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Results for “Delayed Rectifier Potassium Channels”

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PubMed1999

Subunit assembly and domain analysis of electrically silent K+ channel alpha-subunits of the rat Kv9 subfamily.

Alpha-subunits of the voltage-gated potassium channel (Kv) subfamily Kv9 show no channel activity after homomultimeric expression in heterologous expression systems. This report shows that heteromultimeric expression of rKv9.1 and rKv9.3 specifically suppresses the currents mediated by alpha-subunits of the Kv2 and Kv3 subfamilies but does not affect currents mediated by alpha-subunits of the Kv1 and Kv4 subfamilies. To understand the molecular basis of the electrical silence of Kv9 homomultimeric channels, crucial functional domains (amino and carboxy terminus, S4 segment, and pore region) were exchanged between Kv9 alpha-subunits and rKv1.3. Electrophysiological studies of these chimeras revealed that the pore region is involved in determining the nonconductive behavior of homomultimeric Kv9 channels. This analysis was extended by protein interaction assays, aiming to identify the region of Kv9 subunits responsible for the specific suppression of rKv2.1- and rKv3.4-mediated currents. We could show that the amino-terminal domain of Kv9 alpha-subunits does not support homomultimeric assembly but interacts specifically with the rKv2.1 amino-terminal region. Conversely, the specific intersubfamily assembly of rKv3.4 with rKv9.1 or rKv9.3 is governed by the hydrophobic core and not the amino-terminal domain.

Amino Acid Sequence↗

PubMed2002

O2-sensitive K+ channels in immortalised rat chromaffin-cell-derived MAH cells.

The regulation of K(+) channels by O(2) levels is a key link between hypoxia and neurotransmitter release in neuroendocrine cells. Here, we examined the effects of hypoxia on K(+) channels in the immortalised v-myc, adrenal-derived HNK1(+) (MAH) cell line. MAH cells possess a K(+) conductance that is sensitive to Cd(2+), iberiotoxin and apamin, and which is inhibited by ~24 % when exposed to a hypoxic perfusate (O(2) tension 20 mmHg). This conductance was attributed to high-conductance Ca(2+)-activated K(+) (BK) and small-conductance Ca(2+)-activated K(+) (SK) channels, which are major contributors to the O(2)-sensitive K(+) conductance in adrenomedullary chromaffin cells. Under low [Ca(2+)](i) conditions that prevented activation of Ca(2+)-dependent K(+) conductances, a rapidly activating and slowly inactivating K(+) conductance, sensitive to both TEA and 4-aminopyridine (4-AP), but insensitive to 100 nM charybdotoxin (CTX), was identified. This current was also reduced (by ~25 %) when exposed to hypoxia. The hypoxia-sensitive component of this current was greatly attenuated by 10 mM 4-AP, but was only slightly reduced by 10 mM TEA. This suggests the presence of delayed-rectifier O(2)-sensitive channels comprising homomultimeric Kv1.5 or heteromultimeric Kv1.5/Kv1.2 channel subunits. The presence of both Kv1.5 and Kv1.2 alpha-subunits was confirmed using immunocytochemical techniques. We also demonstrated that these K(+) channel subunits are present in neonatal rat adrenomedullary chromaffin cells in situ. These data indicate that MAH cells possess O(2)-sensitive K(+) channels with characteristics similar to those observed previously in isolated chromaffin cells, and therefore provide an excellent model for examining the cellular mechanisms of O(2) sensing in adrenomedullary chromaffin cells.

Adrenal Glands↗

PubMed1998

Functional and molecular differences between voltage-gated K+ channels of fast-spiking interneurons and pyramidal neurons of rat hippocampus.

We have examined gating and pharmacological characteristics of somatic K+ channels in fast-spiking interneurons and regularly spiking principal neurons of hippocampal slices. In nucleated patches isolated from basket cells of the dentate gyrus, a fast delayed rectifier K+ current component that was highly sensitive to tetraethylammonium (TEA) and 4-aminopyridine (4-AP) (half-maximal inhibitory concentrations <0.1 mM) predominated, contributing an average of 58% to the total K+ current in these cells. By contrast, in pyramidal neurons of the CA1 region a rapidly inactivating A-type K+ current component that was TEA-resistant prevailed, contributing 61% to the total K+ current. Both types of neurons also showed small amounts of the K+ current component mainly found in the other type of neuron and, in addition, a slow delayed rectifier K+ current component with intermediate properties (slow inactivation, intermediate sensitivity to TEA). Single-cell RT-PCR analysis of mRNA revealed that Kv3 (Kv3.1, Kv3.2) subunit transcripts were expressed in almost all (89%) of the interneurons but only in 17% of the pyramidal neurons. In contrast, Kv4 (Kv4.2, Kv4.3) subunit mRNAs were present in 87% of pyramidal neurons but only in 55% of interneurons. Selective block of fast delayed rectifier K+ channels, presumably assembled from Kv3 subunits, by 4-AP reduced substantially the action potential frequency in interneurons. These results indicate that the differential expression of Kv3 and Kv4 subunits shapes the action potential phenotypes of principal neurons and interneurons in the cortex.

4-Aminopyridine↗

PubMed1998

Beta-adrenoceptor activation and PKA regulate delayed rectifier K+ channels of vascular smooth muscle cells.

Macroscopic 4-aminopyridine (4-AP)-sensitive, delayed rectifier K+ current of vascular smooth muscle cells is increased during beta-adrenoceptor activation with isoproterenol via a signal transduction pathway involving adenylyl cyclase and cAMP-dependent protein kinase (PKA) (Aiello, E. A., M. P. Walsh, and W. C. Cole. Am. J. Physiol. 268 (Heart Circ. Physiol. 37): H926-H934, 1995.). In this study, we identified the single delayed rectifier K+ (KDR) channel(s) of rabbit portal vein myocytes affected by treatment with isoproterenol or the catalytic subunit of PKA. 4-AP-sensitive KDR channels of 15.3 +/- 0.6 pS (n = 5) and 14.8 +/- 0.6 pS (n = 5) conductance, respectively, were observed in inside-out (I-O) and cell-attached (C-A) membrane patches in symmetrical KCl recording conditions. The kinetics of activation (time constant of 10.7 +/- 3. 02 ms) and inactivation (fast and slow time constants of 0.3 and 2.5 s, respectively) of ensemble currents produced by these channels mimicked those reported for inactivating, 4-AP-sensitive whole cell KDR current of vascular myocytes. Under control conditions, the open probability (NPo) of KDR channels of C-A membrane patches at -40 mV was 0.014 +/- 0.005 (n = 8). Treatment with 1 microM isoproterenol caused a significant, approximately threefold increase in NPo to 0. 041 +/- 0.02 (P < 0.05). KDR channels of I-O patches exhibited rundown after approximately 5 min, which was not affected by ATP (5 mM) in the bath solution. Treatment with the purified catalytic subunit of PKA (50 nM; 5 mM ATP) restored KDR channel activity and caused NPo to increase from 0.011 +/- 0.003 to 0.138 +/- 0.03 (P < 0. 05; n = 11). These data indicate that small-conductance, 15-pS KDR channels are responsible for inactivating the macroscopic delayed rectifier K+ current of rabbit portal vein myocytes and that the activity of these channels is enhanced by a signal transduction mechanism involving beta-adrenoceptors and phosphorylation by PKA at a membrane potential consistent with that observed in the myocytes in situ.

4-Aminopyridine↗

PubMed1995

The voltage-dependent K+ channel (Kv1.5) cloned from rabbit heart and facilitation of inactivation of the delayed rectifier current by the rat beta subunit.

We have isolated a cDNA coding for a delayed rectifier K+ channel (RBKV1.5) from rabbit heart. The amino acid sequence of RBKV1.5 displays a homology to that of other K+ channels of Kv1.5 class. Overall amino acid identity between RBKV1.5 channel and Kv1.5 channel of other species is about 85%. RNA blot analysis revealed the expression of the primary transcript in various rabbit tissues, at the highest level in both the atrium and ventricle. When expressed in Xenopus oocytes, RBKV1.5 current showed a delayed rectifier type characteristics, which was converted to rapidly inactivating currents upon coexpression with a beta subunit.

Amino Acid Sequence↗

PubMed2001

Interaction of a toxin from the scorpion Tityus serrulatus with a cloned K+ channel from squid (sqKv1A).

A toxin from the scorpion Tityus serrulatus (TsTX-Kalpha) blocks native squid K(+) channels and their cloned counterpart, sqKv1A, at pH 8 ((native)K(d) approximately 20 nM; (sqKv1A)K(d) approximately 10 nM). In both cases, decreasing the pH below 7.0 significantly diminishes the TsTX-Kalpha effect (pK = 6.6). In the cloned squid channel, the pH dependence of the block is abolished by a single point mutation (H351G), and no change in toxin affinity was observed at higher pH values (pH > or =8.0). To further investigate the TsTX-Kalpha-sqKv1A interaction, the three-dimensional structure of TsTX-Kalpha was determined in solution by NMR spectroscopy, and a model of the TsTX-Kalpha-sqKv1A complex was generated. As found for other alpha-K toxins such as charybdotoxin (CTX), site-directed mutagenesis at toxin residue K27 (K27A, K27R, and K27E) significantly reduced the toxin's affinity for sqKv1A channels. This is consistent with the TsTX-Kalpha-sqKv1A model reported here, which has K27 of the toxin inserted into the ion conduction pathway of the K(+) channel. This toxin-channel model also illustrates a possible mechanism for the pH-dependent block whereby lysine residues from TsTX-Kalpha (K6 and K23) are repelled by protonated H351 on sqKv1A at low pH.

Amino Acid Sequence↗

PubMed2001

Members of the Kv1 and Kv2 voltage-dependent K(+) channel families regulate insulin secretion.

In pancreatic beta-cells, voltage-dependent K(+) (Kv) channels are potential mediators of repolarization, closure of Ca(2+) channels, and limitation of insulin secretion. The specific Kv channels expressed in beta-cells and their contribution to the delayed rectifier current and regulation of insulin secretion in these cells are unclear. High-level protein expression and mRNA transcripts for Kv1.4, 1.6, and 2.1 were detected in rat islets and insulinoma cells. Inhibition of these channels with tetraethylammonium decreased I(DR) by approximately 85% and enhanced glucose-stimulated insulin secretion by 2- to 4-fold. Adenovirus-mediated expression of a C-terminal truncated Kv2.1 subunit, specifically eliminating Kv2 family currents, reduced delayed rectifier currents in these cells by 60-70% and enhanced glucose-stimulated insulin secretion from rat islets by 60%. Expression of a C-terminal truncated Kv1.4 subunit, abolishing Kv1 channel family currents, reduced delayed rectifier currents by approximately 25% and enhanced glucose-stimulated insulin secretion from rat islets by 40%. This study establishes that Kv2 and 1 channel homologs mediate the majority of repolarizing delayed rectifier current in rat beta-cells and that antagonism of Kv2.1 may prove to be a novel glucose-dependent therapeutic treatment for type 2 diabetes.

1-Methyl-3-isobutylxanthine↗

PubMed2004

Disruption of pancreatic beta-cell lipid rafts modifies Kv2.1 channel gating and insulin exocytosis.

In pancreatic beta-cells, the predominant voltage-gated Ca(2+) channel (Ca(V)1.2) and K(+) channel (K(V)2.1) are directly coupled to SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor) proteins. These SNARE proteins modulate channel expression and gating and closely associate these channels with the insulin secretory vesicles. We show that K(V)2.1 and Ca(V)1.2, but not K(V)1.4, SUR1, or Kir6.2, target to specialized cholesterol-rich lipid raft domains on beta-cell plasma membranes. Similarly, the SNARE proteins syntaxin 1A, SNAP-25, and VAMP-2, but not Munc-13-1 or n-Sec1, are associated with lipid rafts. Disruption of the lipid rafts by depleting membrane cholesterol with methyl-beta-cyclodextrin shunts K(V)2.1, Ca(V)1.2, and SNARE proteins out of lipid rafts. Furthermore, methyl-beta-cyclodextrin inhibits K(V)2.1 but not Ca(V)1.2 channel activity and enhances single-cell exocytic events and insulin secretion. Membrane compartmentalization of ion channels and SNARE proteins in lipid rafts may be critical for the temporal and spatial coordination of insulin release, forming what has been described as the excitosome complex.

Animals↗

PubMed2002

External pore collapse as an inactivation mechanism for Kv4.3 K+ channels.

Kv4 channels are thought to lack a C-type inactivation mechanism (collapse of the external pore) and to inactivate as a result of a concerted action of cytoplasmic regions of the channel. To investigate whether Kv4 channels have outer pore conformational changes during the inactivation process, the inactivation properties of Kv4.3 were characterized in 0 mM and in 2 mM external K+ in whole-cell voltage-clamp experiments. Removal of external K+ increased the inactivation rates and favored cumulative inactivation by repetitive stimulation. The reduction in current amplitude during repetitive stimulation and the faster inactivation rates in 0 mM external K+ were not due to changes in the voltage dependence of channel opening or to internal K+ depletion. The extent of the collapse of the K+ conductance upon removal of external K+ was more pronounced in NMG+-than in Na+-containing solutions. The reduction in the current amplitude during cumulative inactivation by repetitive stimulation is not associated with kinetic changes, suggesting that it is due to a diminished number of functional channels with unchanged gating properties. These observations meet the criteria for a typical C-type inactivation, as removal of external K+ destabilizes the conducting state, leading to the collapse of the pore. A tentative model is presented, in which K+ bound to high-affinity K+-binding sites in the selectivity filter destabilizes an outer neighboring K+ modulatory site that is saturated at approximately 2 mM external K+. We conclude that Kv4 channels have a C-type inactivation mechanism and that previously reported alterations in the inactivation rates after N- and C- termini mutagenesis may arise from secondary changes in the electrostatic interactions between K+-binding sites in the selectivity filter and the neighboring K+-modulatory site, that would result in changes in its K+ occupancy.

Cell Line↗

PubMed2000

Sulfhydryl modification affects coronary artery tension by changing activity of delayed rectifier K+ current.

It has been reported that a change in the cellular redox state may be involved in the regulation of vascular tone, but the underlying mechanism is not fully understood. The present study was designed to investigate the cellular effect of sulfhydryl modifying agents in the coronary artery of rabbit using the tension measurement and whole cell clamping method. The application of diamide, a sulfhydryl oxidizing agent, relaxed the endothelium denuded coronary arteries in a dose dependent manner. The fact that this diamide-induced relaxation was significantly attenuated by a pretreatment of 4-AP, and the coronary arteries precontracted with 100 mM K+ instead of histamine, suggests the involvement of 4-AP sensitive K+ channels in the diamide-induced relaxation of coronary arteries. Whole cell patch clamp studies revealed that the 4-AP sensitive IdK was significantly enhanced by the membrane permeant oxidizing agents, diamide and DTDP, and were reversed by subsequent exposure to the reducing agent, DTT. Neither the membrane impermeant oxidizing or reducing agents, GSSG or GSH, had any effect on the activity of IdK, indicating that intracellular sulfhydryl modification is critical for modulating IdK activity. The Diamide failed to significantly alter the voltage dependence of the activation and inactivation parameters, and did not change the inactivation process, suggesting that diamide increases the number of functional channels without altering their gating properties. Since IdK has been believed to play an important role in regulating membrane potential and arterial tone, our results about the effect of sulfhydryl modifying agents on coronary arterial tone and IdK activity should help understand the pathophysiology of the diseases, where oxidative damage has been implicated.

Animals↗

PubMed1996

Generation and characterization of subtype-specific monoclonal antibodies to K+ channel alpha- and beta-subunit polypeptides.

Molecular characterization of mammalian voltage-sensitive K+ channel genes and their expression became possible with the cloning of the Shaker locus of Drosophila. However, analysis of the expression patterns and subunit composition of native K+ channel protein complexes requires immunological probes specific for the individual K+ channel gene products expressed in excitable tissue. Here, we describe the generation and characterization of monoclonal antibodies (mAbs) against eight distinct mammalian K+ channel polypeptides; the Kv1.1, Kv1.2, Kv1.4, Kv1.5 and Kv1.6 Shaker-related alpha-subunits, the Kv2.1 Shab-related alpha-subunit, and the Kv beta 1 and Kv beta 2 beta-subunits. We characterized the subtype-specificity of these mAbs against native K+ channels in mammalian brain and against recombinant K+ channels expressed in transfected mammalian cells. In addition, we used these mAbs to investigate the cellular and subcellular distribution of the corresponding polypeptides in rat cerebral cortex, as well as their expression levels across brain regions.

Amino Acid Sequence↗

PubMed2004

Single-channel recordings of a rapid delayed rectifier current in adult mouse ventricular myocytes: basic properties and effects of divalent cations.

The rapidly delayed rectifier current (I(Kr)) has been described in ventricular myocytes isolated from many species, as well as from neonatal mice. However, whether I(Kr) is present in the adult mouse heart remains controversial. We used cell-attached patch-clamp recording in symmetrical K(+) solutions to assess the presence and behaviour of single I(Kr) channels in adult mouse cardiomyocytes (mI(Kr)). Of 314 patches, 158 (50.1%) demonstrated mI(Kr) currents as compared with 131 (42.3%) for the I(K1) channel. Single mI(Kr) channel activity was rarely observed at potentials positive to -10 mV. The slope conductance at negative potentials was 12 pS. Upon repolarization, ensemble-averaged mI(Kr) showed slow deactivation with a biexponential time course. A selective I(Kr) blocker, E-4031 (1 microm), completely blocked mI(Kr) channel activity. Extracellular Ca(2+) and Mg(2+) at physiological concentrations shifted the activation by approximately 30 mV, accelerated deactivation kinetics, prolonged long-closed time, and reduced open probability without affecting single-channel conductance, suggesting a direct channel-blocking effect in addition to well-recognized voltage shifts. HERG subunits expressed in Chinese hamster ovary cells produced channels with properties similar to those of mI(Kr), except for the more-negative activation of the HERG channels. Despite the abundant expression of mI(Kr), single-channel events were rarely observed during action-potential clamp and 5 microm E-4031 had no detectable effect on the action potential parameters, confirming that mI(Kr) plays at best a minor role in repolarization of adult mouse cardiomyocytes, probably because the modulatory effects of divalent cations prevent significant mI(Kr) opening under physiological conditions.

Animals↗

PubMed2003

Papaverine blocks hKv1.5 channel current and human atrial ultrarapid delayed rectifier K+ currents.

Papaverine, 1-[(3,4-dimethoxyphenyl)methyl]-6,-7-dimethoxyisoquinoline, has been used as a vasodilator agent and a therapeutic agent for cerebral vasospasm, renal colic, and penile impotence. We examined the effects of papaverine on a rapidly activating delayed rectifier K(+) channel (hKv1.5) cloned from human heart and stably expressed in Ltk(-) cells as well as a corresponding K(+) current (the ultrarapid delayed rectifier, I(Kur)) in human atrial myocytes. Using the whole cell configuration of the patch-clamp technique, we found that papaverine inhibited hKv1.5 current in a time- and voltage-dependent manner with an IC(50) value of 43.4 microM at +60 mV. Papaverine accelerated the kinetics of the channel inactivation, suggesting the blockade of open channels. Papaverine (100 microM) also blocked I(Kur) in human atrial myocytes. These results indicate that papaverine blocks hKv1.5 channels and native hKv1.5 channels in a concentration-, voltage-, state-, and time-dependent manner. This interaction suggests that papaverine could alter cardiac excitability in vivo.

Animals↗

PubMed2000

Potassium (K+) channel expression in basal forebrain cholinergic neurons.

Basal forebrain cholinergic neurons (BFCN) are depleted early in the course of Alzheimer's disease (AD). BFCN voltage-gated K(+) channels regulate acetylcholine release and may play a role in BFCN neurodegeneration. Neuronal voltage-gated K(+) channels are heterotetrameric assemblies of K(v) and accessory subunits. Currently, there is no available information about the K(v) proteins expressed in BFCN. Immunohistochemical techniques were used to investigate the expression of specific K(v) subunits in rat brain BFCN. Our results showed that BFCN express both K(v)3.1 and K(v)2.1 subunits. However, the K(v)2.1 subunit showed a wider distribution in noncholinergic neurons than the K(v)3.1 subunit. K(v)3.1 and K(v)2.1 immunostaining was noticeable not only in neuronal cell bodies but also in the dendritic ramifications of these neurons. Insofar as the K(v)3.1 subunit has been classically associated with "fast-spiking neurons" and BFCN have low firing rates and long-duration action potentials, K(v)3.1 subunits may have functions other than facilitating high-frequency firing in BFCN.

Acetylcholine↗

PubMed2003

Effect of androgen deficiency on mouse ventricular repolarization.

We previously demonstrated that female mouse ventricles have longer action potential durations (APDs) than males. This delayed repolarization results from a lower current density of the ultrarapid delayed rectifier K(+) current (I(K,ur)) and a lower expression level of its underlying K(+) channel (Kv1.5). To evaluate whether this sex difference could be attributable to the action of male sex hormones, we studied the effect of androgen deficiency on ventricular repolarization. We compared cardiac electrophysiological properties in castrated (orchiectomized; ORC) and control (CTL) male mice. Q-Tc intervals as well as APDs measured at 20 %, 50 % and 90 % of repolarization were all significantly longer in ORC than in CTL. The current density of I(K,ur) was significantly lower in ORC than in CTL (at +50 mV, ORC: 29 +/- 4 pA pF(-1), n = 25; CTL: 48 +/- 5 pA pF(-1), n = 17; P = 0.006). In contrast, all the other K(+) currents present in mouse ventricular myocytes were comparable between ORC and CTL. Moreover, results of Western blot analysis showed a lower expression level of Kv1.5 protein in ORC but no difference between the two groups for the other K(+) channels studied. This study demonstrates that androgen deficiency leads to a reduction in the density of I(K,ur) and Kv1.5 in mouse ventricle, and consequently, to prolongation of APD and Q-Tc interval. In conclusion, these findings strongly suggest that male sex hormones contribute to the sex difference that we previously reported in cardiac repolarization in adult mouse heart.

Action Potentials↗

PubMed2003

Electrophysiological differentiation of new neurons in the olfactory bulb.

The subventricular zone produces neuroblasts that migrate to the olfactory bulb (OB) and differentiate into interneurons throughout postnatal life (Altman and Das, 1966; Hinds, 1968; Altman, 1969; Kishi et al., 1990; Luskin, 1993; Lois and Alvarez-Buylla, 1994). Although such postnatally generated interneurons have been characterized morphologically, their physiological differentiation has not been thoroughly described. Combining retroviral-mediated labeling of newly generated neurons with patch-clamp electrophysiology, we demonstrated that soon after new cells enter the layers of the olfactory bulb, they display voltage-dependent currents typical of more mature neurons. We also show that these "newcomers" express functional GABA and glutamate receptor channels, respond synaptically to stimulation of the olfactory nerve, and may establish both axodendritic and dendrodendritic synaptic contacts within the olfactory bulb. These data provide a basic description of the physiology of newly generated cells in the OB and show that such new cells are functional neurons that synaptically integrate into olfactory bulb circuitry soon after their arrival.

Animals↗

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↗

PubMed1999

Role of the outward delayed rectifier K+ current in ceramide-induced caspase activation and apoptosis in cultured cortical neurons.

We studied the novel hypothesis that an up-modulation of channels for outward delayed rectifier K+ current (I(K)) plays a key role in ceramide-induced neuronal apoptosis. Exposure for 6-10 h to the membrane-permeable C2-ceramide (25 microM) or to sphingomyelinase (0.2 unit/ml), but not to the inactive ceramide analogue C2-dihydroceramide (25 microM), enhanced the whole-cell I(K) current without affecting the transient A-type K+ current and increased caspase activity, followed by neuronal apoptosis 24 h after exposure onset. Tetraethylammonium (TEA) or 4-chloro-N,N-diethyl-N-heptylbenzenebutanaminium tosylate (clofilium), at concentrations inhibiting I(K), attenuated the C2-ceramide-induced caspase-3-like activation as well as neuronal apoptosis. Raising extracellular K+ to 25 mM similarly blocked the C2-ceramide-induced cell death; the neuroprotection by 25 mM K+ or TEA was not eliminated by blocking voltage-gated Ca2+ channels. An inhibitor of tyrosine kinases, herbimycin A (10 nM) or lavendustin A (0.1-1 microM), suppressed I(K) enhancement and/or apoptosis induced by C2-ceramide. It is suggested that ceramide-induced I(K) current enhancement is mediated by tyrosine phosphorylation and plays a critical role in neuronal apoptosis.

Animals↗