Characterization of a Hyperpolarization-Activated Inward Current in Cajal-Retzius Cells in Rat Neonatal Neocortex

2000 ◽  
Vol 84 (3) ◽  
pp. 1681-1691 ◽  
Author(s):  
Werner Kilb ◽  
Heiko J. Luhmann
Keyword(s):  
Dissociation Constant ◽  
Reversal Potential ◽  
Hill Coefficient ◽  
Resting Membrane Potential ◽  
Patch Clamp Technique ◽  
Apparent Dissociation Constant ◽  
Membrane Hyperpolarization ◽  
Whole Cell ◽  
Inward Current ◽  
Retzius Cells

Cajal-Retzius cells are among the first neurons appearing during corticogenesis and play an important role in the establishment of cortical lamination. To characterize the hyperpolarization-activated inward current ( I h) and to investigate whether I h contributes to the relatively positive resting membrane potential (RMP) of these cells, we analyzed the properties of I h in visually identified Cajal-Retzius cells in cortical slices from neonatal rats using the whole cell patch-clamp technique. Membrane hyperpolarization to −90 mV activated a prominent inward current that was inhibited by 1 mM Cs+ and was insensitive to 1 mM Ba2+. The activation time constant for I h was strongly voltage dependent. In Na+-free solution, I h was reduced, indicating a contribution of Na+. An analysis of the tail currents revealed a reversal potential of −45.2 mV, corresponding to a permeability coefficient (pNa+/pK+) of 0.13. While an increase in the extracellular K+ concentration ([K+]e) enhances I h, it was reduced by a [K+]e decrease. This [K+]e dependence could not be explained by an effect on the electromotive force on K+ but suggested an additional extracellular binding site for K+ with an apparent dissociation constant of 7.2 mM. Complete Cl−substitution by Br−, I−, or NO3 − had no significant effect on I h, whereas a complete Cl−substitution by the organic compounds methylsulfate, isethionate, or gluconate reduced I h by ∼40%. The I h reduction observed in gluconate could be abolished by the addition of Cl−. The analysis of the [Cl−]e dependence of I h revealed a dissociation constant of 9.8 mM and a Hill-coefficient of 2.5, while the assumption of a gluconate-dependent I h reduction required an unreasonably high Hill-coefficient >20. An internal perfusion with the lidocaine derivative lidocaine N-ethyl bromide blocks I h within 1 min after establishment of the whole cell configuration. An inhibition of I h by 1 mM Cs+ was without an effect on RMP, action potential amplitude, threshold, width, or afterhyperpolarization. We conclude from these results that Cajal-Retzius cells express a prominent I hwith characteristic properties that does not contribute to the RMP.

P2X receptors in mouse Leydig cells

2006 ◽  
Vol 290 (4) ◽  
pp. C1009-C1017 ◽  
Author(s):  
Luiz Artur Poletto Chaves ◽  
Endrigo Piva Pontelli ◽  
Wamberto Antonio Varanda
Keyword(s):  
Leydig Cells ◽  
Pyridoxal Phosphate ◽  
Hill Coefficient ◽  
Disulfonic Acid ◽  
Inward Rectification ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Apparent Dissociation Constant ◽  
Whole Cell

ATP-activated currents were studied in Leydig cells of mice with the patch-clamp technique. Whole cell currents were rapidly activating and slowly desensitizing (55% decrement from the peak value on exposure to 100 μM ATP for 60 s), requiring 3 min of washout to recover 100% of the response. The concentration-response relationships for ATP, adenosine 5′- O-(3-thiotriphosphate) (ATPγS), and 2-methylthio-ATP (2-MeS-ATP) were described by the Hill equation with a concentration evoking 50% of maximal ATP response ( Kd) of 44, 110, and 637 μM, respectively, and a Hill coefficient of 2. The order of efficacy of agonists was ATP ≥ ATPγS > 2-MeS-ATP > 2′,3′- O-(4-benzoylbenzoyl)-ATP (BzATP). αβ-Methylene-ATP (αβ-MeATP), GTP, UTP, cAMP, and adenosine were ineffective. Suramin and pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS) blocked the responses in a concentration-dependent manner. The ATP-activated currents were dependent on extracellular pH, being maximal at pH 6.5 and decreasing with both acidification and alkalinization (apparent dissociation constant (p Ka) of 5.9 and 7.4, respectively). The whole cell current-voltage relationship showed inward rectification and reversed near 0 mV. Experiments performed in bi-ionic conditions for measurement of reversal potentials showed that this channel is highly permeable to calcium [permeability ( P)Ca/ PNa = 5.32], but not to chloride ( PCl/ PNa = 0.03) or N-methyl-d-glucamine (NMDG) ( PNMDG/ PNa = 0.09). Unitary currents recorded in outside-out patches had a chord conductance of 27 pS (between −90 and −50 mV) and were inward rectifying. The average current passing through the excised patch decreased with time [time constant (τ) = 13 s], resembling desensitization of the macroscopic current. These findings indicate that the ATP receptor present in Leydig cells shows properties most similar to those of cloned homomeric P2X2.

Inwardly rectifying K+ channels in esophageal smooth muscle

2000 ◽  
Vol 279 (5) ◽  
pp. G951-G960 ◽  
Author(s):  
Junzhi Ji ◽  
Anne Marie F. Salapatek ◽  
Nicholas E. Diamant
Keyword(s):  
Membrane Potential ◽  
Longitudinal Muscle ◽  
Muscle Cells ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Patch Clamp Technique ◽  
Membrane Hyperpolarization ◽  
Inward Current ◽  
Inwardly Rectifying

The whole cell patch-clamp technique was used to investigate whether there were inwardly rectifying K+(Kir) channels in the longitudinal muscle of cat esophagus. Inward currents were observable on membrane hyperpolarization negative to the K+ equilibrium potential ( E k) in freshly isolated esophageal longitudinal muscle cells. The current-voltage relationship exhibited strong inward rectification with a reversal potential ( E rev) of −76.5 mV. Elevation of external K+ increased the inward current amplitude and positively shifted its E rev after the E k, suggesting that potassium ions carry this current. External Ba2+ and Cs+ inhibited this inward current, with hyperpolarization remarkably increasing the inhibition. The IC50 for Ba2+ and Cs+ at −60 mV was 2.9 and 1.6 mM, respectively. Furthermore, external Ba2+ of 10 μM moderately depolarized the resting membrane potential of the longitudinal muscle cells by 6.3 mV while inhibiting the inward rectification. We conclude that Kir channels are present in the longitudinal muscle of cat esophagus, where they contribute to its resting membrane potential.

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.

Whole cell current analyses of pancreatic acinar AR42J cells. I. Voltage- and Ca(2+)-activated currents

1991 ◽  
Vol 260 (5) ◽  
pp. C934-C948 ◽  
Author(s):  
K. Kusano ◽  
H. Gainer
Keyword(s):  
Reversal Potential ◽  
Outward Current ◽  
Pancreatic Acinar Cells ◽  
Equilibrium Potential ◽  
Channel Blockers ◽  
Whole Cell ◽  
Inward Current ◽  
Tail Current ◽  
Ar42j Cells ◽  
Conductance Increase

Voltage- and Ca(2+)-activated whole cell currents were studied in AR42J cells, a clonal cell line derived from rat pancreatic acinar cells, using a patch electrode voltage-clamp technique. Four kinds of ionic currents were identified by their ionic dependencies, pharmacological properties, and kinetic parameters: 1) an outward current flow due mainly to a voltage-dependent K(+)-conductance increase, 2) an initial transient inward current due to an Na(+)-conductance increase, 3) transient and long-duration inward current due to a Ca(2+)-conductance increase, and 4) a slowly activating inward current that persists over the duration of the depolarizing pulse and deactivates slowly upon repolarization, producing a slow inward tail current. The slow inward tail current was particularly robust and was interpreted as due to a Ca(2+)-activated Cl(-)-conductance increase, since 1) the generation of this current was blocked by removing the extracellular Ca2+, applying Ca(2+)-channel blockers (Cd2+, nifedipine), or by lowering the intracellular Ca2+ concentration [( Ca2+]i) with EGTA; and 2) the reversal potential (Erev) of the slow inward tail current was close to 0 mV in the control condition (152 mM [Cl-]o/154 mM [Cl-]i), and changes of the [Cl-]o/[Cl )i ratio shifted the Erev toward the predicted Cl- equilibrium potential.

Postnatal Development of Electrophysiological Properties of Nucleus Accumbens Neurons

2000 ◽  
Vol 84 (5) ◽  
pp. 2204-2216 ◽  
Author(s):  
Marc L. Belleau ◽  
Richard A. Warren
Keyword(s):  
Nucleus Accumbens ◽  
Postnatal Development ◽  
Action Potentials ◽  
Membrane Resistance ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Projection Neurons ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Wide Range

We have studied the postnatal development of the physiological characteristics of nucleus accumbens (nAcb) neurons in slices from postnatal day 1 ( P1) to P49 rats using the whole cell patch-clamp technique. The majority of neurons (102/108) were physiologically identified as medium spiny (MS) projection neurons, and only these were subjected to detailed analysis. The remaining neurons displayed characteristics suggesting that they were not MS neurons. Around the time of birth and during the first postnatal weeks, the membrane and firing characteristics of MS neurons were quite different from those observed later. These characteristics changed rapidly during the first 3 postnatal weeks, at which point they began to resemble those found in adults. Both whole cell membrane resistance and membrane time constant decreased more than fourfold during the period studied. The resting membrane potential (RMP) also changed significantly from an average of −50 mV around birth to less than −80 mV by the end of the third postnatal week. During the first postnatal week, the current-voltage relationship of all encountered MS neurons was linear over a wide range of membrane potentials above and below RMP. Through the second postnatal week, the proportion of neurons displaying inward rectification in the hyperpolarized range increased steadily and after P15, all recorded MS neurons displayed significant inward rectification. At all ages, inward rectification was blocked by extracellular cesium and tetra-ethyl ammonium and was not changed by 4-aminopyridine; this shows that inward rectification was mediated by the same currents in young and mature MS neurons. MS neurons fired single and repetitive Na+/K+ action potentials as early as P1. Spike threshold and amplitude remained constant throughout development in contrast to spike duration, which decreased significantly over the same period. Depolarizing current pulses from rest showed that immature MS neurons fired action potentials more easily than their older counterparts. Taken together, the results from the present study suggest that young and adult nAcb MS neurons integrate excitatory synaptic inputs differently because of differences in their membrane and firing properties. These findings provide important insights into signal processing within nAcb during this critical period of development.

ATP-induced current in isolated outer hair cells of guinea pig cochlea

1990 ◽  
Vol 63 (5) ◽  
pp. 1068-1074 ◽  
Author(s):  
T. Nakagawa ◽  
N. Akaike ◽  
T. Kimitsuki ◽  
S. Komune ◽  
T. Arima
Keyword(s):  
Guinea Pig ◽  
Hair Cells ◽  
Reversal Potential ◽  
Outer Hair Cells ◽  
Hill Coefficient ◽  
Induced Response ◽  
Induced Current ◽  
Patch Clamp Technique ◽  
Positive Direction ◽  
Guinea Pig Cochlea

1. Electrical and pharmacologic properties of ATP-induced current in outer hair cells isolated from guinea pig cochlea were investigated in the whole-cell recording mode by the use of a conventional patch-clamp technique. 2. Under current-clamp conditions, rapid application of ATP depolarized the outer hair cells resulting in an increase in conductance. The ATP-induced response did not show any desensitization during a continuous application. 3. At a holding potential of -70 mV, the ATP-induced inward current increased in a sigmoidal fashion over the concentration range between 3 microM and 1 mM. The half-maximum concentration (EC50) was 12 microM and the Hill coefficient was 0.93. 4. The ATP-induced current had a reversal potential near 6 mV, which was close to the theoretical value (1 mV) calculated from the Goldman-Hodgkin-Katz equation for permeable intra- and extracellular cations. 5. In the current-voltage (I-V) relationship for the ATP response, a slight inward-going rectification was observed at more positive potentials than the reversal potential. 6. The substitution of extracellular Na+ by equimolar choline+ shifted the reversal potential of the ATP-induced current to more negative values. The substitution of Cs+ in the internal solution by N-methyl-D-glucamine+ (NMG+) shifted it in the positive direction. The reversal potential of ATP-induced current was also shifted to positive values with increasing extracellular Ca2+ concentration. A decrease of intracellular Cl- by gluconate- did not affect the reversal potential, thereby indicating that the ATP-induced current is carried through a large cation channel.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Evidence That the Product of the Human X-Linked Cgd Gene, Gp91-phox, Is a Voltage-Gated H+ Pathway

1999 ◽  
Vol 114 (6) ◽  
pp. 771-786 ◽  
Author(s):  
Lydia M. Henderson ◽  
Robert W. Meech
Keyword(s):  
Cell Membrane ◽  
Time Course ◽  
Lucifer Yellow ◽  
Chinese Hamster ◽  
Reversal Potential ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Whole Cell ◽  
Histidine Residues ◽  
Voltage Gated

Expression of gp91-phox in Chinese hamster ovary (CHO91) cells is correlated with the presence of a voltage-gated H+ conductance. As one component of NADPH oxidase in neutrophils, gp91-phox is responsible for catalyzing the production of superoxide (O2·2). Suspensions of CHO91 cells exhibit arachidonate-activatable H+ fluxes (Henderson, L.M., G. Banting, and J.B. Chappell. 1995. J. Biol. Chem. 270:5909–5916) and we now characterize the electrical properties of the pathway. Voltage-gated currents were recorded from CHO91 cells using the whole-cell configuration of the patch-clamp technique under conditions designed to exclude a contribution from ions other than H+. As in other voltage-gated proton currents (Byerly, L., R. Meech, and W. Moody. 1984. J. Physiol. 351:199–216; DeCoursey, T.E., and V.V. Cherny. 1993. Biophys. J. 65:1590–1598), a lowered external pH (pHo) shifted activation to more positive voltages and caused the tail current reversal potential to shift in the manner predicted by the Nernst equation. The outward currents were also reversibly inhibited by 200 μM zinc. Voltage-gated currents were not present immediately upon perforating the cell membrane, but showed a progressive increase over the first 10–20 min of the recording period. This time course was consistent with a gradual shift in activation to more negative potentials as the pipette solution, pH 6.5, equilibrated with the cell contents (reported by Lucifer yellow included in the patch pipette). Use of the pH-sensitive dye 2′7′ bis-(2-carboxyethyl)-5(and 6) carboxyfluorescein (BCECF) suggested that the final intracellular pH (pHi) was ∼6.9, as though pHi was largely determined by endogenous cellular regulation. Arachidonate (20 μM) increased the amplitude of the currents by shifting activation to more negative voltages and by increasing the maximally available conductance. Changes in external Cl− concentration had no effect on either the time scale or the appearance of the currents. Examination of whole cell currents from cells expressing mutated versions of gp91-phox suggest that: (a) voltage as well as arachidonate sensitivity was retained by cells with only the NH2-terminal 230 amino acids, (b) histidine residues at positions 111, 115, and 119 on a putative membrane-spanning helical region of the protein contribute to H+ permeation, (c) histidine residues at positions 111 and 119 may contribute to voltage gating, (d) the histidine residue at position 115 is functionally important for H+ selectivity. Mechanisms of H+ permeation through gp91-phox include the possible protonation/deprotonation of His-115 as it is exposed alternatively to the interior and exterior faces of the cell membrane (see Starace, D.M., E. Stefani, and F. Bezanilla. 1997. Neuron. 19:1319–1327) and the transfer of protons across an “H-X-X-X-H-X-X-X-H” motif lining a conducting pore.

scholarly journals Characterization of the calcium-activated chloride channel in isolated guinea-pig hepatocytes.

1994 ◽  
Vol 104 (2) ◽  
pp. 357-373 ◽  
Author(s):  
S Koumi ◽  
R Sato ◽  
T Aramaki
Keyword(s):  
Guinea Pig ◽  
Single Channel ◽  
Hill Coefficient ◽  
Disulfonic Acid ◽  
Dependent Manner ◽  
Channel Activity ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Cl Channels ◽  
Inside Out

Macroscopic and unitary currents through Ca(2+)-activated Cl- channels were examined in enzymatically isolated guinea-pig hepatocytes using whole-cell, excised outside-out and inside-out configurations of the patch-clamp technique. When K+ conductances were blocked and the intracellular Ca2+ concentration ([Ca2+]i) was set at 1 microM (pCa = 6), membrane currents were observed under whole-cell voltage-clamp conditions. The reversal potential of the current shifted by approximately 60 mV per 10-fold change in the external Cl- concentration. In addition, the current did not appear when Cl- was omitted from the internal and external solutions, indicating that the current was Cl- selective. The current was activated by increasing [Ca2+]i and was inactivated in Ca(2+)-free, 5 mM EGTA internal solution (pCa > 9). The current was inhibited by bath application of 9-anthracenecarboxylic acid (9-AC) and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) in a voltage-dependent manner. In single channel recordings from outside-out patches, unitary current activity was observed, whose averaged slope conductance was 7.4 +/- 0.5 pS (n = 18). The single channel activity responded to extracellular Cl- changes as expected for a Cl- channel current. The open time distribution was best described by a single exponential function with mean open lifetime of 97.6 +/- 10.4 ms (n = 11), while at least two exponentials were required to fit the closed time distributions with a time constant for the fast component of 21.5 +/- 2.8 ms (n = 11) and that for the slow component of 411.9 +/- 52.0 ms (n = 11). In excised inside-out patch recordings, channel open probability was sensitive to [Ca2+]i. The relationship between [Ca2+]i and channel activity was fitted by the Hill equation with a Hill coefficient of 3.4 and the half-maximal activation was 0.48 microM. These results suggest that guinea-pig hepatocytes possess Ca(2+)-activated Cl- channels.

scholarly journals Mobilization of intracellular calcium by intracellular flash photolysis of caged dihydrosphingosine in cultured neonatal rat sensory neurones.

1998 ◽  
Vol 45 (2) ◽  
pp. 311-326 ◽  
Author(s):  
A Ayar ◽  
N M Thatcher ◽  
U Zehavi ◽  
D R Trentham ◽  
R H Scott
Keyword(s):  
Flash Photolysis ◽  
Reversal Potential ◽  
Neonatal Rat ◽  
Xenon Lamp ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Sensory Neurones ◽  
Whole Cell ◽  
Inward Currents ◽  
Current Activation

The ability of dihydrosphingosine to release Ca2+ from intracellular stores in neurones was investigated by combining the whole cell patch clamp technique with intracellular flash photolysis of caged, N-(2-nitrobenzyl)dihydrosphingosine. The caged dihydrosphingosine (100 microM) was applied to the intracellular environment via the CsCl-based patch pipette solution which also contained 0.3% dimethylformamide and 2 mM dithiothreitol. Cultured dorsal root ganglion neurones from neonatal rats were voltage clamped at -90 mV and inward whole cell Ca2+-activated currents were recorded in response to intracellular photorelease of dihydrosphingosine. Intracellular photorelease of dihydrosphingosine (about 5 microM) was achieved using a Xenon flash lamp. Inward Ca2+-activated currents were evoked in 50 out of 57 neurones, the mean delay to current activation following photolysis was 82+/-13 s. The responses were variable with neurones showing transient, oscillating or sustained inward currents. High voltage-activated Ca2+ currents evoked by 100 ms voltage step commands to 0 mV were not attenuated by photorelease of dihydrosphingosine. Controls showed that alone a flash from the Xenon lamp did not activate currents, and that the unphotolysed caged dihydrosphingosine, and intracellular photolysis of 2-(2-nitrobenzylamino) propanediol also did not evoke responses. The dihydrosphingosine current had a reversal potential of -11+/-3 mV (n = 11), and was carried by two distinct Cl- and cation currents which were reduced by 85% and about 20% following replacement of monovalent cations with N-methyl-D-glucamine or application of the Cl- channel blocker niflumic acid (10 microM) respectively. The responses to photoreleased dihydrosphingosine were inhibited by intracellular application of 20 mM EGTA, 10 microM ryanodine or extracellular application of 10 microM dantrolene, but persisted when Ca2+ free saline was applied to the extracellular environment. Intracellular application of uncaged dihydrosphingosine evoked responses which were attenuated by photolysis of the caged Ca2+ chelator Diazo-2. Experiments also suggested that extracellular application of dihydrosphingosine can activate membrane conductances. We conclude that dihydrosphingosine directly or indirectly mobilises Ca2+ from ryanodine-sensitive intracellular stores in cultured sensory neurones.

Carbachol induces oscillations in membrane potential and intracellular calcium in a colonic tumor cell line, HT-29

1997 ◽  
Vol 273 (4) ◽  
pp. C1186-C1193 ◽  
Author(s):  
P. Sand ◽  
T. Svenberg ◽  
B. Rydqvist
Keyword(s):  
Membrane Potential ◽  
Reversal Potential ◽  
Tumor Cell Line ◽  
Equilibrium Potential ◽  
Patch Clamp Technique ◽  
Control Solution ◽  
Whole Cell ◽  
Calcium Dependent ◽  
High K ◽  
Ht 29

The patch-clamp technique was used to study the effects of carbachol (CCh) on HT-29 cells. During CCh exposure, the cells ( n = 23) depolarized close to the equilibrium potential for Cl−([Formula: see text]; −48 mV) and the membrane potential then started to oscillate (16/23 cells). In voltage-clamp experiments, similar oscillations in whole cell currents could be demonstrated. The whole cell conductance increased from 225 ± 25 pS in control solution to 6,728 ± 1,165 pS (means ± SE, n = 17). In substitution experiments (22 mM Cl− in bath solution,[Formula: see text]= 0 mV), the reversal potential changed from −41.6 ± 2.2 mV (means ± SE, n = 9) to −3.2 ± 2.0 mV (means ± SE, n = 7). When the cells were loaded with the calcium-sensitive fluorescent dye, fluo 3, and simultaneously patch clamped, CCh caused a synchronous oscillating pattern of fluorescence and membrane potential. In cell-attached patches, the CCh-activated currents reversed at a relative membrane potential of 1.9 ± 3.7 mV (means ± SE, n = 11) with control solution in the pipette and at 46.2 ± 5.3 mV (means ± SE, n = 10) with a 15 mM Cl− solution in the pipette. High K+ (144 mM) did not change the reversal potential significantly ( P ≤ 0.05, n = 8). In inside-out patches, calcium-dependent Cl−channels could be demonstrated with a conductance of 19 pS ( n = 7). It is concluded that CCh causes oscillations in membrane potential that involve calcium-dependent Cl−channels and a K+ permeability.

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