Inwardly rectifying chloride channel activity in intestinal pacemaker cells

2005 ◽  
Vol 288 (4) ◽  
pp. G809-G821 ◽  
Author(s):  
Yaohui Zhu ◽  
Andrea Mucci ◽  
Jan D. Huizinga
Keyword(s):  
Single Channel ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Pacemaker Activity ◽  
Channel Activity ◽  
Pacemaker Cells ◽  
Open Probability ◽  
Whole Cell ◽  
Single Channel Activity ◽  
Inwardly Rectifying

Cl− channels are proposed to play a role in gut pacemaker activity, but little is known about the characteristics of Cl− channels in interstitial cells of Cajal (ICC), the intestinal pacemaker cells. The objective of the present study was to identify whole cell Cl− currents in ICC associated with previously observed single-channel activity and to characterize its inward rectification. Whole cell patch-clamp studies showed that ICC express an inwardly rectifying Cl− current that was not sensitive to changes in cation composition of the extracellular solutions. Currents were not affected by replacing all cations with N-methyl-d-glucamine (NMDG+). Whole cell currents followed the Cl− equilibrium potential and were inhibited by DIDS and 9-anthracene carboxylic acid. Ramp protocols of single-channel activity showed that inward rectification was due to reduction in single-channel open probability, not a reduction in single-channel conductance. Single-channel data led to the hypothesis that strong cooperation exists between 30-pS channels that show less cooperation at potentials positive to the reversal potential. Hence, an inwardly rectifying Cl− channel plays a prominent role in determining pacemaker activity in the gut.

Hypoxia modulates nitric oxide-induced regulation of NMDA receptor currents and neuronal cell death

1999 ◽  
Vol 277 (4) ◽  
pp. C673-C683 ◽  
Author(s):  
Muyiwa Gbadegesin ◽  
Stefano Vicini ◽  
Sandra J. Hewett ◽  
David A. Wink ◽  
Michael Espey ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Cell Death ◽  
Steady State ◽  
Nmda Receptor ◽  
Single Channel ◽  
Receptor Activation ◽  
Channel Activity ◽  
Open Probability ◽  
Whole Cell ◽  
Nmda Channel

Nitric oxide (NO) released from a new chemical class of donors enhances N-methyl-d-aspartate (NMDA) channel activity. Using whole cell and single-channel patch-clamp techniques, we have shown that ( Z)-1-[ N-(3-ammoniopropyl)- N-( n-propyl)amino]-NO (PAPA-NO) and diethylamine NO, commonly termed NONOates, potentiate the glutamate-mediated response of recombinant rat NMDA receptors (NR1/NR2A) expressed in HEK-293 cells. The overall effect is an increase in both peak and steady-state whole cell currents induced by glutamate. Single-channel studies demonstrate a significant increase in open probability but no change in the mean single-channel open time or mean channel conductance. Reduction in oxygen levels increased and prolonged the PAPA-NO-induced change in both peak and steady-state glutamate currents in transfected HEK cells. PAPA-NO also enhanced cell death in primary cultures of rodent cortical neurons deprived of oxygen and glucose. This potentiation of neuronal injury was blocked by MK-801, indicating a critical involvement of NMDA receptor activation. The NO-induced increase in NMDA channel activity as well as NMDA receptor-mediated cell death provide firm evidence that NO modulates the NMDA channel in a manner consistent with both a physiological role under normoxic conditions and a pathophysiological role under hypoxic conditions.

Abstract 174: Biophysical Basis of Protein Kinase C Inhibition of the Novel alpha1D Calcium Channel

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Mohamed Chahine ◽  
Yongxia Qu ◽  
Mohamed Boutjdir
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Calcium Channel ◽  
Single Channel ◽  
Ca Channel ◽  
Channel Activity ◽  
Open Probability ◽  
Whole Cell ◽  
Kinase C ◽  
Pkc Activation

The recently reported α 1D calcium channel in the heart is known to be regulated by protein kinase C (PKC) at the whole cell level and has been implicated in atrial fibrillation. The biophysical basis of this regulation at the single channel level is not known. Therefore, the effect of PKC activation was studied on α 1D calcium channel expressed in tsA201 cells using cell-attached method. Unitary currents were recorded in the presence of 70 mM Ba 2+ as the charge carrier. Unitary currents were evoked by 500 ms depolarizing pulses from a holding potential of −80 mV every 0.5 Hz. Under basal condition, channel activity was rare and infrequent, however Bay K 8644 (1 μM) induced channel openings with a conductance of 22.3 pS. Single channel analysis of open and closed time distributions were best fitted with a single exponential. PKC activation by PMA (10 nM), a phorbol ester derivative, resulted in a decrease in open probability and increase in closed-time without any significant effect on the conductance of the α 1D calcium channel. This is consistent with a decreased entry of α 1D Ca channel into open states in the presence of PMA. These data show, for the fist time, 1) the α 1D calcium channel activity at the single channel level and 2) the biophysical basis of by which PKC activation inhibits the α 1D calcium channel. The shortening of the open-time and the lengthening of the closed-time constants and the increase in blank sweeps may explain the inhibition of the α 1D Ca-channel activity and the reduction in whole-cell α 1D Ca current previously reported. Altogether, these data are relevant to the understanding of the patho-physiology of α 1D calcium channel and its regulation by the autonomics.

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.

Peroxynitrite reversibly inhibits Ca2+-activated K+ channels in rat cerebral artery smooth muscle cells

2000 ◽  
Vol 278 (6) ◽  
pp. H1883-H1890 ◽  
Author(s):  
Anna K. Brzezinska ◽  
Debebe Gebremedhin ◽  
William M. Chilian ◽  
Balaraman Kalyanaraman ◽  
Stephen J. Elliott
Keyword(s):  
Single Channel ◽  
Cerebral Arteries ◽  
Ionic Current ◽  
Inhibitory Effect ◽  
Bk Channels ◽  
Bk Channel ◽  
Channel Activity ◽  
Open Probability ◽  
Whole Cell ◽  
Cell Current

Peroxynitrite (ONOO−) is a contractile agonist of rat middle cerebral arteries. To determine the mechanism responsible for this component of ONOO−bioactivity, the present study examined the effect of ONOO− on ionic current and channel activity in rat cerebral arteries. Whole cell recordings of voltage-clamped cells were made under conditions designed to optimize K+ current. The effects of iberiotoxin, a selective inhibitor of large-conductance Ca2+-activated K+ (BK) channels, and ONOO− (10–100 μM) were determined. At a pipette potential of +50 mV, ONOO− inhibited 39% of iberiotoxin-sensitive current. ONOO− was selective for iberiotoxin-sensitive current, whereas decomposed ONOO− had no effect. In excised, inside-out membrane patches, channel activity was recorded using symmetrical K+solutions. Unitary currents were sensitive to increases in internal Ca2+ concentration, consistent with activity due to BK channels. Internal ONOO− dose dependently inhibited channel activity by decreasing open probability and mean open times. The inhibitory effect of ONOO− could be overcome by reduced glutathione. Glutathione, added after ONOO−, restored whole cell current amplitude to control levels and reverted single-channel gating to control behavior. The inhibitory effect of ONOO− on membrane K+ current is consistent with its contractile effects in isolated cerebral arteries and single myocytes. Taken together, our data suggest that ONOO− has the potential to alter cerebral vascular tone by inhibiting BK channel activity.

Biophysical and pharmacological characterization of inwardly rectifying K+ currents in rat spinal cord astrocytes

1995 ◽  
Vol 73 (1) ◽  
pp. 333-346 ◽  
Author(s):  
C. B. Ransom ◽  
H. Sontheimer
Keyword(s):  
Spinal Cord ◽  
Single Channel ◽  
Resting Potential ◽  
Equilibrium Potential ◽  
Dependent Manner ◽  
Rat Spinal Cord ◽  
Open Probability ◽  
Kir Channels ◽  
Inwardly Rectifying ◽  
K Currents

1. Whole cell and cell-attached patch-clamp recordings were obtained from rat spinal cord astrocytes maintained in culture for 6-14 days. It was found that the resting conductance in these astrocytes is primarily due to inwardly rectifying K+ (Kir) channels. 2. Two types of astrocytic Kir channels were identified with single-channel conductances of approximately 28 and approximately 80 pS, respectively. Channels displayed some voltage dependence in their open probability, which was largest (0.8-0.9) near the K+ equilibrium potential (Ek) and decreased at more negative potentials. The resting potential closely followed Ek, so it can be assumed that Kir channels have a high open probability at the resting potential. 3. The conductance of inwardly rectifying K+ currents (Kir) depended strongly on [K+]o and was approximately proportional to the square-root of [K+]o. 4. Kir currents inactivated in a time- and voltage-dependent manner. The Na+ dependence of inactivation was studied with ion substitution experiments. Replacement of [Na+]o with choline or Li+ removed inactivation. This dependence of current inactivation on [Na+]o resembles the previously described block of Kir channels in other systems by [Na+]o. 5. Kir currents were also blocked in a dose-dependent manner by Cs+ (Kd = 189 microM at -140 mV), Ba2+ (Kd = 3.5 microM), and tetraethylammonium (TEA; 90% block at 10 mM) but were insensitive to 4-aminopyridine (4-AP; 5 mM). In the current-clamp mode, Ba2+ and TEA inhibition of Kir currents was associated with a marked depolarization, suggesting that Kir channel activity played a role in the establishment of the negative resting potential typical of astrocytes. 6. These biophysical features of astrocyte inwardly rectifying K+ channels are consistent with those properties required for their proposed involvement in [K+]o clearance: 1) high open probability at the resting potential, 2) increasing conductance with increasing [K+]o, and 3) rectification, e.g., channel closure, at positive potentials. It is proposed, therefore, that the dissipation of [K+]o following neuronal activity is mediated primarily by the activity of astrocytic Kir channels.

Acetylcholine-sensitive potassium channels in human atrial myocytes

1990 ◽  
Vol 259 (6) ◽  
pp. H1730-H1735 ◽  
Author(s):  
R. Sato ◽  
I. Hisatome ◽  
J. A. Wasserstrom ◽  
C. E. Arentzen ◽  
D. H. Singer
Keyword(s):  
Single Channel ◽  
K Channel ◽  
Channel Activity ◽  
Atrial Myocytes ◽  
Open Probability ◽  
Human Atrial Myocytes ◽  
Open Time ◽  
Patch Pipette ◽  
Single Channel Activity ◽  
Gamma S

Single channel recording techniques were used to study acetylcholine (ACh)-sensitive K+ channel activity in human atrial myocytes isolated from specimens obtained during corrective cardiac surgery. Under conditions of cell-attached patch, the presence of ACh in the patch pipette activated K+ channels. Single channel activity occurred in periodic bursts. The channels exhibited a slope conductance of 46 +/- 2 pS inwardly (means +/- SD, n = 4). During a burst, both open and closed time histograms were fitted by a single exponential curve, suggesting the existence of one open and one closed state during a burst. Open probability increased directly with ACh concentration without affecting open time. The channel could be activated by GTP and guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) (in the presence and absence of ACh in the pipette, respectively). Slope conductance, the response to GTP and GTP gamma S, and the independence of activation from Ca2+ were similar to those for other species. In contrast, sensitivity to ACh appeared diminished compared with frog atrial myocytes.

Single-channel properties of a G-protein-coupled inward rectifier potassium channel in brain neurons

1996 ◽  
Vol 75 (1) ◽  
pp. 318-328 ◽  
Author(s):  
J. J. Grigg ◽  
T. Kozasa ◽  
Y. Nakajima ◽  
S. Nakajima
Keyword(s):  
G Protein ◽  
Single Channel ◽  
Inward Rectifier ◽  
Equilibrium Potential ◽  
K Channel ◽  
Open Probability ◽  
K Concentration ◽  
Inwardly Rectifying ◽  
G Protein Coupled ◽  
Cytoplasmic Side

1. In cultured rat locus coeruleus neurons, somatostatin or met-enkephalin induces an inwardly rectifying K+ conductance. This inward rectifier was analyzed at the single-channel level. 2. Using the inside-out patch-clamp, guanosine 5'-triphosphate (GTP) application to the cytoplasmic side in the presence of somatostatin or met-enkephalin in the pipette produced a large increase in channel activity, which disappeared on switching from GTP to guanosine 5'-diphosphate. 3. The unitary conductance was approximately 30 pS at -95 mV with an extracellular K+ concentration of 156 mM and an intracellular K+ concentration of 124 mM at 23 degrees C. The channel showed burst behavior, and the closed time histogram was fit by two exponentials, with the fast time constant being 0.4 ms. The burst time histogram was also fit by two exponentials, with time constants of 0.24 and 2.0 ms (at 10 nM somatostatin). When the somatostatin concentration was changed from 500 to 1 nM, the kinetic behavior of the channel did not change, except that the open probability of the patch was decreased. 4. The current-voltage relation of the unitary channel current showed inward rectification. The reversal potential coincided with the K+ equilibrium potential, and it shifted according to a change in the K+ equilibrium potential. 5. In the presence of external somatostatin, the application of guanosine 5'-O-(3-thiotriphosphate) to the cytoplasmic side induced an irreversible activation of this channel. 6. These results indicate that this K+ channel is the microscopic counterpart of the somatostatin- or met-enkephalin-induced inwardly rectifying K+ current in whole cell recording, and that the channel is activated by a G protein without a diffusible second messenger. Thus this channel is identified as a neuronal G-protein-coupled inward rectifier K+ channel. 7. Analysis of the burst behavior, based on a close-close-open kinetic model, revealed that there are at least four states in the K+ channel, a short gap, a longer closing, a short opening, and a long opening, and that the neuronal inward rectifier is activated at faster rates than the atrial inward rectifier.

ATP-sensitive K+ channels are altered in hypertrophied ventricular myocytes

1988 ◽  
Vol 255 (5) ◽  
pp. H1254-H1258 ◽  
Author(s):  
J. S. Cameron ◽  
S. Kimura ◽  
D. A. Jackson-Burns ◽  
D. B. Smith ◽  
A. L. Bassett
Keyword(s):  
Single Channel ◽  
Single Cells ◽  
Membrane Surface ◽  
Resting Potential ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
K Channel ◽  
Channel Activity ◽  
Protective Function ◽  
In Cells

Unitary K+ currents in single cells isolated from normal and hypertrophied feline left ventricles were studied with regard to ATP sensitivity using patch-clamp single-channel recording. Data were obtained from excised inside-out membrane patches with symmetrical transmembrane K+ concentration [K+] (140 mM) at 22 +/- 1 degree C and in the absence of divalent cations. In the absence of ATP at the intracellular membrane surface, K+ channel activity was observed during depolarizing and hyperpolarizing pulses ranging from 10 to 120 mV from the K+ equilibrium potential. The current voltage (I-V) curve displayed some inward rectification, with slope conductances becoming nonlinear at strong depolarizations. Rectification was particularly pronounced in cells from hypertrophied left ventricles relative to normal. Single-channel conductance determined from the linear portion of the I-V curve was 77 pS in both groups. The channels were blocked by intracellular Ca2+ (1 mM), extracellular tetraethylammonium (TEA; less than or equal to 2 mM) or 4-aminopyridine (0.5 mM); 2 mM ATP produced a total but reversible inhibition. The effect of ATP was to reduce channel openings; conductance was unaffected. The ATP concentration [( ATP]) that induced half-maximal inhibition of channel activity was 75 microM in normal myocytes but was 250 microM in cells from hypertrophied hearts. Rapid channel activation at diminished [ATP] may provide a protective function by maintaining resting potential or promoting vasodilation in hypertrophied myocardium.

A calcium-activated chloride channel in sarcoplasmic reticulum vesicles from rabbit skeletal muscle

1996 ◽  
Vol 270 (6) ◽  
pp. C1675-C1686 ◽  
Author(s):  
J. I. Kourie ◽  
D. R. Laver ◽  
G. P. Ahern ◽  
A. F. Dulhunty
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Lipid Bilayers ◽  
Chloride Channel ◽  
Single Channel ◽  
Single Channels ◽  
Channel Activity ◽  
Open Probability ◽  
Single Channel Activity ◽  
Maximum Conductance

A Ca(2+)-activated Cl- channel is described in sarcoplasmic reticulum (SR) enriched vesicles of skeletal muscle incorporated into lipid bilayers. Small chloride (SCl) channels (n = 20) were rapidly and reversibly activated when cis- (cytoplasmic) [Ca2+] was increased above 10(-7) M, with trans-(luminal) [Ca2+] at either 10(-3) or 10(-7) M. The open probability of single channels increased from zero when cis-[Ca2+] was 10(-7) M to 0.61 +/- 0.12 when [Ca2+] was 10(-4) M. High- and low-conductance levels in single-channel activity were activated at different cis-[Ca2+]. Channel openings to the maximum conductance, 65-75 pS (250/50 mM Cl-, cis/ trans), were active when cis-[Ca2+] was increased above 5 x 10(-6) M. In contrast to the maximum conductance, channel openings to submaximal levels between 5 and 40 pS were activated at a lower cis-[Ca2+] and dominated channel activity between 5 x 10(-7) and 5 x 10(-6) M. Activation of SCl channels was Ca2+ specific and not reproduced by cytoplasmic Mg2+ concentrations of 10(-3) M. We suggest that the SCl channel arises in the SR membrane. The Ca2+ dependence of this channel implies that it is active at [Ca2+] achieved during muscle contraction.

Activation of cardiac ryanodine receptors by the calcium channel agonist FPL-64176

2002 ◽  
Vol 283 (1) ◽  
pp. H331-H338 ◽  
Author(s):  
J. Andrew Wasserstrom ◽  
Leslie A. Wasserstrom ◽  
Andrew J. Lokuta ◽  
James E. Kelly ◽  
Sireen T. Reddy ◽  
...  
Keyword(s):  
Single Channel ◽  
Ryanodine Receptors ◽  
Channel Activity ◽  
Open Probability ◽  
Release Channel ◽  
Cardiac Sarcoplasmic Reticulum ◽  
Significant Difference ◽  
Single Channel Activity ◽  
Control Versus ◽  
Channel Agonist

We investigated the possibility that the Ca2+ channel agonist FPL-64176 (FPL) might also activate the cardiac sarcoplasmic reticulum (SR) Ca2+ release channel ryanodine receptor (RyR). The effects of FPL were tested on single channel activity of purified and crude vesicular RyR (RyR2) isolated from human and dog hearts using the planar lipid bilayer technique. FPL (100–200 μM) increased single channel open probability ( P o) when added to the cytoplasmic side of the channel ( P o = 0.070 ± 0.021 in control RyR2; 0.378 ± 0.086 in 150 μM FPL, n = 9, P < 0.01) by prolonging open times and decreasing closed times without changing current magnitude. FPL had no effect on P o when added to the trans (luminal) side of the bilayer ( P o = 0.079 ± 0.036 in control and 0.103 ± 0.066 in FPL, n = 4, no significant difference). The bell-shaped [Ca2+] dependence of [3H]ryanodine binding and of P o was altered by FPL, suggesting that the mechanism by which FPL increases channel activity is by an increase in Ca2+-induced activation at low [Ca2+] (without a change in threshold) and suppression of Ca2+-induced inactivation at high [Ca2+]. However, the fact that inactivation was restored at elevated [Ca2+] suggests a competitive interaction between Ca2+ and FPL on inactivation. FPL had no effect on RyR skeletal channels (RyR1), where P o was 0.039 ± 0.005 in control versus 0.030 ± 0.006 in 150 μM FPL (no significant difference). These results suggest that, in addition to its ability to activate the L-type Ca2+channels, FPL activates cardiac RyR2 primarily by reducing the Ca2+ sensitivity of inactivation.

Sign in / Sign up

Export Citation Format

Share Document