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  • 1
    Electronic Resource
    Electronic Resource
    Cation specificity and pharmacological properties of the Ca2+-dependent K+ channel of rat cortical collecting ducts (1993)
    Springer
    Pflügers Archiv 422 (1993), S. 481-491 
    Details
    ISSN: 1432-2013
    Keywords: Patch clamp ; Verapamil ; Charyb-dotoxin ; Apamin ; K+ channel blocker ; Permselectivity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The luminal membrane of principal cells of rat cortical collecting duct (CCD) is dominated by a K+ conductance. Two different K+ channels are described for this membrane. K+ secretion probably occurs via a small-conductance Ca2+-independent channel. The function of the second, large-conductance Ca2+-dependent channel is unclear. This study examines properties of this channel to allow a comparison of this K+ channel with the macroscopic K+ conductance of the CCD and with similar K+ channels from other preparations. The channel is poorly active on the cell. It has a conductance of 263±11 pS (n=36, symmetrical K+ concentrations) and of 139±3 pS (n=91) with 145 mmol/l K+ on one side and 3.6 mmol/l K+ on the other side of the membrane. Its open probability is high after excision (0.71±0.03, n=85). The channel flickers rapidly between open and closed states. Its permeability in the cell-free configuration was 7.0±0.2×10−13 cm3/s (n=85). It is inhibited by several typical blockers of K+ channels such as Ba2+, tetraethylammonium, quinine, and quinidine and high concentrations of Mg2+. The Ca2+ antagonists verapamil and diltiazem also inhibit this K+ channel. As is typical for the maxi K+ channel, it is inhibited by charybdotoxin but not by apamin. The selectivity of this large-conductance K+ channel demonstrates significant differences between the permeability sequence (P K 〉 P Rb 〉 P NH4 〉 P Cs=P Li=P Na=P choline=0) and the conductance sequence (g K 〉 g NH4 〉 g Rb 〉 g Li=g choline 〉 g Cs=g Na=0). The only other cations that are significantly conducted by this channel besides K+ (g K at V c =∞ is 279±8 pS, n=88) are NH 4 + (g NH4=127±22 pS, n=10) and Rb+ (g Rb=36±5 pS, n=6). The K+ currents through this channel are reduced by high concentrations of choline+, Cs+, Rb+, and NH 4 + . These properties and the dependence of this channel on Ca2+ and voltage classify it as a “maxi” K+ channel. A possible physiological function of this channel is discussed in the accompanying paper.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00375076
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  • 2
    Electronic Resource
    Electronic Resource
    Regulation and possible physiological role of the Ca2-dependent K+ channel of cortical collecting ducts of the rat (1993)
    Springer
    Pflügers Archiv 422 (1993), S. 492-498 
    Details
    ISSN: 1432-2013
    Keywords: ATP ; pH ; Voltage dependence ; Volume regulation ; Intracellular Ca2+ ; Patch clamp ; Fura-2
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract In the luminal membrane of rat cortical collecting duct (CCD) a big Ca2+-dependent and a small Ca2+-independent K+ channel have been described. Whereas the latter most likely is responsible for the K+ secretion in this nephron segment, the function of the large-conductance K+ channel is unknown. The regulation of this channel and its possible physiological role were examined with the conventional cell-free and the cell-attached nystatin patch-clamp techniques. Patch-clamp recordings were obtained from the luminal membrane of isolated perfused CCD segments and from freshly isolated CCD cells. Intracellular calcium was measured using the calcium-sensitive dye fura-2. The large-conductance K+ channel was strongly voltage- and calcium-dependent. At 3 μmol/l cytosolic Ca2+ activity it was half-maximally activated. At 1 mmol/l it was neither regulated by cytosolic pH nor by ATP. At 1 μmol/l Ca2+ activity the open probability (P o) of this channel was pH-dependent. At pH 7.0 P o was decreased to 4±2% (n=9) and at pH 8.5 it was increased to 425±52% (n=9) of the control. At this low Ca2+ activity the P o of the channel was reduced by 1 mmol/l ATP to 8±4% (n=6). Cell swelling activated the large-conductance K+ channel (n=14) and hyperpolarized the membrane potential of the cells by 9±1 mV (n=23). Intracellular Ca2+ activity increased after hypotonic stress. This increase depended on the extracellular Ca2+ activity. A possible physiological function of the large-conductance K+ channel in rat CCD cells may be the reduction of the intracellular K+ concentration after cell swelling. Once this channel is activated by increases in the cytosolic Ca2+ activity it can be regulated by changes in cellular pH and ATP.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00375077
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  • 3
    Electronic Resource
    Electronic Resource
    Analysing ion channels with hidden Markov models (1994)
    Springer
    Pflügers Archiv 426 (1994), S. 328-332 
    Details
    ISSN: 1432-2013
    Keywords: Cortical collecting duct ; K+ channel ; Rat ; Isolated tubule ; Patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Ion channel current amplitudes (μ) and open probabilities (P o) have been analysed so far by defining a 50% threshold to distinguish between open and closed states of the channels. With this standard method (SM) it is very difficult or even impossible to analyse channels of different size in one membrane patch correctly. A stochastical model, named the hidden Markov model (HMM), separates between observation noise and the stochastic process of opening and closing of ion channels. The HMM allows the independent analysis of μ, P o, and mean dwell times (τ) of different channels in one membrane patch, without defining threshold levels. Using this method errors in the analysis are not summarized like in the SM because all different analysing procedures (e. g. filtering, setting of threshold, fitting processes) are done in one step. Two different K+ channels in excised basolateral membranes of the cortical collecting duct of rat (CCD) were analysed by the SM and the HMM. The μ value of the intermediate-conductance K+ channel (i-K+) was 3.9±0.1 pA (SM) and 3.8±0.2 pA (HMM) for 11 observations. The P o value of this channel was 10.2±4.2% (SM) and 10.1±4.0% (HMM). The mean τ values were 5.4±0.6 ms for the open state and 9.6±2.2 ms and 145±21 ms for the closed states (SM) and 7.8±1.1 ms, 7.7±0.9 ms and 148±24 ms (HMM), respectively. For seven small-conductance K+ (s-K+) channels, which were found in the same membrane patches as the i-K+, an accurate analysis of P o and τ was not possible with the SM. The μ value was 1.0±0.1 (SM), 0.9±0.1 (HMM) pA. P o was 16.6±4.6%, the open τ value was 11.1±2.8 ms, and the closed τ value was 34.9±8.5 ms. The HMM allows the analysis of single-channel currents, P o, and mean τ values when different or more than one ion channel(s) are colocalized in one membrane patch. Where analysis with the SM was possible results did not significantly differ from those obtained with the HMM. Thus for this kind of analysis the method of setting a 50% threshold appears justified.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00374789
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  • 4
    Electronic Resource
    Electronic Resource
    K+ channels in the basolateral membrane of rat cortical collecting duct (1993)
    Springer
    Pflügers Archiv 424 (1993), S. 470-477 
    Details
    ISSN: 1432-2013
    Keywords: Patch clamp ; Verapamil ; Charybdotoxin ; K+ channel blocker ; Ca2+ dependence ; pH dependence ; Cell-attached nystatin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Impalement studies in isolated perfused cortical collecting ducts (CCD) of rats have shown that the basolateral membrane possesses a K+ conductive pathway. In the present study this pathway was investigated at the single-channel level using the patch-clamp technique. Patch-clamp recordings were obtained from enzymatically isolated CCD segments and freshly isolated CCD cells with the conventional cell-free, cell-attached and the cell-attached nystatin method. Two K+ channels were found which were highly active on the cell with a conductance of 67±5 pS (n=18) and 148±4 pS (n=21) with 145 mmol/l K+ in the pipette. In excised patches the first channel had a conductance of 28±2 pS (n=15), whereas the second one had a conductance of 85±1 pS (n=53) at 0 mV clamp voltage with 145 mmol/l K+ on one side and 3.6 mmol/l K+ on the other side of the membrane. So far it has not been possible to characterize the smaller channel further. Excised, and with symmetrical K+ concentrations of 145 mmol/l, the intermediate channel had a linear conductance of 198±19 pS (n=5). After excision in the inside-out configuration the open probability (P o) of this channel was low (0.18±0.05, n=13) whereas in the outside-out configuration this channel had a threefold higher P o (0.57±0.04, n=12). Several inhibitors were tested in excised membranes. Ba2+ (1 mmol/l), tetraethylammonium (TEA+, 10 mmol/l) and verapamil (0.1 mmol/l) all blocked this channel reversibly. Furthermore P o was reversibly reduced by 10 nmol/l charybdotoxin (outside-out). This K+ channel of the basolateral membrane was regulated by cellular pH. P o was reduced to 26±3% at pH 6.5 (n=6) and increased to 216±18% at pH 8.5 (n=7) compared to pH 7.4. Half-maximal inhibition was reached at pH 7.0. The channel had its highest P o at a Ca2+ activity of less than 10−8 mol/l (n=13). Increasing the Ca2+ activity to 1 mmol/l on the cytosolic side of the membrane resulted in a reduction of P o to 13±3% (n=11). Half-maximal inhibition was reached at a Ca2+ activity of 10−5 mol/l. The high activity of both K+ channels of the basolateral membrane on the cell indicates that they may serve for K+ recirculation across the basolateral membrane.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00374910
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