-out macropatches, the current was reversibly reduced > 50% by 2 mM arachidonic acid, and the inhibition developed in < 40 sec. These results suggest that, at concentrations that are likely to be physiologically relevant, arachidonic acid interacts directly with the channel or with a closely associated component. Preliminary mutagenesis of Kv4.2 channels indicates that the N terminal is not required for arachidonic acid action but that the S4-S5 loop may influence the effect.

5,8,11,14-Eicosatetraynoic Acid↗

PubMed2004

Messenger RNA and protein expression analysis of voltage-gated potassium channels in the brain of Abeta(25-35)-treated rats.

Potassium channel dysfunction has been implicated in Alzheimer's disease. In the present study, the expression of voltage-gated potassium channel (Kv) subunits in rat brain was measured after a single intracerebroventricular injection of beta-amyloid peptide 25-35 (Abeta(25-35)). After injection of Abeta, the spatial memory of rats was significantly impaired in the Morris water maze. Expression of five main Kv channel subunits (Kv1.5, Kv2.1, Kv1.4, Kv4.2, and Kv4.3) in mRNA level was assessed by using reverse transcription-polymerase chain reaction (RT-PCR). The mRNA levels of Kv2.1 and Kv1.4 were increased by 72% and 67%, respectively, in hippocampus, and Kv4.2 mRNA was increased by 58% in cortex. No other significant mRNA expression changes were found in Abeta-treated rats. The protein expression of Kv2.1, Kv1.4, and Kv4.2 was detected by using Western blotting. Kv2.1 and Kv1.4 protein levels were increased by 48% and 50%, respectively, in hippocampus of Abeta-treated rats, and Kv4.2 protein was increased by 42% in cerebral cortex. This study indicates that the expression up-regulation of Kv1.4, Kv2.1, and Kv4.2 in Abeta-induced cognitive impairment might play an important role in the pathogenesis of Alzheimer's disease.

Alzheimer Disease↗

PubMed2001

Downregulation of K(+) channel genes expression in type I diabetic cardiomyopathy.

Type I diabetic cardiomyopathy has consistently been shown to be associated with decrease of repolarising K(+) currents, but the mechanisms responsible for the decrease are not well defined. We investigated the streptozotocin (STZ) rat model of type I diabetes. We utilized RNase protection assay and Western blot analysis to investigate the message expression and protein density of key cardiac K(+) channel genes in the diabetic rat left ventricular (LV) myocytes. Our results show that message and protein density of Kv2.1, Kv4.2, and Kv4.3 are significantly decreased as early as 14 days following induction of type I diabetes in the rat. The results demonstrate, for the first time, that insulin-deficient type I diabetes is associated with early downregulation of the expression of key cardiac K(+) channel genes that could account for the depression of cardiac K(+) currents, I(to-f) and I(to-s). These represent the main electrophysiological abnormality in diabetic cardiomyopathy and is known to enhance the arrhythmogenecity of the diabetic heart. The findings also extend the extensive list of gene expression regulation by insulin.

Animals↗

PubMed2001

Modulation of outward potassium currents in aligned cultures of neonatal rat ventricular myocytes during phorbol ester-induced hypertrophy.

Protein kinase C-stimulating phorbol esters induce a strong hypertrophic response when applied in vitro to cardiac ventricular myocytes. The aim of this study was to determine if this in vitro model of hypertrophy is associated with changes in the expression of voltage-gated K(+)channels. Myocytes were isolated from 3--4-day-old neonatal rats and cultured on aligned collagen thin gels. Membrane currents were measured with the use of the whole-cell arrangement of the patch clamp technique and the expression levels of the Kv1.4, Kv4.2 and Kv2.1 alpha subunits quantified using Western blot analysis. Voltage steps positive to -30 mV resulted in the activation of both a transient (I(to)) and a sustained (I(sus)) component of outward K(+)current in the aligned myocytes. Overnight exposure to phorbol 12-myristate 13-acetate (PMA) caused a 55% increase in myocyte size and a three-fold reduction in the peak amplitude of I(to). No differences in the half-maximal voltages required for activation and steady-state inactivation were observed between I(to)measured in control and PMA-treated myocytes. In contrast, PMA treatment resulted in a 62% increase in a tetraethylammonium-sensitive component of I(sus)(TEA-I(sus)) and was associated with the appearance of a slow component of current decay. Expression levels of the Kv1.4 and Kv4.2 alpha subunits were strongly depressed in the hypertrophic myocytes, while the density of the Kv2.1 alpha subunit was enhanced. PMA-induced changes in the Kv alpha subunits were partially prevented through inhibition of the mitogen-activated protein kinase (MAPK) pathway. Thus, PMA-induced hypertrophy of cultured ventricular myocytes is associated with an altered expression of voltage-gated K(+)channels.

Animals↗

PubMed2001

Voltage-gated K+ channels in chemoreceptor sensory neurons of rat petrosal ganglion.

A subpopulation of sensory neurons in the petrosal ganglion transmits information between peripheral chemoreceptors (glomus cells) in the carotid body and relay neurons in the nucleus of the solitary tract. Expression of voltage-gated K+ channels in these neurons was characterized by immunohistochemical localization. Five members of the Kv1 family, Kv1.1, Kv1.2, Kv1.4, Kv1.5 and Kv1.6 and members of two other families, Kv2.1 and Kv4.3, were identified in over 90% of the chemoreceptor neurons. Although the presence of these channel proteins was consistent throughout the population, individual neurons showed considerable variation in K+ current profiles.

Animals↗

PubMed2001

Spatial alterations of Kv channels expression and K(+) currents in post-MI remodeled rat heart.

OBJECTIVE: The hypothesis being tested in the present study is that increased anisotropic properties occurs in the remodeled post-infarction heart due to spatial alterations in Kv channels expression and K(+) currents of the remodeled myocardium. METHODS: Three to 4 weeks post myocardial infarction (MI) in the rat, we measured the two components of the outward K(+) current, I(to-fast (f)) and I(to-slow(s)) in the epicardium (epi) and endocardium (endo) of noninfarcted remodeled left ventricle (LV) using patch clamp techniques. Alterations in mRNA and/or protein levels of potassium channel genes Kv1.4, Kv1.5, Kv2.1, Kv4.2 and Kv4.3 were measured in epi, midmyocardium (mid), and endo regions of LV and in the right ventricle (RV). RESULTS: In sham operated rat heart, the density of I(to-f) was 2.3 times greater in epi compared to endo myocytes. In post-MI heart, the density of I(to-f) and I(to-s) decreased to a similar degree in LV epi and endo but the difference in I(to-f) density between epi and endo persisted. The mRNA and/or protein levels of Kv1.4, Kv2.1, Kv4.2 and Kv4.3 but not Kv1.5 decreased to a varying extent in different regions of LV but not in RV of post-MI heart. CONCLUSIONS: Our results suggest that regional downregulation of Kv channels expression and density of K(+) currents can be a significant determinant of increased spatial electrophysiological heterogeneity and contribute to increased electrical instability of the post-MI heart.

Analysis of Variance↗

PubMed2003

Differential association of the auxiliary subunit Kvbeta2 with Kv1.4 and Kv4.3 K+ channels.

Kvbeta2 subunits associate with Kv1 and Kv4 K+ channels, but the basis of preferential association is not understood. For example, detergent resistance suggests stronger auxiliary subunit association with Kv4.2 than with Kv1.2, but Kvbeta2 preferentially localizes with the latter channels in brain. Here we examine the interaction of Kvbeta2 with two native binding partners in brain: Kv4.3 and Kv1.4. We show that the auxiliary subunit binds more efficiently to Kv1.4 than to Kv4.3 in mammalian cells. However, preexisting Kvbeta2 complexes with Kv1.4 and Kv4.3 have similar detergent sensitivity. Thus, preferential steady state binding may reflect a difference in initial association rather than stability. We also find that that the cytoplasmic C-terminus of Kv4.3 inhibits Kvbeta2 association. Apparently, a region proximal to the Kv4.3 pore contributes to the inefficient auxiliary subunit interaction that produces preferential binding of Kvbeta2 to Kv1 channels.

Binding Sites↗

PubMed2000

Differential targeting of Shaker-like potassium channels to lipid rafts.

Ion channel targeting within neuronal and muscle membranes is an important determinant of electrical excitability. Recent evidence suggests that there exists within the membrane specialized microdomains commonly referred to as lipid rafts. These domains are enriched in cholesterol and sphingolipids and concentrate a number of signal transduction proteins such as nitric-oxide synthase, ligand-gated receptors, and multiple protein kinases. Here, we demonstrate that the voltage-gated K(+) channel Kv2.1, but not Kv4.2, targets to lipid rafts in both heterologous expression systems and rat brain. The Kv2.1 association with lipid rafts does not appear to involve caveolin. Depletion of cellular cholesterol alters the buoyancy of the Kv2.1 associated rafts and shifts the midpoint of Kv2.1 inactivation by nearly 40 mV without affecting peak current density or channel activation. The differential targeting of Kv channels to lipid rafts represents a novel mechanism both for the subcellular sorting of K(+) channels to regions of the membrane rich in signaling complexes and for modulating channel properties via alterations in lipid content.

Animals↗

PubMed1999

Kvbeta1.2 subunit coexpression in HEK293 cells confers O2 sensitivity to kv4.2 but not to Shaker channels.

Voltage-gated K+ (KV) channels are protein complexes composed of ion-conducting integral membrane alpha subunits and cytoplasmic modulatory beta subunits. The differential expression and association of alpha and beta subunits seems to contribute significantly to the complexity and heterogeneity of KV channels in excitable cells, and their functional expression in heterologous systems provides a tool to study their regulation at a molecular level. Here, we have studied the effects of Kvbeta1.2 coexpression on the properties of Shaker and Kv4.2 KV channel alpha subunits, which encode rapidly inactivating A-type K+ currents, in transfected HEK293 cells. We found that Kvbeta1.2 functionally associates with these two alpha subunits, as well as with the endogenous KV channels of HEK293 cells, to modulate different properties of the heteromultimers. Kvbeta1.2 accelerates the rate of inactivation of the Shaker currents, as previously described, increases significantly the amplitude of the endogenous currents, and confers sensitivity to redox modulation and hypoxia to Kv4.2 channels. Upon association with Kvbeta1.2, Kv4.2 can be modified by DTT (1,4 dithiothreitol) and DTDP (2,2'-dithiodipyridine), which also modulate the low pO2 response of the Kv4.2+beta channels. However, the physiological reducing agent GSH (reduced glutathione) did not mimic the effects of DTT. Finally, hypoxic inhibition of Kv4.2+beta currents can be reverted by 70% in the presence of carbon monoxide and remains in cell-free patches, suggesting the presence of a hemoproteic O2 sensor in HEK293 cells and a membrane-delimited mechanism at the origin of hypoxic responses. We conclude that beta subunits can modulate different properties upon association with different KV channel subfamilies; of potential relevance to understanding the molecular basis of low pO2 sensitivity in native tissues is the here described acquisition of the ability of Kv4. 2+beta channels to respond to hypoxia.

2,2'-Dipyridyl↗

PubMed2003

A conserved domain in axonal targeting of Kv1 (Shaker) voltage-gated potassium channels.

Axonal voltage-gated potassium (Kv1) channels regulate action-potential invasion and hence transmitter release. Although evolutionarily conserved, what mediates their axonal targeting is not known. We found that Kv1 axonal targeting required its T1 tetramerization domain. When fused to unpolarized CD4 or dendritic transferrin receptor, T1 promoted their axonal surface expression. Moreover, T1 mutations eliminating Kvbeta association compromised axonal targeting, but not surface expression, of CD4-T1 fusion proteins. Thus, proper association of Kvbeta with the Kv1 T1 domain is essential for axonal targeting.

Amino Acid Motifs↗

PubMed2000

KChAP as a chaperone for specific K(+) channels.

The concept of chaperones for K(+) channels is new. Recently, we discovered a novel molecular chaperone, KChAP, which increased total Kv2.1 protein and functional channels in Xenopus oocytes through a transient interaction with the Kv2.1 amino terminus. Here we report that KChAP is a chaperone for Kv1.3 and Kv4.3. KChAP increased the amplitude of Kv1.3 and Kv4.3 currents without affecting kinetics or voltage dependence, but had no such effect on Kv1.1, 1.2, 1.4, 1.5, 1.6, and 3.1 or Kir2.2, HERG, or KvLQT1. Although KChAP belongs to a family of proteins that interact with transcription factors, upregulation of channel currents was not blocked by the transcription inhibitor actinomycin D. A 98-amino acid fragment of KChAP binds to the channel and is indistinguishable from KChAP in its enhancement of Kv4.3 current and protein levels. Using a KChAP antibody, we have coimmunoprecipitated KChAP with Kv2.1 and Kv4.3 from heart. We propose that KChAP is a chaperone for specific Kv channels and may have this function in cardiomyocytes where Kv4.3 produces the transient outward current, I(to).

Amino Acid Sequence↗

PubMed1999

Expression of voltage-dependent K(+) channel genes in mesenteric artery smooth muscle cells.

Molecular basis of native voltage-dependent K(+) (Kv) channels in smooth muscle cells (SMCs) from rat mesenteric arteries was investigated. The whole cell patch-clamp study revealed that a 4-aminopyridine-sensitive delayed rectifier K(+) current (I(K)) was the predominant K(+) conductance in these cells. A systematic screening of the expression of 18 Kv channel genes using RT-PCR technique showed that six I(K)-encoding genes (Kv1.2, Kv1.3, Kv1.5, Kv2.1, Kv2.2, and Kv3.2) were expressed in mesenteric artery. Although no transient outward Kv currents (I(A)) were recorded in the studied SMCs, transcripts of multiple I(A)-encoding genes, including Kv1.4, Kv3.3, Kv3.4, Kv4.1, Kv4.2, and Kv4.3 as well as I(A)-facilitating Kv beta-subunits (Kvbeta1, Kvbeta2, and Kvbeta3), were detected in mesenteric arteries. Western blot analysis demonstrated that four I(K)-related Kv channel proteins (Kv1.2, Kv1. 3, Kv1.5, and Kv2.1) were detected in mesenteric artery tissues. The presence of Kv1.2, Kv1.3, Kv1.5, and Kv2.1 channel proteins in isolated SMCs was further confirmed by immunocytochemistry study. Our results suggest that the native I(K) in rat mesenteric artery SMCs might be generated by heteromultimerization of Kv genes.

4-Aminopyridine↗