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1.
Piezo1 is a eukaryotic cation-selective mechanosensitive ion channel. To understand channel function in vivo, we first need to analyze and compare the response in the whole cell and the patch. In patches, Piezo1 inactivates and the current is fit well by a 3-state model with a single pressure-dependent rate. However, repeated stimulation led to an irreversible loss of inactivation. Remarkably, the loss of inactivation did not occur on a channel-by-channel basis but on all channels at the same time. Thus, the channels are in common mechanical domain. Divalent ions decreased the unitary conductance from ~68 pS to ~37 pS, irrespective of the cation species. Mg and Ca did not affect inactivation rates, but Zn caused a 3-fold slowing. CytochalasinD (cytoD) does not alter inactivation rates or the transition to the non-inactivating mode but does reduce the steady-state response. Whole-cell currents were similar to patch currents but also had significant differences. In contrast to the patch, cytoD inhibited the current suggesting that the activating forces were transmitted through the actin cytoskeleton. Hypotonic swelling that prestressed the cytoskeleton and the bilayer greatly increased the sensitivity of both control and cytoD cells so there are two pathways to transmit force to the channels. In contrast to patch, removing divalent ions decreased the whole-cell current. The difference between whole cell and patch properties provide new insights into our understanding of the Piezo1 gating mechanisms and cautions against generalization to in situ behavior. 相似文献
2.
Cardiac Na currents and the inactivating, reopening, and waiting properties of single cardiac Na channels 总被引:17,自引:6,他引:17
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Tetrodotoxin (TTX)-sensitive Na currents were examined in single dissociated ventricular myocytes from neonatal rats. Single channel and whole cell currents were measured using the patch-clamp method. The channel density was calculated as 2/micron 2, which agreed with our usual finding of four channels per membrane patch. At 20 degrees C, the single channel conductance was 20 pS. The open time distributions were fit by a single-exponential function with a mean open time of approximately 1.0 ms at membrane potentials from -60 to -40 mV. Averaged single channel and whole cell currents were similar when scaled and showed both fast and slow rates of inactivation. The inactivation and activation gating shifted quickly to hyperpolarized potentials for channels in cell-attached as well as excised patches, whereas a much slower shift occurred in whole cells. Slowly inactivating currents were present in both whole cell and single channel current measurements at potentials as positive as -40 mV. In whole cell measurements, the potential range could be extended, and slow inactivation was present at potentials as positive as -10 mV. The curves relating steady state activation and inactivation to membrane potential had very little overlap, and slow inactivation occurred at potentials that were positive to the overlap. Slow inactivation is in this way distinguishable from the overlap or window current, and the slowly inactivating current may contribute to the plateau of the rat cardiac action potential. On rare occasions, a second set of Na channels having a smaller unit conductance and briefer duration was observed. However, a separate set of threshold channels, as described by Gilly and Armstrong (1984. Nature [Lond.]. 309:448), was not found. For the commonly observed Na channels, the number of openings in some samples far exceeded the number of channels per patch and the latencies to first opening or waiting times were not sufficiently dispersed to account for the slowly inactivating currents: the slow inactivation was produced by channel reopening. A general model was developed to predict the number of openings in each sample. Models in which the number of openings per sample was due to a dispersion of waiting times combined with a rapid transition from an open to an absorbing inactivated state were unsatisfactory and a model that was more consistent with the results was identified. 相似文献
3.
Patch and whole cell calcium currents recorded simultaneously in snail neurons 总被引:10,自引:6,他引:4
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The flow of Ca ions through single Ca channels has been examined. The gigaseal method was used on identifiable snail neurons that were voltage clamped using a two-microelectrode voltage clamp method. Average Ca patch currents and whole cell currents have similar time courses. They are affected similarly by changes in temperature. The differences in amplitude and inactivation between Ba and Ca whole cell currents were present in the patch records. The stationary noise spectra recorded from ensembles of multichannel patches have two components with fast and slow time constants equivalent to two components in the whole cell tail current relaxations. Elementary current amplitudes measured from the variance-mean relationship and from noise spectra gave values comparable to measurements from single channels. The single channel I-V relationship was curvilinear and the maximum slope conductance in 40 mM Cao was 7 pS. The amplitude of unitary currents was unchanged at long times when inactivation had occurred; hence depletion is not involved in this process. Channel density was approximately 3 microns-2 and was the same for Ba and Ca currents. The whole cell asymmetry currents gave very large values for the gating charge per channel. Changes in temperature from 29 to 9 degrees C had only a slight effect on the two Ca tail current tau's at potentials where turn-on of patch and whole cell currents was markedly slowed and the peak amplitudes were reduced by one-third. Single channel recordings were obtained at these two temperatures, and the mean open time and the fast component of the closed times were scarcely affected. Unit amplitudes were reduced by 30% and the slow closed time component was doubled. Therefore, peak currents and the slow closed time component was doubled. Therefore, peak currents were reduced partly as a result of the reduction in unit amplitude, but mainly as a result of a reduction in opening probability, the latter arising from an increase of the long closed times. It is concluded that the behavior of single Ca channels in membrane patches is the same as it is in whole cells. Cooling from 29 to 9 degrees C acts primarily on transitions among closed states and has little effect on the open to closed transition. 相似文献
4.
Halide permeability sequences were obtained from reversal potential measurements of single-channel currents through 10 pS and 20 pS anion channels in human airway epithelial cells. The sequences obtained were Cl- greater than I- greater than Br- greater than or equal to F- for the 10 pS channel and Cl- greater than I- greater than or equal to Br- greater than or equal to F- for the 20 pS channel. However, the permeability differences were not large, the greatest being 0.66 for the ratio of fluoride to chloride permeability in the 20 pS channel. Single-channel currents were also measured with solutions of constant halide concentration but varying ratios of chloride to fluoride ions. An anomalous mole fraction effect was observed for the 20 pS channel but not for the 10 pS channel, suggesting that the former is a multi-ion channel. Comparison of the halide permeability sequences of these two channels with those of whole-cell currents in other epithelial cells does not support their involvement in any of the known whole-cell epithelial currents. 相似文献
5.
Calcium channel selectivity for divalent and monovalent cations. Voltage and concentration dependence of single channel current in ventricular heart cells 总被引:53,自引:22,他引:31
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Single channel and whole cell recordings were used to study ion permeation through Ca channels in isolated ventricular heart cells of guinea pigs. We evaluated the permeability to various divalent and monovalent cations in two ways, by measuring either unitary current amplitude or reversal potential (Erev). According to whole cell measurements of Erev, the relative permeability sequence is Ca2+ greater than Sr2+ greater than Ba2+ for divalent ions; Mg2+ is not measurably permeant. Monovalent ions follow the sequence Li+ greater than Na+ greater than K+ greater than Cs+, and are much less permeant than the divalents. These whole cell measurements were supported by single channel recordings, which showed clear outward currents through single Ca channels at strong depolarizations, similar values of Erev, and similar inflections in the current-voltage relation near Erev. Information from Erev measurements stands in contrast to estimates of open channel flux or single channel conductance, which give the sequence Na+ (85 pS) greater than Li+ (45 pS) greater than Ba2+ (20 pS) greater than Ca2+ (9 pS) near 0 mV with 110-150 mM charge carrier. Thus, ions with a higher permeability, judged by Erev, have lower ion transfer rates. In another comparison, whole cell Na currents through Ca channels are halved by less than 2 microM [Ca]o, but greater than 10 mM [Ca]o is required to produce half-maximal unitary Ca current. All of these observations seem consistent with a recent hypothesis for the mechanism of Ca channel permeation, which proposes that: ions pass through the pore in single file, interacting with multiple binding sites along the way; selectivity is largely determined by ion affinity to the binding sites rather than by exclusion by a selectivity filter; occupancy by only one Ca ion is sufficient to block the pore's high conductance for monovalent ions like Na+; rapid permeation by Ca ions depends upon double occupancy, which only becomes significant at millimolar [Ca]o, because of electrostatic repulsion or some other interaction between ions; and once double occupancy occurs, the ion-ion interaction helps promote a quick exit of Ca ions from the pore into the cell. 相似文献
6.
Piezo ion channels have been found to be essential for mechanical responses in cells. These channels were first shown to exist in Neuro2A cells, and the gene was identified by siRNAs that diminished the mechanical response. Piezo channels are approximately 2500 amino acids long, have between 24–32 transmembrane regions, and appear to assemble into tetramers and require no other proteins for activity. They have a reversal potential around 0 mV and show voltage dependent inactivation. The channel is constitutively active in liposomes, indicating that no cytoskeletal elements are required. Heterologous expression of the Piezo protein can create mechanical sensitivity in otherwise insensitive cells.
Piezo1 currents in outside-out patches were blocked by the extracellular MSC inhibitor peptide GsMTx4. Both enantiomeric forms of GsMTx4 inhibited channel activity in a manner similar to endogenous mechanical channels. Piezo1 can adopt a tonic (non-inactivating) form with repeated stimulation. The transition to the non-inactivating form generally occurs in large groups of channels, indicating that the channels exist in domains, and once the domain is compromised, the members simultaneously adopt new properties.
Piezo proteins are associated with physiological responses in cells, such as the reaction to noxious stimulus of Drosophila larvae. Recent work measuring cell crowding, shows that Piezo1 is essential for the removal of extra cells without apoptosis. Piezo1 mutations have also been linked to the pathological response of red blood cells in a genetic disease called Xerocytosis. These finding suggest that Piezo1 is a key player in cells’ responses to mechanical stimuli. 相似文献
7.
《Channels (Austin, Tex.)》2013,7(4):214-219
Piezo ion channels have been found to be essential for mechanical responses in cells. These channels were first shown to exist in Neuro2A cells, and the gene was identified by siRNAs that diminished the mechanical response. Piezo channels are approximately 2500 amino acids long, have between 24–32 transmembrane regions, and appear to assemble into tetramers and require no other proteins for activity. They have a reversal potential around 0 mV and show voltage dependent inactivation. The channel is constitutively active in liposomes, indicating that no cytoskeletal elements are required. Heterologous expression of the Piezo protein can create mechanical sensitivity in otherwise insensitive cells. Piezo1 currents in outside-out patches were blocked by the extracellular MSC inhibitor peptide GsMTx4. Both enantiomeric forms of GsMTx4 inhibited channel activity in a manner similar to endogenous mechanical channels. Piezo1 can adopt a tonic (non-inactivating) form with repeated stimulation. The transition to the non-inactivating form generally occurs in large groups of channels, indicating that the channels exist in domains, and once the domain is compromised, the members simultaneously adopt new properties. Piezo proteins are associated with physiological responses in cells, such as the reaction to noxious stimulus of Drosophila larvae. Recent work measuring cell crowding, shows that Piezo1 is essential for the removal of extra cells without apoptosis. Piezo1 mutations have also been linked to the pathological response of red blood cells in a genetic disease called Xerocytosis. These finding suggest that Piezo1 is a key player in cells’ responses to mechanical stimuli. 相似文献
8.
The effects of mm‐waves 60.22–62.22 GHz and 75 GHz on A‐type K+ currents and the effects of 61.22 GHz on Ca2+ currents of Lymnaea neurons were investigated using a whole‐cell voltage‐clamp technique. The open end of a rectangular waveguide covered with a thin Teflon film served as a radiator. Specific absorption rates at the waveguide outlet, inserted into physiological solution, were in the range of 0–2400 W/kg. Millimeter wave irradiation increased the peak amplitudes, activation rates, and inactivation rates of both ion currents. The changes in A‐type K+ current were not dependent on the irradiation frequency. It was shown that the changes in the amplitudes and kinetics of both currents resulted from the temperature rise produced by irradiation. No additional effects of irradiation on A‐type K+ current other than thermal were found when tested at the phase transition temperature or in the presence of ethanol. Ethanol reduced the thermal effect of irradiation. Millimeter waves had no effect on the steady‐state activation and inactivation curves, suggesting that the membrane surface charge and binding of calcium ions to the membrane in the area of channel locations did not change. Bioelectromagnetics 20:24–33, 1999. © 1999 Wiley‐Liss, Inc. 相似文献
9.
Wang Z Eldstrom JR Jantzi J Moore ED Fedida D 《American journal of physiology. Heart and circulatory physiology》2004,286(2):H749-H759
Voltage-dependent potassium channel trafficking and localization are regulated by proteins of the cytoskeleton, but the mechanisms by which these occur are still unclear. Using human embryonic kidney (HEK) cells as a heterologous expression system, we tested the role of the actin cytoskeleton in modulating the function of Kv4.2 channels. Pretreatment (>or=1 h) of HEK cells with 5 microM cytochalasin D to disrupt the actin microfilaments greatly augmented whole cell Kv4.2 currents at potentials positive to -20 mV. However, no changes in the voltage dependence of activation and inactivation of macroscopic currents were observed to account for this increase. Similarly, single channel recordings failed to reveal any significant changes in the single channel conductance, open probability, and kinetics. However, the mean patch current was increased from 0.9 +/- 0.2 pA in control to 6.7 +/- 3.0 pA in the presence of cytochalasin D. Imaging experiments revealed a clear increase in the surface expression of the channels and the appearance of "bright spot" features, suggesting that large numbers of channels were being grouped at specific sites. Our data provide clear evidence that increased numbers and altered distribution of Kv4.2 channels at the cell surface are primarily the result of reorganization of the actin cytoskeleton. 相似文献
10.
Outward K+ currents were recorded from 3-day-old embryonic chick ventricular myocytes using the patch clamp method. Two types of macroscopic outward currents were observed, one with rapid activation and de-activation time courses, and the other displaying a slower activation and long-duration tail currents. A time-dependent inactivation at positive potentials was a feature of the rapidly-activating current, allowing resolution of an early outward current. Single K+ channel currents were recorded using the outside-out patch technique. Two classes of K+ channels, which may contribute to the macroscopic currents, were differentiated on the basis of their conductances and kinetics. One class (ca 20 pS conductance) showed a rapid activation upon depolarization, and the other class (ca 60 pS) had a more delayed activation. A time-dependent inactivation of the rapid-activating, single-channel K+ current was also recorded. The two types of K+ channels contribute outward current during the plateau and promote the repolarization of the action potential, and the slowly de-activating K+ current may also be involved in the electrogenesis of automaticity observed in some of these cells. 相似文献
11.
Single channel currents have been recorded from cell-attached patches of tumoral adrenocortical cells. Our experiments suggest the existence of three sets of potassium channels in the surface membrane of these cells. All channel types can be recorded in a given membrane patch but some patches have only one type of single channel currents. One channel type has a unitary conductance of about 103 pS. The other two channels have smaller conductances and opposite voltage dependence. In one case channels open on depolarization and have a single channel conductance of 31.6 pS. In the other case the probability of being in the open state increases on hyperpolarization and the single channel conductance is of 21 pS. These channels seem to be similar to the delayed and anomalous rectifying potassium channels seen in other preparations. The role of membrane ionic permeability in steroid release induced by ACTH is discussed. 相似文献
12.
Ca2+ currents in cerebral artery smooth muscle cells of rat at physiological Ca2+ concentrations 总被引:1,自引:0,他引:1
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《The Journal of general physiology》1996,107(4):459-472
Single Ca2+ channel and whole cell currents were measured in smooth muscle cells dissociated from resistance-sized (100-microns diameter) rat cerebral arteries. We sought to quantify the magnitude of Ca2+ channel currents and activity under the putative physiological conditions of these cells: 2 mM [Ca2+]o, steady depolarizations to potentials between -50 and -20 mV, and (where possible) without extrinsic channel agonists. Single Ca2+ channel conductance was measured over a broad range of Ca2+ concentrations (0.5-80 mM). The saturating conductance ranged from 1.5 pS at 0.5 mM to 7.8 pS at 80 mM, with a value of 3.5 pS at 2 mM Ca (unitary currents of 0.18 pA at -40 mV). Both single channel and whole cell Ca2+ currents were measured during pulses and at steady holding potentials. Ca2+ channel open probability and the lower limit for the total number of channels per cell were estimated by dividing the whole-cell Ca2+ currents by the single channel current. We estimate that an average cell has at least 5,000 functional channels with open probabilities of 3.4 x 10(-4) and 2 x 10(-3) at -40 and -20 mV, respectively. An average of 1-10 (-40 mV and -20 mV, respectively) Ca2+ channels are thus open at physiological potentials, carrying approximately 0.5 pA steady Ca2+ current at -30 mV. We also observed a very slow reduction in open probability during steady test potentials when compared with peak pulse responses. This 4- 10-fold reduction in activity could not be accounted for by the channel's normal inactivation at our recording potentials between -50 and -20 mV, implying that an additional slow inactivation process may be important in regulating Ca2+ channel activity during steady depolarization. 相似文献
13.
Cells can respond to mechanical stress by gating mechanosensitive ion channels (MSCs). The cloning of Piezo1, a eukaryotic cation selective MSC, defines a new system for studying mechanical transduction at the cellular level. Because Piezo1 has electrophysiological properties similar to those of endogenous cationic MSCs that are selectively inhibited by the peptide GsMTx4, we tested whether the peptide targets Piezo1 activity. Extracellular GsMTx4 at micromolar concentrations reversibly inhibited ~80% of the mechanically induced current of outside-out patches from transfected HEK293 cells. The inhibition was voltage insensitive, and as seen with endogenous MSCs, the mirror image d enantiomer inhibited like the l. The rate constants for binding and unbinding based on Piezo1 current kinetics provided association and dissociation rates of 7.0 × 10(5) M(-1) s(-1) and 0.11 s(-1), respectively, and a K(D) of ~155 nM, similar to values previously reported for endogenous MSCs. Consistent with predicted gating modifier behavior, GsMTx4 produced an ~30 mmHg rightward shift in the pressure-gating curve and was active on closed channels. In contrast, streptomycin, a nonspecific inhibitor of cationic MSCs, showed the use-dependent inhibition characteristic of open channel block. The peptide did not block currents of the mechanical channel TREK-1 on outside-out patches. Whole-cell Piezo1 currents were also reversibly inhibited by GsMTx4, and although the off rate was nearly identical to that of outside-out patches, differences were observed for the on rate. The ability of GsMTx4 to target the mechanosensitivity of Piezo1 supports the use of this channel in high-throughput screens for pharmacological agents and diagnostic assays. 相似文献
14.
Two modes of gating during late Na+ channel currents in frog sartorius muscle 总被引:17,自引:6,他引:11
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Na+ currents were measured during 0.4-s depolarizing pulses using the cell-attached variation of the patch-clamp technique. Patches on Cs-dialyzed segments of sartorius muscle of Rana pipiens contained an estimated 25-500 Na+ channels. Three distinct types of current were observed after the pulse onset: a large initial surge of inward current that decayed within 10 ms (early currents), a steady "drizzle" of isolated, brief, inward unitary currents (background currents), and occasional "cloudbursts" of tens to hundreds of sequential unitary inward currents (bursts). Average late currents (background plus bursts) were 0.12% of peak early current amplitude at -20 mV. 85% of the late currents were carried by bursting channels. The unit current amplitude was the same for all three types of current, with a conductance of 10.5 pS and a reversal potential of +74 mV. The magnitudes of the three current components were correlated from patch to patch, and all were eliminated by slow inactivation. We conclude that all three components were due to Na+ channel activity. The mean open time of the background currents was approximately 0.25 ms, and the channels averaged 1.2 openings for each event. Neither the open time nor the number of openings of background currents was strongly sensitive to membrane potential. We estimated that background openings occurred at a rate of 0.25 Hz for each channel. Bursts occurred once each 2,000 pulses for each channel (assuming identical channels). The open time during bursts increased with depolarization to 1-2 ms at -20 mV, whereas the closed time decreased to less than 20 ms. The fractional open time during bursts was fitted with m infinity 3 using standard Na+ channel models. We conclude that background currents are caused by a return of normal Na+ channels from inactivation, while bursts are instances where the channel's inactivation gate spontaneously loses its function for prolonged periods. 相似文献
15.
When transiently expressed in tsA-201 cells, Ca(v)1.4 calcium channels support only modest whole-cell currents with unusually slow voltage-dependent inactivation kinetics. To examine the basis for this unique behavior we used cell-attached patch single-channel recordings using 100 mM external barium as the charge carrier to determine the single-channel properties of Ca(v)1.4 and to compare them to those of the Ca(v)1.2. Ca(v)1.4 channel openings occurred infrequently and were of brief duration. Moreover, openings occurred throughout the duration of the test depolarization, indicating that the slow inactivation kinetics observed at the whole-cell level are caused by sustained channel activity. Ca(v)1.4 and Ca(v)1.2 channels displayed similar latencies to first opening. Because of the rare occurrence of events, the probability of opening could not be precisely determined but was estimated to be <0.015 over a voltage range of -20 to +20 mV. The single-channel conductance of Ca(v)1.4 channels was approximately 4 pS compared with approximately 20 pS for Ca(v)1.2 under the same experimental conditions. Additionally, in the absence of divalent cations, Ca(v)1.4 channels pass cesium ions with a single-channel conductance of approximately 21 pS. Although Ca(v)1.2 opening events were best described kinetically with two open time constants, Ca(v)1.4 open times were best described by a single time constant. BayK8644 slightly enhanced the single-channel conductance in addition to increasing the open time constant for Ca(v)1.4 channels by approximately 45% without, however, causing the appearance of an additional slower gating mode. Overall, our data indicate that single Ca(v)1.4 channels support only minute amounts of calcium entry, suggesting that large numbers of these channels are needed to allow for significant whole-cell current activity, and providing a mechanism to reduce noise in the visual system. 相似文献
16.
F Bezanilla 《Biophysical journal》1987,52(6):1087-1090
Since the work of A. L. Hodgkin and A. F. Huxley (1952. J. Physiol. [Lond.].117:500-544) the squid giant axon has been considered the classical preparation for the study of voltage-dependent sodium and potassium channels. In this preparation much data have been gathered on macroscopic and gating currents but no single sodium channel data have been available. This paper reports patch clamp recording of single sodium channel events from the cut-open squid axon. It is shown that the single channel conductance in the absence of external divalent ions is approximately 14 pS, similar to sodium channels recorded from other preparations, and that their kinetic properties are consistent with previous results on gating and macroscopic currents obtained from the perfused squid axon preparation. 相似文献
17.
Duk-Su Koh Peter Jonas Michael E. Bräu Werner Vogel 《The Journal of membrane biology》1992,130(2):149-162
A novel potassium-selective channel which is active at membrane potentials between -100 mV and +40 mV has been identified in peripheral myelinated axons of Xenopus laevis using the patch-clamp technique. At negative potentials with 105 mM-K on both sides of the membrane, the channel at 1 kHz resolution showed a series of brief openings and closings interrupted by longer closings, resulting in a flickery bursting activity. Measurements with resolution up to 10 kHz revealed a single-channel conductance of 49 pS with 105 mM-K and 17 pS with 2.5 mM-K on the outer side of the membrane. The channel was selective for K ions over Na ions (PNa/PK = 0.033). The probability of being within a burst in outside-out patches varied from patch to patch (> 0.2, but often > 0.9), and was independent of membrane potential. Open-time histograms were satisfactorily described with a single exponential (tau o = 0.09 msec), closed times with the sum of three exponentials (tau c = 0.13, 5.9, and 36.6 msec). Sensitivity to external tetraethylammonium was comparatively low (IC50 = 19.0 mM). External Cs ions reduced the apparent unitary conductance for inward currents at Em = -90 mV (IC50 = 1.1 mM). Ba and, more potently, Zn ions lowered not only the apparent single-channel conductance but also open probability. The local anesthetic bupivacaine with high potency reduced probability of being within a burst (IC50 = 165 nM). The flickering K channel is clearly different from the other five types of K channels identified so far in the same preparation. We suggest that this channel may form the molecular basis of the resting potential in vertebrate myelinated axons. 相似文献
18.
Summary Using whole-cell patch-clamp techniques, we demonstrate that sheep parotid secretory cells have both inwardly and outwardly rectifying currents. The outwardly rectifying current, which is blocked by 10 mmol/liter tetraethylammonium (TEA) applied extracellularly, is probably carried by the 250 pS Ca2+-and voltage-activated K+ (BK) channel which has been described in previous studies. In contrast, the inwardly rectifying current, which is also carried by K+ ions, is not sensitive to TEA. It is similar to the inwardly rectifying currents observed in many excitable tissues in that (i) its conductance is dependent on the square root of the extracellular K+, (ii) the voltage range over which it is activated is influenced by the extracellular K+ concentration and (iii) it is blocked by the addition of Cs+ ions (670 µmol/liter) to the bathing solution. Our previously published cell-attached patch studies have shown that the channel type most commonly observed in the basolateral membrane of unstimulated sheep parotid secretory cells is a K+ channel with a conductance of 30 pS and, in this study, we find that its conductance also depends on the square root of the extracellular K+ concentration. It thus seems likely that it carries the inwardly rectifying K+ current seen in the whole-cell studies. 相似文献
19.
20.
The Ach-evoked Ca2+-activated K+ Current in Mouse Mandibular Secretory Cells. Single Channel Studies
Although acetylcholine (ACh) is able to activate voltage- and Ca2+-sensitive K+ (BK) channels in mouse mandibular secretory cells, our recent whole cell studies have suggested that these channels, like
those in sheep parotid secretory cells, do not contribute appreciably to the conductance that carries the ACh-evoked whole
cell K+ current. In the present study, we have used cell-attached patch clamp methods to identify and characterize the K+ channel type responsible for carrying the bulk of this current. When the cells were bathed in a NaCl-rich solution the predominant
channel type activated by ACh (1 μmol/l or 50 nmol/l) had a conductance only of 40 pS; it was not blocked by TEA but it was
sensitive to quinine and it conducted Rb+ to an appreciable extent. BK channels, which could be seen in some but not all patches from resting cells, also showed increased
activity when ACh was added to the bath, but they were much less conspicuous during ACh stimulation than the 40-pS channels.
When the cells were bathed in a KCl-rich rather than a NaCl-rich solution, a small-conductance K+ channel, sensitive to quinine but not to TEA, was still the most conspicuous channel to be activated by ACh although its
conductance was reduced to 25 pS. Our studies confirm that the ACh-evoked whole-cell K+ current is not carried substantially by BK channels and show that it is carried by a small-conductance K+ channel with quite different properties.
Received: 28 September 1995/Revised: 26 December 1995 相似文献