(Circulation. 1995;92:3497-3504.)
© 1995 American Heart Association, Inc.
Articles |
Delayed Rectifier Channels in Human Ventricular Myocytes
From the Department of Physiology (M.W.V., A.C.G.vG., L.N.B.), Academic Medical Center, and the Department of Clinical and Experimental Cardiology (T.O.), Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
Correspondence to Marieke W. Veldkamp, Department of Physiology, Academic Medical Center, University of Amsterdam, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands.
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Abstract |
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Background Previous studies have shown that in heart there are two kinetically distinct components of delayed rectifier current: a rapidly activating component (IKr) and a more slowly activating component (IKs). The presence of IKr and/or IKs appears to be species dependent. We studied the nature of the delayed rectifier current in human ventricle in whole-cell and single-channel experiments.
Methods and Results Ventricular myocytes were obtained from hearts of patients with ischemic or dilated cardiomyopathy. Single-channel currents and whole-cell tail currents were recorded at negative potentials directly after return from a depolarizing step. Single-channel currents were measured in the cell-attached patch configuration with 140 mmol/L K+ in the pipette. In the present study, we identified a voltage-dependent channel with a single-channel conductance of 12.9±0.8 pS (mean±SEM, n=5) and a reversal potential near to the K+ equilibrium potential, suggesting that the channel is selective to K+ ions. Channel activity was observed only after a depolarizing step and increased with the duration and amplitude of the depolarization, indicating time- and voltage-dependent activation. Activation at +30 mV was complete within 300 milliseconds, and the time constant of activation, determined in the whole-cell configuration, was 101±25 milliseconds (mean±SEM, n=4). The voltage dependence of activation could be described by a Boltzmann equation with a half-activation potential of -29.9 mV and a slope factor of 9.5 mV. The addition of the class III antiarrhythmic drug E-4031 completely blocked channel activity in one patch. No indications for the presence of IKs were found in these experiments.
Conclusions The conformity between the properties of IKr and those of the K+ channel in the present study strongly suggests that IKr is present in human ventricle.
Key Words: ventricles • potassium • delayed rectifyer current
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Introduction |
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TopAbstractIntroductionMethods Results Discussion References |
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A delicate balance between small inward and outward currents exists during the plateau phase of the ventricular action potential. Activation of the delayed rectifier current (IK) initiates the onset of final repolarization. Consequently, the kinetics of this current are an important parameter for action potential duration and refractoriness.1 2 3 4 The delayed rectifier current has gained a lot of interest because blockade of this current by specific class III antiarrhythmic agents may provide a tool for action potential prolongation with concomitant increase in refractory period,5 6 7 which is considered to have an antiarrhythmic effect.8 Voltage-clamp experiments with the class III antiarrhythmic drug E-4031 have revealed the existence of two kinetically and pharmacologically distinct types of delayed rectifier current: a rapidly activating component (IKr), which is blocked by E-4031, and a more slowly activating component (IKs), which is insensitive to E-4031.9 10 Furthermore, IKr activates at relatively negative potentials and shows inward rectification at positive potentials, whereas IKs activates at more positive potentials and has a linear current-voltage relation. The presence of IKr and IKs appears to be species dependent. Although in some species these two types coexist,1 9 10 11 12 other species display either IKs13 14 or IKr.15 16 17 18 19 20 21 The aim of the present study was to investigate at the single-channel level the type of IK that is present in isolated human ventricular myocytes. Data on IK in human heart are scarce. Recently, it was shown that IK is present in both human atrial22 and ventricular23 myocytes, although in ventricle, IK was very small and could be identified in only a minority of the cells. The observation that IK in atrial myocytes is partially blocked by E-4031 demonstrates that both IKr and IKs are present in human atrium.24 Whether IK in human ventricle consists of one or two components has not been elucidated. Results of the present study show for the first time measurements of the delayed rectifier current at the single-channel level in human ventricle. It appears that this current is comparable to IKr in other mammalian species.
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Methods |
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TopAbstractIntroductionMethods Results Discussion References |
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Preparation of Single Ventricular Myocytes
Ventricular myocytes were isolated from explanted human hearts by an enzymatic dissociation procedure. The hearts were obtained from nine patients with end-stage heart failure due to ischemic cardiomyopathy (n=3) or dilated cardiomyopathy (n=6) who were undergoing heart transplantation. Informed consent was obtained before heart transplantation. Directly after explantation, the heart was transported to the laboratory in oxygenated Tyrode's solution (4°C). A portion of the left ventricular free wall was dissected and mounted on a Langendorff perfusion apparatus, where it was perfused through a branch of the left anterior descending coronary artery with solutions in the following sequence: (1) normal Tyrode's solution (10 minutes), (2) Ca2+-free Tyrode's solution (10 minutes), and (3) Ca2+-free Tyrode's solution to which collagenase type B (0.1 to 0.25 mg/mL, Boehringer Mannheim), collagenase type P (0.1 mg/mL, Boehringer Mannheim), and trypsin inhibitor (0.15 mg/mL, Boehringer Mannheim) had been added (20 to 30 minutes). Portions of the left ventricular wall that were clearly digested by the enzymes were cut into small pieces that were gently agitated in a small beaker with Kraft-Brühe (KB)25 solution to obtain single cells. Tissue pieces were taken throughout the ventricular wall, so cell suspension contained cells from epicardial, midcardial, and endocardial origins. During the entire isolation procedure, the solutions were oxygenated and temperature was maintained at 37°C. Cells were allowed to sediment for 15 minutes, after which half of the supernatant was replaced by normal Tyrode's solution to increase the extracellular Ca2+ concentration. This procedure was repeated three times before the experiments.
Solutions and Reagents
The composition of the solutions was
as follows (in mmol/L):
normal Tyrode's solution—NaCl 140, KCl 5.4, CaCl2 1.8,
MgCl2 1.0, glucose 5.5, and HEPES 5.0, pH adjusted to 7.4
with NaOH; Ca2+-free Tyrode's solution—NaCl 140,
KCl 5.4,
MgCl2 0.5, KH2PO4 1.2, glucose 5.5,
and HEPES 5.0, pH adjusted to 6.9 with NaOH; and KB solution—KCl 85,
K2HPO4 30, MgSO4 5.0, glucose 20,
pyruvic acid 5.0, creatine 5.0, taurine 5.0, EGTA 0.5,
ß-hydroxybutyric acid 5.0, succinic acid 5.0,
Na2 · ATP 2.0 and 50 g/L polyvinyl
pyrrolidone · 40, pH adjusted to 6.9 with KOH. Pipette solution for
whole-cell recording of IK tail
currents contained (in mmol/L) KCl 140, K2ATP 5.0, and
HEPES 10, pH adjusted to 7.2 with KOH. Pipette solution for
whole-cell recording of action potentials contained (in
mmol/L) KCl 140, and HEPES 10, pH adjusted to 7.2 with KOH. Pipette
solution for cell-attached patch recording of
single-channel currents contained (in mmol/L) KCl 140,
CaCl2 2.0, MgCl2 1.0, and HEPES 10, pH adjusted
to 7.4 with KOH. E-4031 (Eisai) was prepared as a 1-mmol/L stock
solution.
Recording Procedures
Small aliquots of cell suspension were
introduced to a cell
chamber placed on the temperature-controlled stage of an inverted
microscope (Nikon Diaphot). The isolated myocytes were allowed to
adhere to the bottom of the cell chamber for 3 minutes, after which
perfusion with normal Tyrode's solution was started. The cells were
continuously superfused with normal Tyrode's solution at a rate of
1 mL/min. The temperature of the bath was maintained at 34±2°C.
Currents were recorded in the whole-cell or cell-attached
patch configuration of the patch-clamp technique26
with a laboratory-made patch-clamp amplifier. The pipettes were
pulled from borosilicate glass by a laboratory-made one-stage
puller. The tips of the pipettes were heat polished. After filling with
the appropiate pipette solution, the pipettes had a resistance of 3 to
5 M
.
Action potentials were elicited at 0.5 Hz (0.44 to 0.65
Hz) at a
stimulus strength of 1.5 times diastolic threshold. Action
potential duration was determined with a device that measured the time
when the membrane potential was above an adjustable trigger level. The
level was set to 95% of the amplitude of the action potential. The
voltage-clamp protocol for channel activation consisted of
depolarizing steps of variable amplitude and duration at 1.5- to
3.5-second intervals. Single-channel and whole-cell tail
currents were measured at negative membrane potentials directly after
return from the depolarizing step. With 140 mM K+ in the
pipette, single-channel currents were recorded as inward
currents, and channel openings are plotted as downward deflections in
the recordings. The patch membrane potential was obtained as
the difference between an average resting membrane potential of -70 mV
(-71.2±1.14 [mean±SEM], n=15) and the
patch pipette potential.
Whole-cell and single-channel currents were stored on digital
audio tape with the use of a digital tape recorder (DTR 1200,
Biologic). Membrane currents were off-line filtered (low-pass)
at 0.5 kHz (single-channel currents) or 2 kHz (whole-cell
currents) with a two-pole Butterworth filter, digitized at a
sampling interval of 667 µsec with an AD converter board (National
Instruments), and stored on computer hard disk (Apple Macintosh) for
subsequent data analysis.
Data Analysis
Single-channel recordings were corrected for
leakage
current and capacitive transients and evaluated with the use of a
laboratory-made analysis program. Correction for capacitive
transients was accomplished by subtraction of an exponential function.
The time constant and amplitude of the exponential function were
adjusted until a flat baseline was obtained. Whole-cell currents
were not compensated for capacitance and series resistance. The series
resistance varied between 6.2 and 13 M.
In whole-cell experiments, the degree of activation was determined by measuring the amplitude of the tail current at a fixed potential (-50 mV), directly after a depolarizing step. Tail current amplitude was defined as the difference between the peak current and the steady state current at -50 mV. The amplitude of the tail current is considered to be proportional to the degree of activation during the preceding depolarization. Analogous to this, to estimate the degree of activation in single-channel experiments, one may construct ensemble average currents and measure peak amplitudes. However, the construction of ensemble averaged currents was hampered by the presence of inward rectifier channel activity. Therefore, we estimated the degree of activation in the following, alternative way:
When the voltage, at which current tails are measured, is constant, one may also expect that the time constant of current decay is constant. Therefore, the area-under-the-current tails is also proportional to the degree of activation. The area-under-the-tail current is given by:
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where A is the area-under-the-tail current, ñopen is the mean number of channels open during time T, and i is the single-channel current amplitude. In the experiments presented here, T was the first 800 ms after return to -70 mV. Because T is constant and i is constant (at one particular voltage), ñopen is directly proportional to A and thus to the degree of activation. Consequently, we used ñopen as a measure for the degree of activation in the single-channel experiments.
The mean number of open channels in the patch membrane was derived from amplitude histograms constructed from several traces. The area under each peak in the amplitude histogram, representing the fraction of time that 0, 1, 2, ..., n channels are simultaneously in the open state, was calculated from gaussian functions fitted through these peaks. The height, width, and mean of the gaussian functions were adjusted to the data by the eye. The mean number of open channels was then calculated in the following way:
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where n is the number of channels simultaneously in the open state, nmax is the total number of channels in the patch membrane, and Pn is the fraction of time that n channels are simultaneously in the open state.
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Results |
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TopAbstractIntroductionMethods Results Discussion References |
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Fig 1
shows an action potential recorded in a
cell from a patient with dilated cardiomyopathy at
a 1870-ms pacing cycle length. The action potential configuration is
characterized by a prominent notch during phase 1 and a
long-lasting plateau phase. The action potential duration was 1120
milliseconds (ms). In a total of five cells driven at 0.5 Hz (0.44
to 0.65 Hz), action potential duration was 1247±32 ms
(mean±SEM),
substantially longer than reported in normal human single
ventricular cells.23
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Depolarization Activates a 12.4-pS K+
Channel
Fig 2 shows a typical example of membrane
currents
recorded at -70 mV from a cell-attached patch of a human
ventricular myocyte. Most patches contained inward
rectifier (IKl) channels, identified by openings
of long duration and a single-channel conductance of 41.5 pS (see
Fig 4). This value is close to the values reported for
IKl channels in rabbit
ventricle27 28 29 but somewhat larger
than values reported
for IKl channels in guinea pig
ventricle.30 31 32
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) and inward rectifier (
) channel plotted against the
applied membrane potential for data from five patches. Data from each
experiment were fitted separately by linear regression. Straight line
through the pooled data represents a linear function with
values for slope conductance and extrapolated reversal potential that
are the mean of five experiments. The slope conductance was 12.9±0.8
and 41.5±3.8 pS for the delayed rectifier and inward rectifier
channel, respectively. The extrapolated reversal potential was
-15±7
and -16±5 mV for the delayed rectifier and inward rectifier
channel,
respectively.
To activate
delayed rectifier (IK)
channels, the patch membrane was depolarized from -70 mV to +30 mV
for
500 ms (Fig 2
). During the depolarization to +30 mV, no
channel
activity could be observed. However, after repolarization to -70 mV, a
series of short channel openings was seen, superimposed on either the
open or closed state of the IKl channel.
The unitary current amplitude was approximately one third of that of
the IKl channel. Unlike
IKl channel activity, this type of channel
activity disappeared shortly after repolarization and could be evoked
again only by another depolarizing step. In 6 of 26 ptaches (23%), we
observed this type of channel activity. Although channel openings could
not be observed in any of these patches during depolarization, these
data suggest that this channel is activated during
depolarization and that the decay of channel activity seen after return
to -70 mV is due to deactivation.
Fig 3A shows
another example of this type of channel
activity recorded at -70 mV after a depolarizing step to +30 mV.
In this experiment, the patch was exposed to 10-7 mol/L
E-4031, a class III antiarrhythmic drug that has been shown to
specifically block a component of the delayed rectifier current.
Approximately 1 minute after the superfusion with E-4031, channel
openings occurred less frequently (Fig 3B) and disappeared
completely
after another minute (Fig 3C). Washout did not restore channel
activity
within the duration of the experiment. Because channel activity was not
restored after washout of E-4031, one might argue that rundown is
involved. However, in none of our single-channel experiments was a
significant decrease in channel activity observed during the
experiment. In this particular experiment, channel activity before the
addition of E-4031 had been stable for almost 1 hour, and it is
unlikely that channel activity started to run down exactly at the
moment of addition of the drug. Finally, the time needed for the drug
to block the channels after switching solutions was comparable to the
time needed to block IK channels in rabbit
ventricular myocytes at the same perfusion
rate.29 We refer to this channel as an
IK channel.
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To determine the single-channel
conductance of this channel, we
depolarized the patch membrane to +30 mV and measured the unitary
current amplitude after repolarization to various test potentials (Fig
4). The unitary current amplitude of both the
IKl channel and the IK
channel increased with hyperpolarization (Fig 4A).
The decay of IK channel activity was much faster
at -110 mV than at more depolarized potentials (eg, -70 mV),
suggesting a voltage dependence of deactivation. Fig 4C shows
the
current-voltage relation of both channels. When the data from five
experiments were fit by linear regression, a slope conductance of
12.9±0.8 pS for the IK channel and 41.5±3.8
pS
for the IKl channel was determined.
Extrapolation of the current-voltage curve yielded an estimated
reversal potential of -15±7 mV for the IK
channel, which is near the K+ equilibrium potential (0
mV with 140 mmol/L K+ in the patch pipette). This result
suggests that the investigated channel is selective to K+
ions.
Time Course of Activation: Single-Channel
Analysis
The time course of activation of the human
IK channel was studied by depolarizing
cell-attached patches to +30 mV for various durations and by
measuring single-channel activity after return to -70 mV (as a
measure for the degree of activation). The single-channel currents
(Fig 5, left) were recorded from a cell-attached
patch containing one IKl and one
IK channel. Fig 5 (right) shows the
accompanying
amplitude histograms. In this experiment, a 50-ms lasting
depolarization was too short to elicit IK
channel activity (Fig 5, top left). After repolarization to
-70 mV,
only IKl channel activity was observed, which
resulted in two peaks in the amplitude histogram. One peak corresponds
to the closed IKl channel (0 pA), and one peak
corresponds to the open IKl channel (-2.1 pA).
After return to -70 mV from a 100-ms lasting depolarization, some
traces contained IK channel activity, indicated
by two extra peaks in the amplitude histogram (Fig 5, right,
arrows).
The peak at -0.7 pA corresponds to the open IK
channel when the IKl channel is closed, and the
peak at -2.7 pA corresponds to the open IK
channel when the IKl channel is also open.
Lengthening of the depolarizing pulse to 200 ms resulted in a further
increase in the number of traces containing IK
channel activity. This increase in IK channel
activity can be appreciated by the increase in the area under the peaks
corresponding to the open IK channel in the
amplitude histogram (Fig 5, right). Depolarizations of 300 ms
in
duration did not result in a further increase in
IK channel activity. To determine the degree of
activation during depolarization, the mean number of open channels
(ñopen) during the first 800 ms after return to -70
mV was calculated from the amplitude histograms (see Methods). Values
for ñopen were normalized to the maximum value
(ñopen,max) and were plotted versus the
duration of the depolarization (Fig 6). The time
dependence of activation shows that the increase in
ñopen saturates for depolarizations of 250 ms and
longer, indicating that activation of the IK
channel is complete within this length of time. Similar results were
obtained in an additional two patches. It was not possible to determine
the time constant of activation for these single-channel data. Due
to the limited number of events, there is considerable scatter in the
ñopen values, which makes it difficult to reliably
fit the data points with an exponential function.
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Time Course of Activation: Whole-Cell Tail Current
Analysis
To determine the time course of activation of the macroscopic
IK, whole-cell tail currents were studied. In 9
of 12 cells (75%), small, slowly decaying tail currents were observed
after return from a depolarizing step. In 4 of these cells, tail
current analysis was performed. Fig 7A shows
superimposed whole-cell current traces obtained during
depolarizations from -50 mV to +30 mV of various durations and
subsequent tail currents (IK,tail) after return
to -50 mV. No measures were taken to suppress other
voltage-dependent currents active at depolarized potentials.
Increasing the duration of the depolarizing step clearly increased
IK,tail amplitude up to depolarizations of 300
ms in duration (Fig 7B). When the normalized tail current
amplitude was
plotted versus the duration of the depolarization (Fig 7C),
data points
could be fitted with a monoexponential function of the
form:
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where
A is a free parameter and
represents the maximum value of the exponential function,
t is the duration of the depolarizing step, and 
is the
time constant of activation at this potential (+30 mV). For this
particular experiment, was 140 ms. This type of experiment was
performed in a total of four cells, and the average time constant of
activation was 101±25 ms (mean±SEM).
Voltage Dependence of Activation
Cell-attached patches were
depolarized for 500 ms to various
potentials and channel activity after return to -70 mV was measured to
determine the voltage dependence of activation (Fig 8).
The single-channel currents shown in Fig 8 (left) were recorded
from a cell-attached patch containing one
IKl channel and one IK
channel. Fig 8 (right) shows the accompanying amplitude
histograms.
After return to -70 mV from a depolarization to -50 mV, only a few
traces displayed IK channel activity (Fig
8, top
left). The very small peak indicated by an arrow in the amplitude
histogram corresponds to one open IK channel
when the IKl channel is also open (Fig 8,
top
right). Depolarization to -20 mV clearly increased
IK channel activity, as expressed by the two
peaks in the amplitude histogram. One peak corresponds to the open
IK channel when the IKl
channel is closed, and one peak corresponds to the open
IK channel when the IKl
channel is also open. Depolarization to +10 mV resulted in a further
increase in IK channel activity, as may be
appreciated from the increase in the area under the peaks corresponding
to the open IK channel. Thus,
IK channel activity increases when the preceding
depolarizing step is more positive.
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The ñopen during
the first 800 ms after return to the
resting membrane potential was calculated from the amplitude histograms
and was used as a measure for the degree of activation during the
preceding depolarization. Normalized ñopen values
were plotted versus the potential during depolarization (Fig
9). The data points were fitted by a Boltzmann
equation:
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where C is a free parameter and represents the maximum value of the Boltzmann function, V1/2 is the half-activation potential, Vm is the membrane potential, and K is the slope factor. We found a V1/2 of -29.9 mV and a K of 9.5 mV.
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Discussion |
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TopAbstractIntroductionMethods Results Discussion References |
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In the present study, we describe for the first time measurements of single-channel currents through native delayed rectifier channels in the membrane of human ventricular myocytes. The investigated channel was identified as a delayed rectifier channel on the basis of the following observations: (1) channel activity was observed only after return from a depolarizing step and increased with the duration of the depolarizing step, indicating time-dependent activation on depolarization (Figs 2
and 5); (2) the channel had a slope conductance of
12.9±0.8 pS (mean±SEM)
and an (extrapolated) reversal potential of -15±7 mV, which is
close
to the expected K+ equilibrium potential and suggests that
the channel is highly permeable for K+ ions (Fig
4); (3)
increasing the amplitude of the depolarizing step increased channel
activity, indicating voltage dependence of activation (Figs 8
and 9);
and (4) after return from a depolarizing step, channel activity
recorded at negative membrane potentials decayed with time,
indicating deactivation upon repolarization (Figs 2 and
4).
Comparison to Rapidly Activating Delayed Rectifier Currents in
Other Species
In several species, the cardiac delayed rectifier
current is
composed of a slowly activating component (IKs)
and a more rapidly activating component (IKr),
with different kinetic, conductive, and pharmacological properties.
IKr activates at more negative
potentials than IKs and displays marked
inward-going rectification, whereas IKs has
a linear current-voltage relation. IKr is
specifically blocked by the class III antiarrhythmic agent E-4031, and
IKs is not. The presence of
IKr and/or IKs appears to
depend on species. IKr is the only component in
cat ventricular19 20 21 and rabbit
sinoatrial,
atrioventricular, and Purkinje fiber
myocytes,15 16 17 18 whereas
IKs is the
only component in frog atrial myocytes.13 14 Both
components have been identified in myocytes from guinea pig atrium and
ventricle,9 10 12 sheep Purkinje
fiber,1 and
chick atrium.11 Recently, the presence of both components
has been demonstrated in human atrium.24
The rapid time
course of activation found in the present study
strongly suggests that the IK channel in human
ventricle resembles IKr. Activation at +30 mV
was complete within 300 ms, and the time constant of activation,
obtained by whole-cell tail current analysis, was 101±25
ms (mean±SEM). This value compares well with the activation time
constants at comparable voltages reported for
IKr in human atrium (100 ms),24
mouse atrial tumor myocytes (120 ms),33 and a rapidly
activating IK channel we previously identified
in rabbit ventricle (187 ms)29 but are slower than the
activation time constants for IKr in guinea pig
(20 ms)9 10 and rabbit sinoatrial node (40
ms).16
Other characteristics of this human IK channel are also consistent with those of IKr. The single-channel conductance of 12.9 pS is close to the conductances found for IK channels with relatively rapid kinetics in rabbit sinoatrial node (11.1 pS)16 and ventricle (13.7 pS)29 and guinea pig atrium (10 pS)34 assessed under similar conditions.
IK channel activity could be evoked at
voltages
positive to -60 mV and saturated for depolarizations to 0 mV (Fig
8).
Therefore, the IK channel in the present
study shows a similar voltage dependence as IKr.
Although slightly more negative, the half-maximal activation
potential in human ventricle (-29.9 mV in the present study)
compares with that for IKr in rabbit sinus node
(-25.1 mV)16 and guinea pig atrium and ventricle
(-19.3
and -21.5 mV).9 10 Also, the slope factor (9.5
mV in the
present study) was close to that found in the other studies
(ranging from 5.2 to 7.5 mV).9 10 16 The
fact that we
failed to observe channel openings during depolarizations at positive
potentials may point to inward rectifying properties of this channel,
which is also characteristic of
IKr.9 10 16 24 35
Finally, the
observation that channel activity was completely blocked by E-4031 also
favors a resemblance to IKr.
In the past few years, a number of voltage-gated K+ channels have been cloned from the cardiovascular systems of rat and humans.36 37 38 These cloned channels can approximately be divided into two groups: (1) channels with fast activation and fast inactivation kinetics and (2) channels with fast activation kinetics and no or little, slow inactivation. The latter group consists of delayed rectifier-type channels, but none of these channels resembles the channel presented in the present study because the activation time course of these cloned channels is at least one order of magnitude faster than the human IK channel in this study.
In our experiments, we
found no indication of the presence of
IKs at the single-channel level. It has been
reported, however, that the single-channel conductance of
IKs is very small (<1 pS).39
Therefore, it is conceivable that IKs is
present in human ventricle but remains unnoticed in
single-channel recordings. Also, no observations were made
in the whole-cell recordings that pointed to the presence
of IKs. When IKs
contributes significantly to membrane current, this current should have
been recognized in whole-cell currents. However, the outward
current during depolarization was very small (Fig 6A), even at
longer
depolarizations (900 ms). Moreover, the increase in whole-cell tail
current amplitude saturated for depolarizations of 300 ms in duration.
These observations suggest that IKs in human
ventricle is small or absent. Similar findings were obtained in the
whole-cell voltage clamp study of Beuckelmann et al.23
They showed that in human ventricle, IK is
small (or absent) and is completely activated within 250 ms,
suggesting the presence of an IK current
different from IKs.
It must be noted, however, that
the ventricular myocytes in
this study had extremely long action potentials (Fig 1), even
for
cardiomyopathy. The low stimulus frequency is
certainly one of the causes of prolonged action potential duration. The
experimental temperature was approximately 3°C below the
physiological temperature and is another factor for
action potential prolongation. In addition, repolarization in these
cells may have been altered by the isolation procedure. Therefore, it
cannot be excluded that the absence of the slow component of
IK may be due to the diseased state of the cells
or to altered membrane properties caused by the isolation procedure
rather than true absence of this component. Also, the possibility of
rundown of IKs channel activity in the
whole-cell configuration cannot be ignored. Therefore, we cannot
exclude a role for IKs in the nondiseased human
ventricle.
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Acknowledgments |
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The authors thank Jacques de Bakker, Jessica Vermeulen, Ton Baartscheer, and Cees Schumacher for their help in the preparation of the cells. The hearts were made available by the Department of Cardiology, A2U, Utrecht.
Received February 27, 1995; revision received July 20, 1995; accepted July 23, 1995.
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References |
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TopAbstractIntroductionMethods Results Discussion References |
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