Basolateral potassium channels in renal proximal tubule

1987 ◽  
Vol 253 (3) ◽  
pp. F476-F487 ◽  
Author(s):  
H. Sackin ◽  
L. G. Palmer
Keyword(s):  
Single Channel ◽  
Basolateral Membrane ◽  
K Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
K Channels ◽  
Open Time ◽  
Excised Patches ◽  
The Mean ◽  
Inside Out

Potassium (K+) channels in the basolateral membrane of unperfused Necturus proximal tubules were studied in both cell-attached and excised patches, after removal of the tubule basement membrane by manual dissection without collagenase. Two different K+ channels were identified on the basis of their kinetics: a short open-time K+ channel, with a mean open time less than 1 ms, and a long open-time K+ channel with a mean open time greater than 20 ms. The short open-time channel occurred more frequently than the longer channel, especially in excised patches. For inside-out excised patches with Cl- replaced by gluconate, the current-voltage relation of the short open-time K+ channel was linear over +/- 60 mV, with a K+-Na+ selectivity of 12 +/- 2 (n = 12), as calculated from the reversal potential with oppositely directed Na+ and K+ gradients. With K-Ringer in the patch pipette and Na-Ringer in the bath, the conductance of the short open-time channel was 47 +/- 2 pS (n = 15) for cell-attached patches, 26 +/- 2 pS (n = 15) for patches excised (inside out) into Na-Ringer, and 36 +/- 6 pS (n = 3) for excised patches with K-Ringer on both sides. These different conductances can be partially explained by a dependence of single-channel conductance on the K+ concentration on the interior side of the membrane. In experiments with a constant K+ gradient across excised patches, large changes in Na+ at the interior side of the membrane produced no change in single-channel conductance, arguing against a direct block of the K+ channel by Na+. Finally, the activity of the short open-time channel was voltage gated, where the mean number of open channels decreased as a linear function of basolateral membrane depolarization for potentials between -60 and 0 mV. Depolarization from -60 to -40 mV decreased the mean number of open K+ channels by 28 +/- 8% (n = 6).

Electrical characteristics of inward-rectifying K+ channels in isolated bullfrog oxyntic cells

1991 ◽  
Vol 261 (2) ◽  
pp. G206-G212 ◽  
Author(s):  
H. Mieno ◽  
G. Kajiyama
Keyword(s):  
Single Channel ◽  
Rana Catesbeiana ◽  
Basolateral Membrane ◽  
Inward Rectification ◽  
K Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Open Probability ◽  
K Channels ◽  
Patch Clamp Method

The properties of K+ channels in the isolated oxyntic cells of the bullfrog (Rana catesbeiana) were investigated using the patch-clamp method. Two types of K+ channels on the basolateral membrane were identified on the basis of their electrophysiological and pharmacological properties. The K+ channel most frequently observed has a single-channel conductance of 61.0 +/- 2.9 pS (n = 10) and is activated by an increase in intracellular Ca2+. The other K+ channel has a single-channel conductance of 30.3 +/- 2.7 pS (n = 7), which is activated by adenosine 3',5'-cyclic monophosphate (cAMP). The physiological and pharmacological characteristics common to the two K+ channels are inward-going rectification with a high selectivity for K+ and indirect inhibition by omeprazole. The inward rectification is controlled by intracellular Mg2+ in such a way that the more Mg2+ is applied intracellularly, the more their inward-rectifying property is enhanced. The finding that bethanechol and cAMP increase the open probability of these K+ channels as well as activating the acid secretion indicates that there may be a relationship between these two processes in the oxyntic cells.

ATP-sensitive K(+)-selective channels of inner medullary collecting duct cells

1994 ◽  
Vol 267 (3) ◽  
pp. F489-F496 ◽  
Author(s):  
S. C. Sansom ◽  
T. Mougouris ◽  
S. Ono ◽  
T. D. DuBose
Keyword(s):  
Single Channel ◽  
Collecting Duct ◽  
K Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Inner Medullary Collecting Duct ◽  
Outward Currents ◽  
Inward Currents ◽  
Excised Patches ◽  
Medullary Collecting Duct

The inner medullary collecting duct (IMCD) in vivo has the capacity to either secrete or reabsorb K+. However, a selective K+ conductance has not been described previously in the IMCD. In the present study, the patch-clamp method was used to determine the presence and properties of K(+)-selective channels in the apical membrane of the inner medullary collecting duct cell line, mIMCD-3. Two types of K(+)-selective channels were observed in both cell-attached and excised patches. The most predominant K+ channel, a smaller conductance K+ channel (SK), was present in cell-attached patches with 140 mM KCl (high bath K+) but not with 135 mM NaCl plus 5 mM KCl (low bath K+) in the bathing solution. The single-channel conductance of SK was 36 pS with inward currents and 29 pS with outward currents in symmetrical 140 mM KCl. SK was insensitive to both voltage and Ca2+. However, SK was inhibited significantly by millimolar concentrations of ATP in excised patches. A second K(+)-selective channel [a larger K+ channel (BK)] displayed a single-channel conductance equal to 132 pS with inward currents and 90 pS with outward currents in symmetrical 140 mM KCl solutions. BK was intermittently activated in excised inside-out patches by Mg(2+)-ATP in concentrations from 1 to 5 mM. With complete removal of Mg2+, BK was insensitive to ATP. BK was also insensitive to potential and Ca2+ and was observed in cell-attached patches with 140 mM KCl in the bath solution. Both channels were blocked reversibly by 1 mM Ba2+ from the intracellular surface but not by external Ba2+.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium-dependent inactivation of inwardly rectifying K+ channel in a tumor mast cell line

1992 ◽  
Vol 262 (1) ◽  
pp. C84-C90 ◽  
Author(s):  
M. Mukai ◽  
I. Kyogoku ◽  
M. Kuno
Keyword(s):  
Mast Cell ◽  
Cell Line ◽  
Single Channel ◽  
K Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Whole Cell ◽  
Mast Cell Line ◽  
Inwardly Rectifying ◽  
Inside Out

Antigenic stimulation of rat basophilic leukemia (RBL-2H3) cells, a tumor mast cell line, is associated with an increase in intracellular free Ca2+ concentrations ([Ca2+]i) and membrane polarization. We recorded whole cell and single-channel currents through the inwardly rectifying K+ channel, a major resting conductance of cells, using the patch-clamp technique, and we examined interactions between channel activity and [Ca2+]i. With 10 microM Ca2+ in the pipette, the amplitude of whole cell currents gradually declined within 5 min to 48 +/- 13% of that immediately after rupture of the patch membrane, in the presence of 1 mM ATP which minimized intrinsic rundown. In inside-out patches, activity of the channel was reduced by increasing the concentration of Ca2+ in the internal medium, both in the presence and absence of 1 mM ATP, with no apparent change in single-channel conductance. Time-averaged mean current activity in inside-out patches in the presence of 5 microM Ca2+ was less than 50% of that with Ca2+ of 100 nM or less. These results suggest that a rise in [Ca2+]i leads to a closure of the inwardly rectifying K+ channel. In some inside-out patches, inward currents characterized by burst composed of rapid transitions between open and closed states were observed (flickering currents). Single-channel properties of the flickering currents are similar to the inwardly rectifying K+ channel except for kinetics (single-channel conductance of 24.5 +/- 7.9 pS, inward rectification, and permeability to K+).(ABSTRACT TRUNCATED AT 250 WORDS)

Chloride efflux inhibits single calcium channel open probability in vertebrate photoreceptors: Chloride imaging and cell-attached patch-clamp recordings

2000 ◽  
Vol 17 (2) ◽  
pp. 197-206 ◽  
Author(s):  
WALLACE B. THORESON ◽  
RON NITZAN ◽  
ROBERT F. MILLER
Keyword(s):  
Time Distribution ◽  
Single Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Open Probability ◽  
Pipette Solution ◽  
Open Time ◽  
The Mean ◽  
Channel Properties ◽  
Distribution Fit

The present study uses cell-attached patch-recording techniques to study the single-channel properties of Ca2+ channels in isolated salamander photoreceptors and investigate their sensitivity to reductions in intracellular Cl−. The results show that photoreceptor Ca2+ channels possess properties similar to L-type Ca2+ channels in other preparations, including (1) enhancement of openings by the dihydropyridine agonist, (−)BayK8644; (2) suppression by a dihydropyridine antagonist, nisoldipine; (3) single-channel conductance of 22 pS with 82 mM Ba2+ as the charge carrier; (4) mean open probability of 0.1; (5) open-time distribution fit with a single exponential (τ0 = 1.1 ms) consistent with a single open state; and (6) closed time distribution fit with two exponentials (τc1 = 0.7 ms, τc2 = 25.4 ms) consistent with at least two closed states. Using a Cl−-sensitive dye to measure intracellular [Cl−], it was found that perfusion with gluconate-containing, low Cl− medium depleted intracellular [Cl−]. It was therefore possible to reduce intracellular [Cl−] by perfusion with a low Cl− solution while maintaining the extracellular channel surface in high Cl− pipette solution. Under these conditions, the single-channel conductance was unchanged, but the mean open probability fell to 0.03. This reduction can account for the 66% reduction in whole-cell Ca2+ currents produced by perfusion with low Cl− solutions. Examination of the open and closed time distributions suggests that the reduction in open probability arises from increases in closed-state dwell times. Changes in intracellular [Cl−] may thus modulate photoreceptor Ca2+ channels.

scholarly journals Light-dependent K+ Channels in the Mollusc Onchidium Simple Photoreceptors Are Opened by cGMP

2002 ◽  
Vol 120 (4) ◽  
pp. 581-597 ◽  
Author(s):  
Tsukasa Gotow ◽  
Takako Nishi
Keyword(s):  
K Channel ◽  
Single Channels ◽  
Open Probability ◽  
Mean Values ◽  
K Channels ◽  
Open Time ◽  
Hyperpolarizing Response ◽  
The Mean ◽  
K Concentration ◽  
Inside Out

Light-dependent K+ channels underlying a hyperpolarizing response of one extraocular (simple) photoreceptor, Ip-2 cell, in the marine mollusc Onchidium ganglion were examined using cell-attached and inside-out patch-clamp techniques. A previous report (Gotow, T., T. Nishi, and H. Kijima. 1994. Brain Res. 662:268–272) showed that a depolarizing response of the other simple photoreceptor, A-P-1 cell, results from closing of the light-dependent K+ channels that are activated by cGMP. In the cell-attached patch recordings of Ip-2 cells, external artificial seawater (ASW) was replaced with a modified ASW containing 150 mM K+ and 200 mM Mg2+ to suppress any synaptic input and to maintain the membrane potential constant. When Ip-2 cells were equilibrated with this modified ASW, the internal K+ concentration was estimated to be 260 mM. Light-dependent single-channels in the cell-attached patch on these cells were opened by light but scarcely by voltage. After confirming the light-dependent channel activity in the cell-attached patches, an application of cGMP to the excised inside-out patches newly activated a channel that disappeared on removal of cGMP. Open and closed time distributions of this cGMP-activated channel could be described by the sum of two exponents with time constants τo1, τo2 and τc1, τc2, respectively, similar to those of the light-dependent channel. In both the channels, τo1 and τo2 in ms ranges were similar to each other, although τc2 over tens of millisecond ranges was different. τo1, τo2, and the mean open time τo were both independent of light intensity, cGMP concentration, and voltage. In both channels, the open probability increased as the membrane was depolarized, without changing any of τo2 or τo. In both, the reversal potentials using 200- and 450-mM K+-filled pipettes were close to the K+ equilibrium potentials, suggesting that both the channels are primarily K+ selective. Both the mean values of the channel conductance were estimated to be the same at 62 and 91 pS in 200- and 450-mM K+ pipettes at nearly 0 mV, respectively. Combining these findings with those in the above former report, it is concluded that cGMP is a second messenger which opens the light-dependent K+ channel of Ip-2 to cause hyperpolarization, and that the channel is the same as that of A-P-1 closed by light.

scholarly journals A voltage-clamp study of isolated stingray horizontal cell non-NMDA excitatory amino acid receptors

1989 ◽  
Vol 61 (1) ◽  
pp. 162-172 ◽  
Author(s):  
T. J. O'Dell ◽  
B. N. Christensen
Keyword(s):  
Amino Acid ◽  
Dicarboxylic Acid ◽  
Single Channel ◽  
Excitatory Amino Acid ◽  
Rank Order ◽  
Horizontal Cell ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Open Time ◽  
The Mean

1. Horizontal cells enzymatically isolated from retinas of the Atlantic stingray (Dasyatis sabina) were voltage-clamped using the patch electrode in the whole-cell mode. A rapid microsuperfusion system was used to apply excitatory amino acid agonists and antagonists. 2. The isolated cells responded to glutamate (GLU), kainate (KA), quisqualate (QA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA). Responses elicited by GLU, QA, and AMPA but not KA exhibited a concentration-dependent and concanavalin A- (Con-A) sensitive desensitization. No responses were elicited by aspartate, N-methyl-D-aspartate, or quinolinate at concentrations as high as 1.0 mM. 3. Judging from the concentration producing one-half of the maximal current response (EC50), the rank order affinities of the agonists was QA greater than or equal to GLU greater than AMPA greater than KA. Whereas KA had the lowest affinity of the agonists tested it was the most efficacious, producing the largest currents. Hill coefficients of the concentration-response data were near two for KA and GLU and near one for QA and AMPA. 4. The agonists differed in their sensitivity to various excitatory amino acid receptor antagonists. Kynurenate (KYN) produced a nearly complete block of horizontal cell responses to GLU and KA at concentrations that had little effect on QA and AMPA. Piperidine-2,3-dicarboxylic acid (cis-PDA), 1-(4-chlorobenzoyl)-piperazine-2,3-dicarboxylic acid (pCB-PzDA), and folic acid were less potent antagonists than KYN but were also better blockers of KA and GLU responses than of QA- and AMPA-elicited responses. 5. When QA, AMPA, or GLU were applied in combination with 55.0 microM KA the current was less than that produced by KA alone. The rank order potency for the inhibition of KA-elicited responses was QA greater than AMPA greater than GLU. In the presence of low concentrations of KA (1.0-20.0 microM), QA- and AMPA-elicited responses were potentiated. This potentiation was prevented by KYN. 6. Single-channel conductance and mean open time were estimated from the current noise fluctuations in the presence of agonist. The mean single-channel conductance for QA was 9 pS that was almost twice as large as the conductance for KA (5.9 pS) and GLU (5.7 pS). The mean open time in the presence of QA or GLU was approximately 1 ms, which was about one-half of that for KA (2.0 ms). 7. These results are best explained by the existence of a single receptor protein with multiple but not identical ligand-binding sites.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Biophysical and Molecular Mechanisms Underlying the Modulation of Heteromeric Kir4.1–Kir5.1 Channels by Co2 and Ph

2000 ◽  
Vol 116 (1) ◽  
pp. 33-46 ◽  
Author(s):  
Zhenjiang Yang ◽  
Haoxing Xu ◽  
Ningren Cui ◽  
Zhiqiang Qu ◽  
Sengthong Chanchevalap ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Single Channel ◽  
Ph Sensitivity ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Kir Channels ◽  
Kir Channel ◽  
Open Time ◽  
Excised Patches ◽  
Single Channel Currents

CO2 chemoreception may be related to modulation of inward rectifier K+ channels (Kir channels) in brainstem neurons. Kir4.1 is expressed predominantly in the brainstem and inhibited during hypercapnia. Although the homomeric Kir4.1 only responds to severe intracellular acidification, coexpression of Kir4.1 with Kir5.1 greatly enhances channel sensitivities to CO2 and pH. To understand the biophysical and molecular mechanisms underlying the modulation of these currents by CO2 and pH, heteromeric Kir4.1–Kir5.1 were studied in inside-out patches. These Kir4.1–Kir5.1 currents showed a single channel conductance of 59 pS with open-state probability (Popen) ∼ 0.4 at pH 7.4. Channel activity reached the maximum at pH 8.5 and was completely suppressed at pH 6.5 with pKa 7.45. The effect of low pH on these currents was due to selective suppression of Popen without evident effects on single channel conductance, leading to a decrease in the channel mean open time and an increase in the mean closed time. At pH 8.5, single-channel currents showed two sublevels of conductance at ∼1/4 and 3/4 of the maximal openings. None of them was affected by lowering pH. The Kir4.1–Kir5.1 currents were modulated by phosphatidylinositol-4,5-bisphosphate (PIP2) that enhanced baseline Popen and reduced channel sensitivity to intracellular protons. In the presence of 10 μM PIP2, the Kir4.1–Kir5.1 showed a pKa value of 7.22. The effect of PIP2, however, was not seen in homomeric Kir4.1 currents. The CO2/pH sensitivities were related to a lysine residue in the NH2 terminus of Kir4.1. Mutation of this residue (K67M, K67Q) completely eliminated the CO2 sensitivity of both homomeric Kir4.1 and heteromeric Kir4.1–Kir5.1. In excised patches, interestingly, the Kir4.1–Kir5.1 carrying K67M mutation remained sensitive to low pHi. Such pH sensitivity, however, disappeared in the presence of PIP2. The effect of PIP2 on shifting the titration curve of wild-type and mutant channels was totally abolished when Arg178 in Kir5.1 was mutated. Thus, these studies demonstrate a heteromeric Kir channel that can be modulated by both acidic and alkaline pH, show the modulation of pH sensitivity of Kir channels by PIP2, and provide information of the biophysical and molecular mechanisms underlying the Kir modulation by intracellular protons.

Activation of ATP-sensitive K channels in heart cells by pinacidil: dependence on ATP

1989 ◽  
Vol 257 (6) ◽  
pp. H2092-H2096 ◽  
Author(s):  
J. P. Arena ◽  
R. S. Kass
Keyword(s):  
Single Channel ◽  
K Channel ◽  
Heart Cells ◽  
Channel Activity ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Atp Binding Site ◽  
Atp Concentration ◽  
Ventricular Cells ◽  
Inside Out

We have investigated the effects of pinacidil on channel activity recorded from inside-out patches of membrane excised from guinea pig ventricular cells. If the cytosolic ATP concentration is greater than 0 but less than 500 microM, pinacidil increases the activity of a channel identified as the ATP-sensitive K channel (IKATP) by its single-channel conductance, its inhibition by ATP, and its sensitivity to glybenclamide. When ATP is greater than 3.0 mM the effects of pinacidil are inhibited. Our experiments show that pinacidil enhances the activity of IKATP in heart cells, but that the action of the drug depends on the ATP concentration of the cytosolic solutions. The results suggest that pinacidil acts indirectly, perhaps at an ATP-binding site that regulates this channel.

Sites of antagonist action on N-methyl-D-aspartic acid receptors studied using fluctuation analysis and a rapid perfusion technique

1988 ◽  
Vol 60 (2) ◽  
pp. 645-663 ◽  
Author(s):  
M. L. Mayer ◽  
G. L. Westbrook ◽  
L. Vyklicky
Keyword(s):  
Nmda Receptor ◽  
Kynurenic Acid ◽  
Single Channel ◽  
Nmda Antagonist ◽  
Fluctuation Analysis ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Open Time ◽  
Voltage Dependent ◽  
The Mean

1. Mouse hippocampal neurons in dissociated culture were grown at low density on previously plated hippocampal glial cell cultures and voltage clamped using the tight seal whole-cell patch-clamp technique. Flow pipes were used to rapidly exchange the extracellular solution, and to apply N-methyl-D-aspartic acid (NMDA) and some NMDA antagonists. Fluctuation analysis was used to estimate changes in the behavior of NMDA-activated ion channels during application of antagonists. In the presence of NMDA control spectra were well fit by single Lorentzian functions consistent with mean open times of 5-6 ms. 2. Two antagonists thought to act at the NMDA receptor agonist recognition site, 2-amino-5-phosphonovaleric acid (AP5) and kynurenic acid, did not produce changes in the mean open time or single channel conductance, consistent with their action as competitive antagonists. Onset of antagonism and recovery from the action of both AP5 and kynurenic acid was rapid and complete within 1 s. However, raising the extra-cellular glycine concentration, from 1 microM to 1 mM, reduced the potency of 100 microM kynurenic acid as an NMDA antagonist, suggesting that kynurenate has an additional action as a competitive antagonist at the glycine modulatory site on NMDA receptor channels. 3. In the presence of 150 microM magnesium NMDA spectra recorded at -60 mV were fit by double Lorentzian functions, consistent with single-channel events consisting of bursts of openings lasting 3.3 ms in duration, interrupted by blocking and unblocking events of average duration 0.18 ms. The onset and recovery from magnesium antagonism was rapid, and complete within 1 s, but was highly voltage dependent and at +40 mV magnesium (150 microM) failed to produce NMDA antagonism. These results are consistent with a voltage-dependent channel block of NMDA receptor channels produced by binding of magnesium to a site within the ion channel. 4. Zinc (30 microM) was a potent NMDA antagonist at both -60 and +40 mV, and at either potential appeared to reduce the mean open time of NMDA-activated ion channels from about 5 ms to approximately 3 ms. Over the frequency range measured, 1-1,000 Hz, NMDA spectra were well fit by single Lorentzians during zinc antagonism, in contrast to results obtained with magnesium. The mean single channel conductance also decreased in the presence of zinc to approximately 75% of control. Onset of antagonism and recovery from the action of zinc was rapid and complete within 1 s.(ABSTRACT TRUNCATED AT 400 WORDS)

Two distinct mechanisms are responsible for single K channel block by internal tetraethylammonium ions

1989 ◽  
Vol 256 (3) ◽  
pp. C683-C687 ◽  
Author(s):  
D. Yamamoto ◽  
N. Suzuki
Keyword(s):  
Single Channel ◽  
K Channel ◽  
Delayed Rectifier ◽  
Channel Block ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Average Current ◽  
Inside Out ◽  
Cytoplasmic Side ◽  
Tetraethylammonium Ions

Tetraethylammonium (TEA) ions blocked the unitary currents through the delayed rectifier potassium channels of Drosophila neurons from the cytoplasmic side of inside-out membrane patches by two distinct mechanisms. First, TEA attenuated the single-channel conductance, probably by producing very rapid block-unblock reactions at the inner mouth of the potassium pore. Second, TEA markedly enhanced the slow inactivation, making the incidence of channel openings highly nonrandom; blank traces with no channel openings during repetitive depolarizations showed a significant tendency to be clustered in the presence of TEA. This second action accounts for almost half of the reduction of average current produced by 10 mM internal TEA.

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