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K+ Currents (k+ + current)
Kinds of K+ Currents Selected AbstractsBLOCK OF Na+ AND K+ CURRENTS IN RAT VENTRICULAR MYOCYTES BY QUINACAINOL AND QUINIDINECLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 1-2 2005Michael K Pugsley SUMMARY 1.,The electrophysiological actions of quinacainol were investigated on sodium (INa), transient outward (ito) and sustained-outward plateau (iKsus) potassium currents in rat isolated cardiac myocytes using the whole-cell patch-clamp technique and compared with quinidine. 2.,Quinacainol blocked sodium currents in a concentration-dependent manner and with a potency similar to that of quinidine (mean (±SEM) EC50 50 ± 12 vs 95 ± 25 µmol/L for quinidine and quinacainol, respectively). However, quinacainol had a considerably prolonged onset and recovery from block compared with quinidine. 3.,Neither quinacainol nor quinidine significantly changed the steady state voltage dependence of activation of sodium currents. Quinidine produced a hyperpolarizing shift in the voltage dependence for sodium current inactivation, but no such shift was observed with quinacainol at doses that produced a substantial current block. 4.,Although quinacainol did not effectively block voltage-dependent potassium currents, even at concentrations as high as 1.5 mmol/L, quinidine, at a half-maximal sodium channel-blocking concentration, reduced peak ito current amplitude, increased the rate of inactivation of ito and blocked iKsus. 5.,These results indicate that quinacainol, a quinidine analogue, blocks sodium currents in cardiac myocytes with little effect on ito or iKsus potassium currents, which suggests that quinacainol may be exerting class 1c anti-arrhythmic actions. [source] Block of HERG-Carried K+ Currents by the New Repolarization Delaying Agent H 345/52JOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 6 2003Gregory J. Amos M.D. Ph.D. Introduction: The aim of this study was to analyze the block of HERG-carried membrane currents caused by H 345/52, a new antiarrhythmic compound with low proarrhythmic activity, in transfected mouse fibroblasts. Methods and Results: Using the whole-cell configuration of the voltage patch clamp technique, it was demonstrated that H 345/52 concentration-dependently blocked HERG-carried currents with an IC50 of 230 nM. H 345/52 preferentially bound to the open channel with unusually rapid kinetics and was trapped by channel closure. Voltage-independent behavior of H 345/52 was observed during both square-pulse and action potential clamp protocols. In contrast, the Class III agents dofetilide (10 nM) and almokalant (250 nM) demonstrated significant membrane potential-dependent effects during square-pulse clamp protocols. When using action potential clamp protocols, voltage dependence was seen with dofetilide but not with almokalant. Mathematical simulations of human ventricular action potentials predicted that the different voltage-dependent behaviors would not produce marked variations in action potential duration prolongation patterns. Conclusion: We propose that block of IKr is the principal mechanism by which H 345/52 delays repolarization in human myocardium. The voltage independence of HERG/IKr block is unlikely to underlie the low proarrhythmic potential, and ancillary effects on other membrane currents must be considered. (J Cardiovasc Electrophysiol, Vol. 14, pp. 651-658, June 2003) [source] Purinergic activation of a leak potassium current in freshly dissociated myocytes from mouse thoracic aortaACTA PHYSIOLOGICA, Issue 2 2009S. Hayoz Abstract Aim:, Exogenous ATP elicits a delayed calcium-independent K+ current on freshly isolated mouse thoracic aorta myocytes. We investigated the receptor, the intracellular pathway and the nature of this current. Methods:, The patch-clamp technique was used to record ATP-elicited delayed K+ current in freshly dissociated myocytes. Results:, ATP-elicited delayed K+ current was not inhibited by a ,cocktail' of K+ channel blockers (4-AP, TEA, apamin, charybdotoxin, glibenclamide). The amplitude of the delayed K+ current decreased after the reduction of extracellular pH from 7.4 to 6.5. These two characteristics suggest that this current could be carried by the TASK subfamily of ,twin-pore potassium channels' (K2P). Purinergic agonists including dATP, but not ADP, activated the delayed K+ current, indicating that P2Y11 is the likely receptor involved in its activation. The PKC activator phorbol ester 12,13-didecanoate stimulated this current. In addition, the PKC inhibitor Gö 6850 partially inhibited it. Real-time quantitative PCR showed that the genes encoding TASK-1 and TASK-2 are expressed. Conclusion:, Our results indicate that blocker cocktail-insensitive delayed K+ current in freshly dissociated aortic myocytes is probably carried by the TASK subfamily of twin-pore channels. [source] Protein kinase A modulates A-type potassium currents of larval zebrafish (Danio rerio) white muscle fibresACTA PHYSIOLOGICA, Issue 2 2009C. A. Coutts Abstract Aims:, Potassium (K+) channels are involved in regulating cell excitability and action potential shape. To our knowledge, very little is known about the modulation of A-type K+ currents in skeletal muscle fibres. Therefore, we sought to determine whether K+ currents of zebrafish white skeletal muscle were modulated by protein kinase A (PKA). Methods:, Pharmacology and whole-cell patch clamp were used to examine A-type K+ currents and action potentials associated with zebrafish white skeletal muscle fibres. Results:, Activation of PKA by a combination of forskolin + 3-isobutyl-1-methylxanthine (Fsk + IBMX) decreased the peak current density by ,60% and altered the inactivation kinetics of A-type K+ currents. The specific PKA inhibitor H-89 partially blocked the Fsk + IBMX-induced reduction in peak current density, but had no effect on the change in decay kinetics. Fsk + IBMX treatment did not shift the activation curve, but it significantly reduced the slope factor of activation. Activation of PKA by Fsk + IBMX resulted in a negative shift in the V50 of inactivation. H-89 prevented all Fsk + IBMX-induced changes in the steady-state properties of K+ currents. Application of Fsk + IBMX increased action potential amplitude, but had no significant effect on action potential threshold, half width or recovery rate, when fibres were depolarized with single pulses, paired pulses or with high-frequency stimuli. Conclusion:, PKA modulates the A-type K+ current in zebrafish skeletal muscle and affects action potential properties. Our results provide new insights into the role of A-type K+ channels in muscle physiology. [source] Androgen modulates the kinetics of the delayed rectifying K+ current in the electric organ of a weakly electric fishDEVELOPMENTAL NEUROBIOLOGY, Issue 12 2007M. Lynne McAnelly Abstract Weakly electric fish such as Sternopygus macrurus utilize a unique signal production system, the electric organ (EO), to navigate within their environment and to communicate with conspecifics. The electric organ discharge (EOD) generated by the Sternopygus electric organ is quasi-sinusoidal and sexually dimorphic; sexually mature males produce long duration EOD pulses at low frequencies, whereas mature females produce short duration EOD pulses at high frequencies. EOD frequency is set by a medullary pacemaker nucleus, while EOD pulse duration is determined by the kinetics of Na+ and K+ currents in the electric organ. The inactivation of the Na+ current and the activation of the delayed rectifying K+ current of the electric organ covary with EOD frequency such that the kinetics of both currents are faster in fish with high (female) EOD frequency than those with low (male) EOD frequencies. Dihydrotestosterone (DHT) implants masculinize the EOD centrally by decreasing frequency at the pacemaker nucleus (PMN). DHT also acts at the electric organ, broadening the EO pulse, which is at least partly due to a slowing of the inactivation kinetics of the Na+ current. Here, we show that chronic DHT treatment also slows the activation and deactivation kinetics of the electric organ's delayed rectifying K+ current. Thus, androgens coregulate the time-varying kinetics of two distinct ion currents in the EO to shape a sexually dimorphic communication signal. © 2007 Wiley Periodicals, Inc. Develop Neurobiol, 2007 [source] Development of ionic currents of zebrafish slow and fast skeletal muscle fibersDEVELOPMENTAL NEUROBIOLOGY, Issue 3 2006Christopher A. Coutts Abstract Voltage-gated Na+ and K+ channels play key roles in the excitability of skeletal muscle fibers. In this study we investigated the steady-state and kinetic properties of voltage-gated Na+ and K+ currents of slow and fast skeletal muscle fibers in zebrafish ranging in age from 1 day postfertilization (dpf) to 4,6 dpf. The inner white (fast) fibers possess an A-type inactivating K+ current that increases in peak current density and accelerates its rise and decay times during development. As the muscle matured, the V50s of activation and inactivation of the A-type current became more depolarized, and then hyperpolarized again in older animals. The activation kinetics of the delayed outward K+ current in red (slow) fibers accelerated within the first week of development. The tail currents of the outward K+ currents were too small to allow an accurate determination of the V50s of activation. Red fibers did not show any evidence of inward Na+ currents; however, white fibers expressed Na+ currents that increased their peak current density, accelerated their inactivation kinetics, and hyperpolarized their V50 of inactivation during development. The action potentials of white fibers exhibited significant changes in the threshold voltage and the half width. These findings indicate that there are significant differences in the ionic current profiles between the red and white fibers and that a number of changes occur in the steady-state and kinetic properties of Na+ and K+ currents of developing zebrafish skeletal muscle fibers, with the most dramatic changes occurring around the end of the first day following egg fertilization. © 2005 Wiley Periodicals, Inc. J Neurobiol, 2006 [source] Electrophysiological and morphological characterization of dentate astrocytes in the hippocampusDEVELOPMENTAL NEUROBIOLOGY, Issue 2 2005Masako Isokawa Abstract We studied electrophysiological and morphological properties of astrocytes in the dentate gyrus of the rat hippocampus in slices. Intracellular application of Lucifer yellow revealed two types of morphology: one with a long process extruding from the cell body, and the other with numerous short processes surrounding the cell body. Their electrophysiological properties were either passive, that is, no detectable voltage-dependent conductance, or complex, with Na+/K+ currents similar to those reported in the Ammon's horn astrocytes. We did not find any morphological correlate to the types of electrophysiological profile or dye coupling. Chelation of cytoplasmic calcium ([Ca2+]i) by BAPTA increased the incidence of detecting a low Na+ conductance and transient outward K+ currents. However, an inwardly rectifying K+ current (Kir), a hallmark of differentiated CA1/3 astrocytes, was not a representative K+ -current in the complex dentate astrocytes, suggesting that these astrocytes could retain an immature form of K-currents. Dentate astrocytes may possess a distinct current profile that is different from those in CA1/3 Ammon's horn. © 2005 Wiley Periodicals, Inc. J Neurobiol, 2005 [source] Epileptiform Activity Induced by Pharmacologic Reduction of M-Current in the Developing Hippocampus in VitroEPILEPSIA, Issue 1 2006Fernando Peña Summary:,Purpose: Benign familial neonatal convulsions (BFNCs), an inheritable epilepsy that occurs in neonates but not in adults, is caused by hypofunctional mutations in genes codifying for the M-type K+ current. In an attempt to develop an in vitro model of this disease, we tested whether blocking M-current with linopirdine induces epileptiform activity in brain slices from animals of different ages. Methods: Horizontal hippocampus,entorhinal cortex slices were obtained from neonatal (1,2 weeks after birth) and adult (8,9 weeks after birth) rats. Extracellular field recordings of the CA1 region were performed. After recording control conditions, linopirdine was added to the bath, and field activity was recorded continuously for 3 h. 4-Aminopyridine, a drug commonly used to induce epileptiform activity in vitro, was used as a control for our experimental conditions. Results: Bath perfusion of linopirdine induced epileptiform activity only in slices from neonatal rats. Epileptiform activity consisted of interictal-like and ictal-like activity. In slices from adult rats, linopirdine induced erratic interictal-like activity. In contrast, 4-aminopyridine was able to induce epileptiform activity in slices from both neonatal and adult rats. Conclusions: We demonstrated that blockade of M-current in vitro produces epileptiform activity with a developmental pattern similar to that observed in BNFCs. This could be an in vitro model that can be used to study the cellular mechanisms of epileptogenesis and the developmental features of BFNCs, as well as to develop some therapeutic strategies. [source] Inhibitory Effect of Lamotrigine on A-type Potassium Current in Hippocampal Neuron,Derived H19-7 CellsEPILEPSIA, Issue 7 2004Chin-Wei Huang Summary:,Purpose: We investigated the effects of lamotrigine (LTG) on the rapidly inactivating A-type K+ current (IA) in embryonal hippocampal neurons. Methods: The whole-cell configuration of the patch-clamp technique was applied to investigate the ion currents in cultured hippocampal neuron,derived H19-7 cells in the presence of LTG. Effects of various related compounds on IA in H19-7 cells were compared. Results: LTG (30 ,M,3 mM) caused a reversible reduction in the amplitude of IA. The median inhibitory concentration (IC50) value required for the inhibition of IA by LTG was 160 ,M. 4-Aminopyridine (1 mM), quinidine (30 ,M), and capsaicin (30 ,M) were effective in suppressing the amplitude of IA, whereas tetraethylammonium chloride (1 mM) and gabapentin (100 ,M) had no effect on it. The time course for the inactivation of IA was changed to the biexponential process during cell exposure to LTG (100 ,M). LTG (300 ,M) could shift the steady-state inactivation of IA to a more negative membrane potential by approximately ,10 mV, although it had no effect on the slope of the inactivation curve. Moreover, LTG (100 ,M) produced a significant prolongation in the recovery of IA inactivation. Therefore in addition to the inhibition of voltage-dependent Na+ channels, LTG could interact with the A-type K+ channels to suppress the amplitude of IA. The blockade of IA by LTG does not simply reduce current magnitude, but alters current kinetics, suggesting a state-dependent blockade. LTG might have a higher affinity to the inactivated state than to the resting state of the IA channel. Conclusions: This study suggests that in hippocampal neurons, during exposure to LTG, the LTG-mediated inhibition of these K+ channels could be one of the ionic mechanisms underlying the increased neuronal excitability. [source] PACAP inhibits delayed rectifier potassium current via a cAMP/PKA transduction pathway: evidence for the involvement of IK in the anti-apoptotic action of PACAPEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 6 2004Y. A. Mei Abstract Activation of potassium (K+) currents plays a critical role in the control of programmed cell death. Because pituitary adenylate cyclase-activating polypeptide (PACAP) has been shown to inhibit the apoptotic cascade in the cerebellar cortex during development, we have investigated the effect of PACAP on K+ currents in cultured cerebellar granule cells using the patch-clamp technique in the whole-cell configuration. Two types of outward K+ currents, a transient K+ current (IA) and a delayed rectifier K+ current (IK) were characterized using two different voltage protocols and specific inhibitors of K+ channels. Application of PACAP induced a reversible reduction of the IK amplitude, but did not affect IA, while the PACAP-related peptide vasoactive intestinal polypeptide had no effect on either types of K+ currents. Repeated applications of PACAP induced gradual attenuation of the electrophysiological response. In the presence of guanosine 5,-[,thio]triphosphate (GTP,S), PACAP provoked a marked and irreversible IK depression, whereas cell dialysis with guanosine 5,-[,thio]diphosphate GDP,S totally abolished the effect of PACAP. Pre-treatment of the cells with pertussis toxin did not modify the effect of PACAP on IK. In contrast, cholera toxin suppressed the PACAP-induced inhibition of IK. Exposure of granule cells to dibutyryl cyclic adenosine monophosphate (dbcAMP) mimicked the inhibitory effect of PACAP on IK. Addition of the specific protein kinase A inhibitor H89 in the patch pipette solution prevented the reduction of IK induced by both PACAP and dbcAMP. PACAP provoked a sustained increase of the resting membrane potential in cerebellar granule cells cultured either in high or low KCl-containing medium, and this long-term depolarizing effect of PACAP was mimicked by the IK specific blocker tetraethylammonium chloride (TEA). In addition, pre-incubation of granule cells with TEA suppressed the effect of PACAP on resting membrane potential. TEA mimicked the neuroprotective effect of PACAP against ethanol-induced apoptotic cell death, and the increase of caspase-3 activity observed after exposure of granule cells to ethanol was also significantly inhibited by TEA. Taken together, the present results demonstrate that, in rat cerebellar granule cells, PACAP reduces the delayed outward rectifier K+ current by activating a type 1 PACAP (PAC1) receptor coupled to the adenylyl cyclase/protein kinase A pathway through a cholera toxin-sensitive Gs protein. Our data also show that PACAP and TEA induce long-term depolarization of the resting membrane potential, promote cell survival and inhibit caspase-3 activity, suggesting that PACAP-evoked inhibition of IK contributes to the anti-apoptotic effect of the peptide on cerebellar granule cells. [source] SK channels and the varieties of slow after-hyperpolarizations in neuronsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2003Fivos Vogalis Abstract Action potentials and associated Ca2+ influx can be followed by slow after-hyperpolarizations (sAHPs) caused by a voltage-insensitive, Ca2+ -dependent K+ current. Slow AHPs are a widespread phenomenon in mammalian (including human) neurons and are present in both peripheral and central nervous systems. Although, the molecular identity of ion channels responsible for common membrane potential mechanisms has been largely determined, the nature of the channels that underlie the sAHPs in neurons, both in the brain and in the periphery, remains unresolved. This short review discusses why there is no clear molecular candidate for sAHPs. [source] A Novel Background Potassium Channel in Rat Atrial CellsEXPERIMENTAL PHYSIOLOGY, Issue 4 2000Z. Shui A K+ channel activated by intracellular ATP has been observed in inside-out patches from rat atrial cells. The channel has a slope conductance of 130 ± 5 pS in symmetrical 140 mM K+ solution, and is almost independent of voltage over the range from -80 to +80 mV. There is no detectable inactivation during application of ATP over a few minutes. In the presence of 3 mM intracellular ATP, channel openings occur as bursts with a mean open time of 1.7 ms, a mean closed time of 0.4 ms, a mean burst duration of 18 ms and a mean burst interval of 41 ms. Kinetic analysis suggests that ATP mainly affects the burst duration and the burst interval of the channel. Based on the properties above, the channel differs from other known K+ channels in cardiac cells and may contribute to background K+ current. [source] Enhancement of neuronal outward delayed rectifier K+ current by human monocyte-derived macrophagesGLIA, Issue 14 2009Dehui Hu Abstract Macrophages are critical cells in mediating the pathology of neurodegenerative disorders and enhancement of neuronal outward potassium (K+) current has implicated in neuronal apoptosis. To understand how activated macrophages induce neuronal dysfunction and injury, we studied the effects of lipopolysaccharide (LPS)-stimulated human monocytes-derived macrophage (MDM) on neuronal outward delayed rectifier K+ current (IK) and resultant change on neuronal viability in primary rat hippocampal neuronal culture. Bath application of LPS-stimulated MDM-conditioned media (MCM) enhanced neuronal IK in a concentration-dependentmanner, whereas non-stimulated MCM failed to alter neuronal IK. The enhancement of neuronal IK was repeated in a macrophage-neuronal co-culture system. The link of stimulated MCM (MCM(+))-associated enhancement of IK to MCM(+)-induced neuronal injury, as detected by PI/DAPI (propidium iodide/4,,6-diamidino-2-phenylindol) staining and MTT assay, was demonstrated by experimental results showing that addition of IK blocker tetraethylammonium to the culture protected hippocampal neurons from MCM(+)-associated challenge. Further investigation revealed elevated levels of Kv 1.3 and Kv 1.5 channel expression in hippocampal neurons after addition of MCM(+) to the culture. These results suggest that during brain inflammation macrophages, through their capacity of releasing bioactive molecules, induce neuronal injury by enhancing neuronal IK and that modulation of Kv channels is a new approach to neuroprotection. © 2009 Wiley-Liss, Inc. [source] Molecular cloning, genomic organization and functional characterization of a new short-chain potassium channel toxin-like peptide BmTxKS4 from Buthus martensii Karsch(BmK)JOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 4 2004Sheng Jiqun Abstract Scorpion venom contains many small polypeptide toxins, which can modulate Na+, K+, Cl,, and Ca2+ ion,channel conductance in the cell membrane. A full-length cDNA sequence encoding a novel type of K+ -channel toxin (named BmTxKS4) was first isolated and identified from a venom gland cDNA library of Buthus martensii Karsch (BmK). The encoded precursor contains 78 amino acid residues including a putative signal peptide of 21 residues, propeptide of 11 residues, and a mature peptide of 43 residues with three disulfide bridges. BmTxKS4 shares the identical organization of disulfide bridges with all the other short-chain K+ -channel scorpion toxins. By PCR amplification of the genomic region encoding BmTxKS4, it was shown that BmTxKS4 composed of two exons is disrupted by an intron of 87 bp inserted between the first and the second codes of Phe (F) in the encoding signal peptide region, which is completely identical with that of the characterized scorpion K+ -channel ligands in the size, position, consensus junctions, putative branch point, and A+T content. The GST-BmTxKS4 fusion protein was successfully expressed in BL21 (DE3) and purified with affinity chromatography. About 2.5 mg purified recombinant BmTxKS4 (rBmTxKS4) protein was obtained by treating GST-BmTxKS4 with enterokinase and sephadex chromatography from 1 L bacterial culture. The electrophysiological activity of 1.0,M rBmTxKS4 was measured and compared by whole cell patch-clamp technique. The results indicated that rBmTxKS4 reversibly inhibited the transient outward K+ current (Ito), delayed inward rectifier K+ current (Ik1), and prolonged the action potential duration of ventricular myocyte, but it has no effect on the action potential amplitude. Taken together, BmTxKS4 is a novel subfamily member of short-strain K+ -channel scorpion toxin. © 2004 Wiley Periodicals, Inc. J Biochem Mol Toxicol 18:187,195, 2004; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20026 [source] Changes in Left Ventricular Repolarization and Ion Channel Currents Following a Transient Rate Increase Superimposed on Bradycardia in Anesthetized DogsJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 6 2000MICHAEL RUBART M.D. Electrical Remodeling of the Heart due to Rate. Introduction: We previously demonstrated in dogs that a transient rate increase superimposed on bradycardia causes prolongation of ventricular refractoriness that persists for hours after resumption of bradycardia. In this study, we examined changes in membrane currents that are associated with this phenomenon. Methods and Results: The whole cell, patch clamp technique was used to record transmembrane voltages and currents, respectively, in single mid-myocardial left ventricular myocytes from dogs with 1 week of complete AV block; dogs either underwent 1 hour of left ventricular pacing at 120 beats/min or did not undergo pacing. Pacing significantly heightened mean phase 1 and peak plateau amplitudes by ,6 and ,3 mV, respectively (P < 0.02). and prolonged action potential duration at 90% repolarization from 235 ± 8 msec to 278 ± 8 msec (1 Hz; P = 0.02). Rapid pacing-induced changes in transmembrane ionic currents included (1) a more pronounced cumulative inactivation of the 4-aminopyridine-sensitive transient outward K+ current, I to over the range of physiologic frequencies, resulting from a ,30% decrease in the population of quickly reactivating channels; (2) increases in peak density of L-type Ca2+ currents, Ica.I.' by 15% to 35% between +10 and +60 mV; and (3) increases in peak density of the Ca2+ -activated chloride current, ICl.Ca' by 30% to 120% between +30 and +50 mV. Conclusion: Frequency-dependent reduction in Ito combined with enhanced ICa.I. causes an increase in net inward current that may he responsible for the observed changes in ventricular repolarization. This augmentation of net cation influx is partially antagonized by an increase in outward ICa.Cl. [source] Few cultured rat primary sensory neurons express a tolbutamide-sensitive K+ currentJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 2 2002Violeta Ristoiu Abstract The response of dorsal root ganglion (DRG) neurons to metabolic inhibition is known to involve calcium-activated K+ channels; in most neuronal types ATP-sensitive K+ channels (KATP) also contribute, but this is not yet established in the DRG. We have investigated the presence of a KATP current using whole-cell recordings from rat DRG neurons, classifying the neurons functionally by their "current signature" (Petruska et al, J Neurophysiol 84: 2365,2379, 2000). We clearly identified a KATP current in only 1 out of 62 neurons, probably a nociceptor. The current was activated by cyanide (2 mM NaCN) and was sensitive to 100 ,M tolbutamide; the relation between reversal potential and external K+ concentration indicated it was a K+ current. In a further two neurons, cyanide activated a K+ current that was only partially blocked by tolbutamide, which may also be an atypical KATP current. We conclude that KATP channels are expressed in normal DRG, but in very few neurons and only in nociceptors. [source] The Kv4.2 mediates excitatory activity-dependent regulation of neuronal excitability in rat cortical neuronsJOURNAL OF NEUROCHEMISTRY, Issue 3 2008Bin Shen Abstract Neuronal excitability can cooperate with synaptic transmission to control the information storage. This regulation of neuronal plasticity can be affected by alterations in neuronal inputs and accomplished by modulation of voltage-dependent ion channels. In this study, we report that enhanced excitatory input negatively regulated neuronal excitability. Enhanced excitatory input by glutamate, electric field stimulation or high K+ increased transient outward K+ current, whereas did not affect the delayed rectifier K+ current in rat cultured cortical neurons. Both the voltage-dependent K+ channel 4.2 and 4.3 subunits contributed to the increase. The increase in the K+ current density by Kv4.2 was ascribed to its cytoplasmic membrane translocation, which was mediated by NMDA type of glutamate receptor. Furthermore, enhanced excitatory input inhibited neuronal excitability. Taken together, our results suggest that excitatory neurotransmission affects neuronal excitability via the regulation of the K+ channel membrane translocation. [source] Electrophysiological Remodeling in Human Atrial FibrillationPACING AND CLINICAL ELECTROPHYSIOLOGY, Issue 7p2 2003DAVID R. VAN WAGONER Atrial fibrillation (AF) is a progressive disease characterized by cumulative electrophysiological and structural remodeling of the atria. Cellular electrophysiological studies have revealed marked reductions in the densities of the L-type voltage-gated Ca2+ current, ICa,L, the transient outward K+ current, ITO, and the ultra-rapid delayed rectifier K+ current, IKur, in atrial myocytes from patients in persistent or permanent AF. The density of the muscarinic K+ current (IKACh) is also reduced, however the inward rectifier K+ current (IK1) density is increased. The net shortening or lengthening of the action potential is dependent on the balance between changes in inward and outward currents. The prominent reduction in ICa,L appears to be sufficient to explain the observed decreases in action potential duration and effective refractory period that are characteristic of the fibrillating atria. Earlier studies have shown that calcium overload and perturbations in calcium handling play prominent roles in AF induced atrial remodeling. More recently, we have shown that AF is associated with evidence of oxidative injury to atrial tissue, and suggested that oxidative stress may directly contribute to the pathophysiology of AF. It is anticipated that insights gleaned from mechanistic studies will facilitate the development of improved pharmacological approaches to treat AF and to prevent the progression of arrhythmia. (PACE 2003; 26[Pt. II]:1572,1575) [source] Calcium-dependent K current in plasma membranes of dermal cells of developing bean cotyledonsPLANT CELL & ENVIRONMENT, Issue 2 2004W.-H. ZHANG ABSTRACT In developing seeds of bean (Phaseolus vulgaris L.), phloem-imported assimilates (largely sucrose and potassium) are released from coats to seed apoplasm and subsequently retrieved by the dermal cell complexes of cotyledons. To investigate the mechanisms of K+ uptake by the cotyledons, protoplasts of dermal cell complexes were isolated and whole-cell currents across their plasma membranes were measured with the patch-clamp technique. A weakly rectified cation current displaying a voltage-dependent blockade by external Ca2+ and acidic pH, dominated the conductance of the protoplasts. The P haseolus v ulgaris Cotyledon Dermal-cell pH and Calcium-dependent Cation Conductance (Pv-CD-pHCaCC) was highly selective for K+ over Ca2+ and Cl,. For K+ current through Pv-CD-pHCaCC a sigmoid shaped current,voltage (I,V) curve was observed with negative conductance at voltages between ,200 and ,140 mV. This negative K+ conductance was Ca2+ dependent. With other univalent cations (Na+, Rb+, NH4+) the currents were smaller and were not Ca2+ dependent. Reversal potentials remained constant when external K+ was substituted with these cations, suggesting that Pv-CD-pHCaCC channels were non-selective. The Pv-CD-pHCaCC would provide a pathway for K+ and other univalent cation influx into developing cotyledons. These cation influxes could be co-ordinated with sucrose influx via pH and Ca2+dependence. [source] Mechanisms by which atrial fibrillation-associated mutations in the S1 domain of KCNQ1 slow deactivation of IKs channelsTHE JOURNAL OF PHYSIOLOGY, Issue 17 2008Lioara Restier The slow delayed rectifier K+ current (IKs) is a major determinant of action potential repolarization in the heart. IKs channels are formed by coassembly of pore-forming KCNQ1 ,-subunits and ancillary KCNE1 ,-subunits. Two gain of function mutations in KCNQ1 subunits (S140G and V141M) have been associated with atrial fibrillation (AF). Previous heterologous expression studies found that both mutations caused IKs to be instantaneously activated, presumably by preventing channel closure. The purpose of this study was to refine our understanding of the channel gating defects caused by these two mutations located in the S1 domain of KCNQ1. Site-directed mutagenesis was used to replace S140 or V141 with several other natural amino acids. Wild-type and mutant channels were heterologously expressed in Xenopus oocytes and channel function was assessed with the two-microelectrode voltage clamp technique. Long intervals between voltage clamp pulses revealed that S140G and V141M KCNQ1-KCNE1 channels are not constitutively active as previously reported, but instead exhibit extremely slow deactivation. The slow component of IKs deactivation was decreased 62-fold by S140G and 140-fold by the V141M mutation. In addition, the half-point for activation of these mutant IKs channels was ,50 mV more negative than wild-type channels. Other substitutions of S140 or V141 in KCNQ1 caused variable shifts in the voltage dependence of activation, but slowed IKs deactivation to a much lesser extent than the AF-associated mutations. Based on a published structural model of KCNQ1, S140 and V141 are located near E160 in S2 and R237 in S4, two charged residues that could form a salt bridge when the channel is in the open state. In support of this model, mutational exchange of E160 and R237 residues produced a constitutively open channel. Together our findings suggest that altered charge-pair interactions within the voltage sensor module of KCNQ1 subunits may account for slowed IKs deactivation induced by S140 or V141. [source] Temperature-sensitive TREK currents contribute to setting the resting membrane potential in embryonic atrial myocytesTHE JOURNAL OF PHYSIOLOGY, Issue 15 2008Hengtao Zhang TREK channels belong to the superfamily of two-pore-domain K+ channels and are activated by membrane stretch, arachidonic acid, volatile anaesthetics and heat. TREK-1 is highly expressed in the atrium of the adult heart. In this study, we investigated the role of TREK-1 and TREK-2 channels in regulating the resting membrane potential (RMP) of isolated chicken embryonic cardiac myocytes. At room temperature, the average RMP of embryonic day (ED) 11 atrial myocytes was ,22 ± 2 mV. Raising the temperature to 35°C hyperpolarized the membrane to ,69 ± 2 mV and activated a large outwardly rectifying K+ current that was relatively insensitive to conventional K+ channel inhibitors (TEA, 4-AP and Ba2+) but completely inhibited by tetracaine (200 ,m), an inhibitor of TREK channels. The heat-induced hyperpolarization was mimicked by 10 ,m arachidonic acid, an agonist of TREK channels. There was little or no inwardly rectifying K+ current (IK1) in the ED11 atrial cells. In marked contrast, ED11 ventricular myocytes exhibited a normal RMP (,86.1 ± 3.4 mV) and substantial IK1, but no temperature- or tetracaine-sensitive K+ currents. Both RT-PCR and real-time PCR further demonstrated that TREK-1 and TREK-2 are highly and almost equally expressed in ED11 atrium but much less expressed in ED11 ventricle. In addition, immunofluorescence demonstrated TREK-1 protein in the membrane of atrial myocytes. These data indicate the presence and function of TREK-1 and TREK-2 in the embryonic atrium. Moreover, we demonstrate that TREK-like currents have an essential role in determining membrane potential in embryonic atrial myocytes, where IK1 is absent. [source] Depletion of membrane cholesterol eliminates the Ca2+ -activated component of outward potassium current and decreases membrane capacitance in rat uterine myocytesTHE JOURNAL OF PHYSIOLOGY, Issue 2 2007A. Shmygol Changes in membrane cholesterol content have potent effects on cell signalling and contractility in rat myometrium and other smooth muscles. We have previously shown that depletion of cholesterol with methyl-,-cyclodextrin (MCD) disrupts caveolar microdomains. The aim of this work was to determine the mechanism underlying the increase in Ca2+ signalling and contractility occurring in the myometrium with MCD. Patch clamp data obtained on freshly isolated myocytes from the uterus of day 19,21 rats showed that outward K+ current was significantly reduced by MCD. Membrane capacitance was also reduced. Cholesterol-saturated MCD had no effect on the amplitude of outward current suggesting that the reduction in the outward current was due to cholesterol depletion induced by MCD rather than a direct inhibitory action of MCD on the K+ channels. Confocal visualization of the membrane bound indicator Calcium Green C18, revealed internalization of the surface membrane with MCD treatment. Large conductance, Ca2+ -sensitive K+ channel proteins have been shown to localize to caveolae. When these channels were blocked by iberiotoxin outward current was significantly reduced in the uterine myocytes; MCD treatment reduced the density of outward current. Following reduction of outward current by MCD pretreatment, iberiotoxin was unable to produce any additional decrease in the current, suggesting a common target. MCD treatment also increased the amplitude and frequency of spontaneous rises in cytosolic Ca2+ level ([Ca2+]i transients) in isolated myocytes. In intact rat myometrium, MCD treatment increased Ca2+ signalling and contractility, consistent with previous findings, and this effect was also found to be reduced by BK channel inhibition. These data suggest that (1) disruption of cholesterol-rich microdomains and caveolae by MCD leads to a decrease in the BK channel current thus increasing cell excitability, and (2) the changes in membrane excitability produced by MCD underlie the changes found in Ca2+ signalling and uterine contractility. [source] Tyrosine kinase and phosphatase regulation of slow delayed-rectifier K+ current in guinea-pig ventricular myocytesTHE JOURNAL OF PHYSIOLOGY, Issue 2 2006Sergey Missan The objective of this study was to investigate the involvement of tyrosine phosphorylation in the regulation of the cardiac slowly activating delayed-rectifier K+ current (IKs) that is important for action potential repolarization. Constitutive IKs recorded from guinea-pig ventricular myocytes was suppressed by broad-spectrum tyrosine kinase (TK) inhibitors tyrphostin A23 (IC50, 4.1 ± 0.6 ,m), tyrphostin A25 (IC50, 12.1 ± 2.1 ,m) and genistein (IC50, 64 ± 4 ,m), but was relatively insensitive to the inactive analogues tyrphostin A1, tyrphostin A63, daidzein and genistin. IKs was unaffected by AG1478 (10 ,m), an inhibitor of epidermal growth factor receptor TK, and was strongly suppressed by the Src TK inhibitor PP2 (10 ,m) but not by the inactive analogue PP3 (10 ,m). The results of experiments with forskolin, H89 and bisindolylmaleimide I indicate that the suppression of IKs by TK inhibitors was not mediated via inhibition of (IKs -stimulatory) protein kinases A and C. To evaluate whether the suppression was related to lowered tyrosine phosphorylation, myocytes were pretreated with TK inhibitors and then exposed to the phosphotyrosyl phosphatase inhibitor orthovanadate (1 mm). Orthovanadate almost completely reversed the suppression of IKs induced by broad-spectrum TK inhibitors at concentrations around their IC50 values. We conclude that basal IKs is strongly dependent on tyrosine phosphorylation of Ks channel (or channel-regulatory) protein. [source] A voltage-dependent K+ current contributes to membrane potential of acutely isolated canine articular chondrocytesTHE JOURNAL OF PHYSIOLOGY, Issue 1 2004Jim R. Wilson The electrophysiological properties of acutely isolated canine articular chondrocytes have been characterized using patch-clamp methods. The ,steady-state' current,voltage relationship (I,V) of single chondrocytes over the range of potentials from ,100 to +40 mV was highly non-linear, showing strong outward rectification positive to the zero-current potential. Currents activated at membrane potentials negative to ,50 mV were time independent, and the I,V from ,100 to ,60 mV was linear, corresponding to an apparent input resistance of 9.3 ± 1.4 G, (n= 23). The outwardly rectifying current was sensitive to the K+ channel blocking ion tetraethylammonium (TEA), which had a 50% blocking concentration of 0.66 mm (at +50 mV). The ,TEA-sensitive' component of the outwardly rectifying current had time- and membrane potential-dependent properties, activated near ,45 mV and was half-activated at ,25 mV. The reversal potential of the ,TEA-sensitive' current with external K+ concentration of 5 mm and internal concentration of 145 mm, was ,84 mV, indicating that the current was primarily carried by K+ ions. The resting membrane potential of isolated chondrocytes (,38.1 ± 1.4 mV; n= 19) was depolarized by 14.8 ± 0.9 mV by 25 mm TEA, which completely blocked the K+ current of these cells. These data suggest that this voltage-sensitive K+ channel has an important role in regulating the membrane potential of canine articular chondrocytes. [source] Flufenamic acid blocks depolarizing afterpotentials and phasic firing in rat supraoptic neuronesTHE JOURNAL OF PHYSIOLOGY, Issue 2 2002Masoud Ghamari-Langroudi Depolarizing afterpotentials (DAPs) that follow action potentials in magnocellular neurosecretory cells (MNCs) are thought to underlie the generation of phasic firing, a pattern that optimizes vasopressin release from the neurohypophysis. Previous work has suggested that the DAP may result from the Ca2+ -dependent reduction of a resting K+ conductance. Here we examined the effects of flufenamic acid (FFA), a blocker of Ca2+ -dependent non-selective cation (CAN) channels, on DAPs and phasic firing using intracellular recordings from supraoptic MNCs in superfused explants of rat hypothalamus. Application of FFA, but not solvent (0.1 % DMSO), reversibly inhibited (IC50+ 13.8 ,m; R+ 0.97) DAPs and phasic firing with a similar time course, but had no significant effects (P > 0.05) on membrane potential, spike threshold and input resistance, nor on the frequency and amplitude of spontaneous synaptic potentials. Moreover, FFA did not affect (P > 0.05) the amplitude, duration, undershoot, or frequency-dependent broadening of action potentials elicited during the spike trains used to evoke DAPs. These findings suggest that FFA inhibits the DAP by directly blocking the channels responsible for its production, rather than by interfering with Ca2+ influx. They also support a role for DAPs in the generation of phasic firing in MNCs. Finally, the absence of a depolarization and increased membrane resistance upon application of FFA suggests that the DAP in MNCs may not be due to the inhibition of resting K+ current, but to the activation of CAN channels. [source] Contribution of Kv4 channels toward the A-type potassium current in murine colonic myocytesTHE JOURNAL OF PHYSIOLOGY, Issue 2 2002Gregory C. Amberg A rapidly inactivating K+ current (A-type current; IA) present in murine colonic myocytes is important in maintaining physiological patterns of slow wave electrical activity. The kinetic profile of colonic IA resembles that of Kv4-derived currents. We examined the contribution of Kv4 ,-subunits to IA in the murine colon using pharmacological, molecular and immunohistochemical approaches. The divalent cation Cd2+ decreased peak IA and shifted the voltage dependence of activation and inactivation to more depolarized potentials. Similar results were observed with La3+. Colonic IA was sensitive to low micromolar concentrations of flecainide (IC50= 11 ,M). Quantitative PCR indicated that in colonic and jejunal tissue, Kv4.3 transcripts demonstrate greater relative abundance than transcripts encoding Kv4.1 or Kv4.2. Antibodies revealed greater Kv4.3-like immunoreactivity than Kv4.2-like immunoreactivity in colonic myocytes. Kv4-like immunoreactivity was less evident in jejunal myocytes. To address this finding, we examined the expression of K+ channel-interacting proteins (KChIPs), which act as positive modulators of Kv4-mediated currents. Qualitative PCR identified transcripts encoding the four known members of the KChIP family in isolated colonic and jejunal myocytes. However, the relative abundance of KChIP transcript was 2.6-fold greater in colon tissue than in jejunum, as assessed by quantitative PCR, with KChIP1 showing predominance. This observation is in accordance with the amplitude of the A-type current present in these two tissues, where colonic myocytes possess densities twice that of jejunal myocytes. From this we conclude that Kv4.3, in association with KChIP1, is the major molecular determinant of IA in murine colonic myocytes. [source] The Action of a Novel Fluoroquinolone Antibiotic Agent Antofloxacin Hydrochloride on Human-Ether- à-go-go- Related Gene Potassium ChannelBASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 2 2010Jia Guo In this study, the effects of a novel fluoroquinolone, antofloxacin hydrochloride (AX) on human- ether-à-go-go -related gene (HERG) encoding potassium channels and the biophysical mechanisms of drug action were performed with whole-cell patch-clamp technique in transiently transfected HEK293 cells. The administration of AX caused voltage- and time-dependent inhibition of HERG K+ current (IHERG/MiRP1) in a concentration-dependent manner but did not markedly modify the properties of channel kinetics, including activation, inactivation, deactivation and recovery from inactivation as well. In comparison with sparfloxacin (SPX), levofloxacin lactate (LVFX), the potency of AX to inhibit HERG tail currents was the least one, with an IC50 value of 460.37 ,M. By contrast, SPX was the most potent compound, displaying an IC50 value of 2.69 ,M whereas LVFX showed modest potency, with an IC50 value of 43.86 ,M, respectively. Taken together, our data suggest that AX only causes a minor reduction of IHERG/MiRP1 at the estimated free plasma level. [source] Electrophysiological Effects of the Anti-Cancer Drug Lapatinib on Cardiac RepolarizationBASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 1 2010Hyang-Ae Lee Although lapatinib is associated with a risk of QT prolongation, the effects of the drug on cellular cardiac electrical properties and on action potential duration (APD) have not been studied. To evaluate the potential effects of lapatinib on cardiac repolarization, we investigated its electrophysiological effects using a whole-cell patch,clamp technique in transiently transfected HEK293 cells expressing human ether-à-go-go (hERG; to examine the rapidly activating delayed rectifier K+ current, IKr), KCNQ1/KCNE1 (to examine the slowly activating delayed rectifier K+ current, IKs), KCNJ2 (to examine the inwardly rectifying K+ current, IK1), or SCN5A (to examine the inward Na+ current, INa) and in rat cardiac myocytes (to examine the inward Ca2+ current, ICa). We also examined its effects on the APD at 90% (APD90) in isolated rabbit Purkinje fibres. In ion channel studies, lapatinib inhibited the hERG current in a concentration-dependent manner, with a half-maximum inhibition concentration (IC50) of 0.8 ± 0.09 ,m. In contrast, at concentrations up to 3 ,m, lapatinib did not significantly reduce the INa, IK1 or ICa amplitudes; at 3 ,m, it did slightly inhibit the IKs amplitude (by 19.4 ± 4.7%; p < 0.05). At 5 ,m, lapatinib induced prolongation of APD90 by 16.1% (p < 0.05). These results suggest that the APD90 -prolonging effect of lapatinib on rabbit Purkinje fibres is primarily a result of inhibition of the hERG current and IKs, but not INa, IK1 or ICa. [source] Emodin Inhibits Voltage-Dependent Potassium Current in Guinea Pig Gallbladder Smooth MuscleBASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 3 2009Zhi-Xuan Wu We studied the effects of emodin on the contraction of gallbladder smooth muscle and voltage-dependent K+ current in gallbladder smooth muscle cells. Gallbladder muscle strips were obtained from adult guinea pigs and the resting tension was recorded. Gallbladder smooth muscle cells were isolated by enzymatic digestion, and K+ current was recorded by the whole-cell patch clamp method. Emodin increased the resting tension of gallbladder smooth muscle strips and inhibited voltage-dependent K+ current in a dose-dependent manner. When 10 µM emodin was applied to gallbladder smooth muscle cells for 3,6 min., the amplitude of voltage-dependent K+ current was decreased by 31.5 ± 0.5% at +40 mV, and this inhibitory effect mostly recovered after washout. The steady-state inactivation curves were shifted in a hyperpolarizing direction by emodin. In the presence of the protein kinase C inhibitors staurosporine and chelerythrine, the effect of emodin on voltage-dependent K+ current was significantly attenuated. In conclusion, emodin promotes gallbladder contraction, mainly by inhibiting voltage-dependent K+ current via the protein kinase C pathway. These findings provide theoretical foundation for the application of emodin in gallbladder motility disorders. [source] Glutamate-induced post-activation inhibition of locus coeruleus neurons is mediated by AMPA/kainate receptors and sodium-dependent potassium currentsBRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2009Teresa Zamalloa Background and purpose:, Locus coeruleus (LC) neurons respond to sensory stimuli with a glutamate-triggered burst of spikes followed by an inhibition. The aim of our work was to characterize the inhibitory effect of glutamate in the LC. Experimental approach:, Single-unit extracellular and patch-clamp recordings were performed to examine glutamate responses in rat brain slices containing the LC. Key results:, Glutamate caused an initial activation followed by a late post-activation inhibition (PAI). Both effects were blocked by an AMPA/kainate receptor antagonist but not by NMDA or metabotropic glutamate receptor antagonists. All glutamate receptor agonists were able to activate neurons, but only AMPA and quisqualate caused inhibition. In neurons clamped at ,60 mV, glutamate and AMPA induced inward, followed by outward, currents, with the latter reversing polarity at ,110 mV. Glutamate-induced PAI was not modified by ,2 -adrenoceptor, µ opioid, A1 adenosine and GABAA/B receptor antagonists or Ca2+ -dependent release blockade, but it was reduced by raising the extracellular K+ concentration. Glutamate-induced PAI was not affected by several potassium channel, Na+/K+ pump, PKC and neuronal NO synthase inhibitors or lowering the extracellular Ca2+ concentration. The Na+ -activated K channel opener bithionol concentration-dependently potentiated glutamate-induced PAI, whereas partial (80%) Na+ replacement reduced glutamate- and AMPA-induced PAI. Finally, reverse transcription polymerase chain reaction assays showed the presence of mRNA for the Ca2+ -impermeable GluR2 subunit in the LC. Conclusions and implications:, Glutamate induces a late PAI in the LC, which may be mediated by a novel postsynaptic Na+ -dependent K+ current triggered by AMPA/kainate receptors. Mandarin translation of abstract [source] | ||||||||||||||||||||||||||||