scholarly journals Permissive Modulation of Sphingosine-1-Phosphate-Enhanced Intracellular Calcium on BKCa Channel of Chromaffin Cells

2021 ◽  
Vol 22 (4) ◽  
pp. 2175
Author(s):  
Adonis Z. Wu ◽  
Tzu-Lun Ohn ◽  
Ren-Jay Shei ◽  
Huei-Fang Wu ◽  
Yong-Cyuan Chen ◽  
...  
Keyword(s):  
Chromaffin Cells ◽  
Low Dose ◽  
Single Channel ◽  
Cell System ◽  
K Channel ◽  
Lipid Mediator ◽  
Bkca Channel ◽  
High Dose ◽  
Open Probability ◽  
Sphingosine 1 Phosphate

Sphingosine-1-phosphate (S1P), is a signaling sphingolipid which acts as a bioactive lipid mediator. We assessed whether S1P had multiplex effects in regulating the large-conductance Ca2+-activated K+ channel (BKCa) in catecholamine-secreting chromaffin cells. Using multiple patch-clamp modes, Ca2+ imaging, and computational modeling, we evaluated the effects of S1P on the Ca2+-activated K+ currents (IK(Ca)) in bovine adrenal chromaffin cells and in a pheochromocytoma cell line (PC12). In outside-out patches, the open probability of BKCa channel was reduced with a mean-closed time increment, but without a conductance change in response to a low-concentration S1P (1 µM). The intracellular Ca2+ concentration (Cai) was elevated in response to a high-dose (10 µM) but not low-dose of S1P. The single-channel activity of BKCa was also enhanced by S1P (10 µM) in the cell-attached recording of chromaffin cells. In the whole-cell voltage-clamp, a low-dose S1P (1 µM) suppressed IK(Ca), whereas a high-dose S1P (10 µM) produced a biphasic response in the amplitude of IK(Ca), i.e., an initial decrease followed by a sustained increase. The S1P-induced IK(Ca) enhancement was abolished by BAPTA. Current-clamp studies showed that S1P (1 µM) increased the action potential (AP) firing. Simulation data revealed that the decreased BKCa conductance leads to increased AP firings in a modeling chromaffin cell. Over a similar dosage range, S1P (1 µM) inhibited IK(Ca) and the permissive role of S1P on the BKCa activity was also effectively observed in the PC12 cell system. The S1P-mediated IK(Ca) stimulation may result from the elevated Cai, whereas the inhibition of BKCa activity by S1P appears to be direct. By the differentiated tailoring BKCa channel function, S1P can modulate stimulus-secretion coupling in chromaffin cells.

scholarly journals Membrane-delimited Inhibition of Maxi-K Channel Activity by the Intermediate Conductance Ca2+-activated K Channel

2006 ◽  
Vol 127 (2) ◽  
pp. 159-169 ◽  
Author(s):  
Jill Thompson ◽  
Ted Begenisich
Keyword(s):  
Single Channel ◽  
Expression System ◽  
Chemical Activation ◽  
K Channel ◽  
Channel Activity ◽  
Single Family ◽  
Open Probability ◽  
K Channels ◽  
Channel Proteins ◽  
K Activity

The complexity of mammalian physiology requires a diverse array of ion channel proteins. This diversity extends even to a single family of channels. For example, the family of Ca2+-activated K channels contains three structural subfamilies characterized by small, intermediate, and large single channel conductances. Many cells and tissues, including neurons, vascular smooth muscle, endothelial cells, macrophages, and salivary glands express more than a single class of these channels, raising questions about their specific physiological roles. We demonstrate here a novel interaction between two types of Ca2+-activated K channels: maxi-K channels, encoded by the KCa1.1 gene, and IK1 channels (KCa3.1). In both native parotid acinar cells and in a heterologous expression system, activation of IK1 channels inhibits maxi-K activity. This interaction was independent of the mode of activation of the IK1 channels: direct application of Ca2+, muscarinic receptor stimulation, or by direct chemical activation of the IK1 channels. The IK1-induced inhibition of maxi-K activity occurred in small, cell-free membrane patches and was due to a reduction in the maxi-K channel open probability and not to a change in the single channel current level. These data suggest that IK1 channels inhibit maxi-K channel activity via a direct, membrane-delimited interaction between the channel proteins. A quantitative analysis indicates that each maxi-K channel may be surrounded by four IK1 channels and will be inhibited if any one of these IK1 channels opens. This novel, regulated inhibition of maxi-K channels by activation of IK1 adds to the complexity of the properties of these Ca2+-activated K channels and likely contributes to the diversity of their functional roles.

Permeation and gating properties of a cloned renal K+ channel

1995 ◽  
Vol 268 (2) ◽  
pp. C389-C401 ◽  
Author(s):  
S. Chepilko ◽  
H. Zhou ◽  
H. Sackin ◽  
L. G. Palmer
Keyword(s):  
Single Channel ◽  
Reversal Potential ◽  
Cation Binding ◽  
K Channel ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Membrane Hyperpolarization ◽  
Channel Current ◽  
Inward Currents ◽  
Kinetics Of

The renal K+ channel (ROMK2) was expressed in Xenopus oocytes, and the patch-clamp technique was used to assess its conducting and gating properties. In cell-attached patches with 110 mM K+ in the bath and pipette, the reversal potential was near zero and the inward conductance (36 pS) was larger than the outward conductance (17 pS). In excised inside-out patches the channels showed rectification in the presence of 5 mM Mg2+ on the cytoplasmic side but not in Mg(2+)-free solution. Inward currents were also observed when K+ was replaced in the pipette by Rb+, NH4+, or thallium (Tl+). The reversal potentials under these conditions yielded a selectivity sequence of Tl+ > K+ > Rb+ > NH4+. On the other hand, the slope conductances for inward current gave a selectivity sequence of K+ = NH4+ > Tl+ > Rb+. The differences in the two sequences can be explained by the presence of cation binding sites within the channel, which interact with Rb+ and Tl+ more strongly and with NH4+ less strongly than with K+. Two other ions, Ba2+ and Cs+, blocked the channel from the outside. The effect of Ba2+ (1 mM) was to reduce the open probability of the channels, whereas Cs+ (10 mM) reduced the apparent single-channel current. The effects of both blockers are enhanced by membrane hyperpolarization. The kinetics of the channel were also studied in cell-attached patches. With K+ in the pipette the distribution of open times could be described by a single exponential (tau 0 = 25 ms), whereas two exponentials (tau 1 = 1 ms, tau 2 = 30 ms) were required to describe the closed-time distribution. Hyperpolarization of the oocyte membrane decreased the open probability and tau 0, and increased tau 1, tau 2, and the number of long closures. The presence of Tl+ in the pipette significantly altered the kinetics, reducing tau 0 and eliminating the long-lived closures. These results suggest that the gating of the channel may depend on the nature of the ion in the pore.

scholarly journals Effect of Auxin (IAA) on the Fast Vacuolar (FV) Channels in Red Beet (Beta vulgaris L.) Taproot Vacuoles

2020 ◽  
Vol 21 (14) ◽  
pp. 4876
Author(s):  
Zbigniew Burdach ◽  
Agnieszka Siemieniuk ◽  
Waldemar Karcz
Keyword(s):  
Single Channel ◽  
Outward Current ◽  
K Channel ◽  
Single Channels ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Inward Current ◽  
Bath Solution ◽  
Outward Currents ◽  
Inward Currents

In contrast to the well-studied effect of auxin on the plasma membrane K+ channel activity, little is known about the role of this hormone in regulating the vacuolar K+ channels. Here, the patch-clamp technique was used to investigate the effect of auxin (IAA) on the fast-activating vacuolar (FV) channels. It was found that the macroscopic currents displayed instantaneous currents, which at the positive potentials were about three-fold greater compared to the one at the negative potentials. When auxin was added to the bath solution at a final concentration of 1 µM, it increased the outward currents by about 60%, but did not change the inward currents. The imposition of a ten-fold vacuole-to-cytosol KCl gradient stimulated the efflux of K+ from the vacuole into the cytosol and reduced the K+ current in the opposite direction. The addition of IAA to the bath solution with the 10/100 KCl gradient decreased the outward current and increased the inward current. Luminal auxin reduced both the outward and inward current by approximately 25% compared to the control. The single channel recordings demonstrated that cytosolic auxin changed the open probability of the FV channels at the positive voltages to a moderate extent, while it significantly increased the amplitudes of the single channel outward currents and the number of open channels. At the positive voltages, auxin did not change the unitary conductance of the single channels. We suggest that auxin regulates the activity of the fast-activating vacuolar (FV) channels, thereby causing changes of the K+ fluxes across the vacuolar membrane. This mechanism might serve to tightly adjust the volume of the vacuole during plant cell expansion.

scholarly journals Potassium channel types in arterial chemoreceptor cells and their selective modulation by oxygen.

1992 ◽  
Vol 100 (3) ◽  
pp. 401-426 ◽  
Author(s):  
M D Ganfornina ◽  
J López-Barneo
Keyword(s):  
Time Course ◽  
Single Channel ◽  
K Channel ◽  
Open Probability ◽  
K Channels ◽  
Glomus Cells ◽  
Control Value ◽  
Voltage Dependent ◽  
K Current ◽  
Chemoreceptor Cells

Single K+ channel currents were recorded in excised membrane patches from dispersed chemoreceptor cells of the rabbit carotid body under conditions that abolish current flow through Na+ and Ca2+ channels. We have found three classes of voltage-gated K+ channels that differ in their single-channel conductance (gamma), dependence on internal Ca2+ (Ca2+i), and sensitivity to changes in O2 tension (PO2). Ca(2+)-activated K+ channels (KCa channels) with gamma approximately 210 pS in symmetrical K+ solutions were observed when [Ca2+]i was greater than 0.1 microM. Small conductance channels with gamma = 16 pS were not affected by [Ca2+]i and they exhibited slow activation and inactivation time courses. In these two channel types open probability (P(open)) was unaffected when exposed to normoxic (PO2 = 140 mmHg) or hypoxic (PO2 approximately 5-10 mmHg) external solutions. A third channel type (referred to as KO2 channel), having an intermediate gamma(approximately 40 pS), was the most frequently recorded. KO2 channels are steeply voltage dependent and not affected by [Ca2+]i, they inactivate almost completely in less than 500 ms, and their P(open) reversibly decreases upon exposure to low PO2. The effect of low PO2 is voltage dependent, being more pronounced at moderately depolarized voltages. At 0 mV, for example, P(open) diminishes to approximately 40% of the control value. The time course of ensemble current averages of KO2 channels is remarkably similar to that of the O2-sensitive K+ current. In addition, ensemble average and macroscopic K+ currents are affected similarly by low PO2. These observations strongly suggest that KO2 channels are the main contributors to the macroscopic K+ current of glomus cells. The reversible inhibition of KO2 channel activity by low PO2 does not desensitize and is not related to the presence of F-, ATP, and GTP-gamma-S at the internal face of the membrane. These results indicate that KO2 channels confer upon glomus cells their unique chemoreceptor properties and that the O2-K+ channel interaction occurs either directly or through an O2 sensor intrinsic to the plasma membrane closely associated with the channel molecule.

scholarly journals Gating of Recombinant Small-Conductance Ca-activated K+ Channels by Calcium

1998 ◽  
Vol 111 (4) ◽  
pp. 565-581 ◽  
Author(s):  
Birgit Hirschberg ◽  
James Maylie ◽  
John P. Adelman ◽  
Neil V. Marrion
Keyword(s):  
Time Course ◽  
Single Channel ◽  
Cell Types ◽  
K Channel ◽  
Open Probability ◽  
K Channels ◽  
Sensitive Clone ◽  
Single Channel Currents ◽  
Open States ◽  
Small Conductance

Small-conductance Ca-activated K+ channels play an important role in modulating excitability in many cell types. These channels are activated by submicromolar concentrations of intracellular Ca2+, but little is known about the gating kinetics upon activation by Ca2+. In this study, single channel currents were recorded from Xenopus oocytes expressing the apamin-sensitive clone rSK2. Channel activity was detectable in 0.2 μM Ca2+ and was maximal above 2 μM Ca2+. Analysis of stationary currents revealed two open times and three closed times, with only the longest closed time being Ca dependent, decreasing with increasing Ca2+ concentrations. In addition, elevated Ca2+ concentrations resulted in a larger percentage of long openings and short closures. Membrane voltage did not have significant effects on either open or closed times. The open probability was ∼0.6 in 1 μM free Ca2+. A lower open probability of ∼0.05 in 1 μM Ca2+ was also observed, and channels switched spontaneously between behaviors. The occurrence of these switches and the amount of time channels spent displaying high open probability behavior was Ca2+ dependent. The two behaviors shared many features including the open times and the short and intermediate closed times, but the low open probability behavior was characterized by a different, long Ca2+-dependent closed time in the range of hundreds of milliseconds to seconds. Small-conductance Ca- activated K+ channel gating was modeled by a gating scheme consisting of four closed and two open states. This model yielded a close representation of the single channel data and predicted a macroscopic activation time course similar to that observed upon fast application of Ca2+ to excised inside-out patches.

Evidence for KCNQ1 K+ channel expression in rat zymogen granule membranes and involvement in cholecystokinin-induced pancreatic acinar secretion

2008 ◽  
Vol 294 (4) ◽  
pp. C879-C892 ◽  
Author(s):  
Wing-Kee Lee ◽  
Blazej Torchalski ◽  
Eleni Roussa ◽  
Frank Thévenod
Keyword(s):  
Single Channel ◽  
Reversal Potential ◽  
Fluid Secretion ◽  
Pancreatic Tissue ◽  
Bk Channel ◽  
K Channel ◽  
Amylase Secretion ◽  
Zymogen Granule ◽  
Open Probability ◽  
Pancreatic Acini

Secretion of enzymes and fluid induced by Ca2+ in pancreatic acini is not completely understood and may involve activation of ion conductive pathways in zymogen granule (ZG) membranes. We hypothesized that a chromanol 293B-sensitive K+ conductance carried by a KCNQ1 protein is expressed in ZG membranes (ZGM). In suspensions of rat pancreatic ZG, ion flux was determined by ionophore-induced osmotic lysis of ZG suspended in isotonic salts. The KCNQ1 blocker 293B selectively blocked K+ permeability (IC50 of ∼10 μM). After incorporation of ZGM into planar bilayer membranes, cation channels were detected in 645/150 mM potassium gluconate cis/trans solutions. Channels had linear current-voltage relationships, a reversal potential ( Erev) of −20.9 ± 0.9 mV, and a single-channel K+ conductance ( gK) of 265.8 ± 44.0 pS ( n = 39). Replacement of cis 500 mM K+ by 500 mM Na+ shifted Erev to −2.4 ± 3.6 mV ( n = 3), indicating K+ selectivity. Single-channel analysis identified several K+ channel groups with distinct channel behaviors. K+ channels with a gK of 651.8 ± 88.0 pS, Erev of −22.9 ± 2.2 mV, and open probability ( Popen) of 0.43 ± 0.06 at 0 mV ( n = 6) and channels with a gK of 155.0 ± 11.4 pS, Erev of −18.3 ± 1.8 mV, and Popen of 0.80 ± 0.03 at 0 mV ( n = 3) were inhibited by 100 μM 293B or by the more selective inhibitor HMR-1556 but not by the maxi-Ca2+-activated K+ channel (BK channel) inhibitor charybdotoxin (5 nM). KCNQ1 protein was demonstrated by immunoperoxidase labeling of pancreatic tissue, immunogold labeling of ZG, and immunoblotting of ZGM. 293B also inhibited cholecystokinin-induced amylase secretion of permeabilized acini (IC50 of ∼10 μM). Thus KCNQ1 may account for ZG K+ conductance and contribute to pancreatic hormone-stimulated enzyme and fluid secretion.

Ba(2+)-sensitive K+ channels in basal membrane of confluent Madin-Darby canine kidney cells

1994 ◽  
Vol 267 (6) ◽  
pp. F1007-F1014
Author(s):  
X. Y. Wang ◽  
P. J. Harris ◽  
R. E. Kemm
Keyword(s):  
Single Channel ◽  
Mdck Cells ◽  
Physiological Role ◽  
K Channel ◽  
Madin Darby Canine Kidney ◽  
Open Probability ◽  
Cytosolic Ph ◽  
K Channels ◽  
Darby Canine Kidney ◽  
Canine Kidney

A method is described for gaining access to the basolateral membranes of confluent Madin-Darby canine kidney (MDCK) cells by surgical reflection of the cell layer overlying fluid-filled domes. Single-channel recordings from cell-attached inside-out and outside-out configurations revealed two K+ channels located in the basal membranes of the highly differentiated monolayers. With 140 mmol/l KCl in pipette, the intermediate-conductance K+ channel displayed outward rectification in cell-attached configuration with channel conductances of 65 pS for outward part and 17 pS for inward part. In excised-patch recording, this channel had a conductance of 92 pS with 140 mmol/l KCl on the extracellular side of the patch and 5 mmol/l KCl on the cytosolic side. The maximum conductance obtained in symmetrical KCl (140 mmol/l) solution was 140 pS. Ba2+ (1 mmol/l) and tetraethylammonium (5 mmol/l) blocked this channel reversibly. Channel open probability (Po) was reduced from 0.41 at cytosolic pH 7.4 to 0.14 at pH 6.8 and increased to 0.64 at pH 8.0. The channel activity was significantly inhibited by elevation of intracellular Ca2+. A small-conductance K+ channel was also observed mainly in excised patches with single-channel conductance of 48 pS in symmetrical KCl solutions. However, the activity of this channel was partially obscured by the intermediate-conductance K+ channel and further analysis was not possible. A physiological role of these channels in mediating K+ recycling through the monolayer is suggested.

Large conducting potassium channel reconstituted from airway smooth muscle

1992 ◽  
Vol 262 (3) ◽  
pp. L327-L336 ◽  
Author(s):  
D. Savaria ◽  
C. Lanoue ◽  
A. Cadieux ◽  
E. Rousseau
Keyword(s):  
Smooth Muscle ◽  
Lipid Bilayers ◽  
Airway Smooth Muscle ◽  
Single Channel ◽  
Specific Inhibitor ◽  
Physiological Role ◽  
Membrane Receptors ◽  
K Channel ◽  
Open Probability ◽  
K Channels

Microsomal fractions were prepared from canine and bovine airway smooth muscle (ASM) by differential and gradient centrifugations. Surface membrane vesicles were characterized by binding assays and incorporated into planar lipid bilayers. Single-channel activities were recorded in symmetric or asymmetric K+ buffer systems and studied under voltage and Ca2+ clamp conditions. A large-conductance K(+)-selective channel (greater than 220 pS in 150 mM K+) displaying a high Ca2+, low Ba2+, and charybdotoxin (CTX) sensitivity was identified. Time analysis of single-channel recordings revealed a complex kinetic behavior compatible with the previous schemes proposed for Ca(2+)-activated K+ channels in a variety of biological surface membranes. We now report that the open probability of the channel at low Ca2+ concentration is enhanced on in vitro phosphorylation, which is mediated via an adenosine 3',5'-cyclic monophosphate-dependent protein kinase. In addition to this characterization at the molecular level, a second series of pharmacological experiments were designed to assess the putative role of this channel in ASM strips. Our results show that 50 nM CTX, a specific inhibitor of the large conducting Ca(2+)-dependent K+ channel, prevents norepinephrine transient relaxation on carbamylcholine-precontracted ASM strips. It was also shown that CTX reversed the steady-state relaxation induced by vasoactive intestinal peptide and partially antagonized further relaxation induced by cumulative doses of this potent bronchodilatator. Thus it is proposed that the Ca(2+)-activated K+ channels have a physiological role because they are indirectly activated on stimulation of various membrane receptors via intracellular mechanisms.

Potent inhibition of the aortic smooth muscle maxi-K channel by clinical doses of ethanol

2000 ◽  
Vol 279 (4) ◽  
pp. C1107-C1115 ◽  
Author(s):  
F. S. Walters ◽  
M. Covarrubias ◽  
J. S. Ellingson
Keyword(s):  
Smooth Muscle ◽  
Lipid Bilayers ◽  
Single Channel ◽  
K Channel ◽  
Open Probability ◽  
K Channels ◽  
Aortic Smooth Muscle ◽  
Slope Factor ◽  
The Mean ◽  
Relationship Change

We investigated the effects of clinically relevant ethanol concentrations (5–20 mM) on the single-channel kinetics of bovine aortic smooth muscle maxi-K channels reconstituted in lipid bilayers (1:1 palmitoyl-oleoyl-phosphatidylethanolamine: palmitoyl-oleoyl-phosphatidylcholine). Ethanol at 10 and 20 mM decreased the channel open probability ( P o) by 75 ± 20.3% mainly by increasing the mean closed time (+82 to +960%, n = 7). In some instances, ethanol also decreased the mean open time (−40.8 ± 22.5%). The P o-voltage relation in the presence of 20 mM ethanol exhibited a rightward shift in the midpoint of voltage activation (Δ V ½ ≅ 17 mV), a slightly steeper relationship (change in slope factor, Δ k, ≅ −2.5 mV), and a decreased maximum P o (from ∼0.82 to ∼0.47). Interestingly, channels inhibited by ethanol at low Ca2+ concentrations (2.5 μM) were very resistant to ethanol in the presence of increased Ca2+ (≥ 20 μM). Alcohol consumption in clinically relevant amounts may alter the contribution of maxi-K channels to the regulation of arterial tone.

Na+-induced inward rectification in the two-pore domain K+ channel, TASK-2

2005 ◽  
Vol 288 (1) ◽  
pp. F162-F169 ◽  
Author(s):  
Michael J. Morton ◽  
Sarah Chipperfield ◽  
Abdulrahman Abohamed ◽  
Asipu Sivaprasadarao ◽  
Malcolm Hunter
Keyword(s):  
Single Channel ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Selectivity Filter ◽  
Inward Rectification ◽  
K Channel ◽  
Open Probability ◽  
Whole Cell ◽  
Single Channel Currents ◽  
Pore Domain

TASK-2 is a member of the two-pore domain K+ (K2P) channel family that is expressed at high levels in several epithelia, including the proximal tubule. In common with the other TASK channels, TASK-2 is sensitive to changes in extracellular pH. We have expressed human TASK-2 in Chinese hamster ovary cells and studied whole cell and single-channel activity by patch clamp. The open probability of K2P channels is generally independent of voltage, yielding linear current-voltage ( I- V) curves. Despite these properties, we found that these channels showed distinct inward rectification immediately on the establishment of whole cell clamp, which became progressively less pronounced with time. This rectification was due to intracellular Na+ but was unaffected by polyamines or Mg2+ (agents that cause rectification in Kir channels). Rectification was concentration- and voltage-dependent and could be reversibly induced by switching between Na+-rich and Na+-free bath solutions. In excised inside-out patches, Na+ reduced the amplitude of single-channel currents, indicative of rapid block and unblock of the pore. Mutations in the selectivity filter abolished Na+-induced rectification, suggesting that Na+ binds within the selectivity filter in wild-type channels. This sensitivity to intracellular Na+ may be an additional potential regulatory mechanism of TASK-2 channels.

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