Download Effects of endothelin-1 on calcium and potassium currents in

Pflügers Arch - Eur J Physiol (2000) 441:144–149
DOI 10.1007/s004240000400
O R I G I N A L A RT I C L E
J. Magyar · N. Iost · Á. Körtvély · T. Bányász
L. Virág · P. Szigligeti · A. Varró · M. Opincariu
J. Szécsi · J.G. Papp · P.P. Nánási
Effects of endothelin-1 on calcium and potassium currents
in undiseased human ventricular myocytes
Received: 27 March 2000 / Received after revision: 30 May 2000 / Accepted: 29 June 2000 / Published online: 7 September 2000
© Springer-Verlag 2000
Abstract Endothelins have been reported to exert a
wide range of electrophysiological effects in mammalian
cardiac cells. These results are controversial and human
data are not available. Our aim was to study the effects
of endothelin-1 (ET-1, 8 nmol/l) on the L-type calcium
current (ICa-L) and various potassium currents (rapid
component of the delayed rectifier, IKr; transient outward
current, Ito; and the inward rectifier K current, IK1) in
isolated human ventricular cardiomyocytes. Cells were
obtained from undiseased donor hearts using collagenase
digestion via the segment perfusion technique. The
whole-cell configuration of the patch-clamp technique
was applied to measure ionic currents at 37°C. ET-1
significantly decreased peak ICa-L from 10.2±0.6 to
6.8±0.8 pA/pF at +5 mV (66.7% of control, P<0.05,
n=5). This reduction of peak current was accompanied
by a lengthening of inactivation. The voltage dependence
of steady-state activation and inactivation was not altered by ET-1. IKr, measured as tail current amplitudes at
–40 mV, decreased from 0.31±0.02 to 0.06±0.02 pA/pF
(20.3% of control, P<0.05, n=4) after exposure to ET-1.
ET-1 failed to change the peak amplitude of Ito, measured at +50 mV (9.3±4.6 and 9.0±4.4 pA/pF before and
after ET-1, respectively), or steady-state IK1 amplitude,
measured at the end of a 400-ms hyperpolarization to
J. Magyar · Á. Körtvély · T. Bányász · P. Szigligeti · P.P. Nánási (✉)
Department of Physiology, University Medical School of Debrecen,
H-4012 Debrecen, PO Box 22, Hungary
e-mail: [email protected]
Tel.: +36-52-416634, Fax: +36-52-432289
L. Virág · A. Varró · J.G. Papp
Department of Pharmacology and Pharmacotherapy,
Faculty of Medicine, University of Szeged, H-6701 Szeged,
PO Box 427, Hungary
N. Iost · J.G. Papp
Research Unit for Cardiovascular Pharmacology,
Hungarian Academy of Sciences, H-6701 Szeged, PO Box 427,
Hungary
M. Opincariu · J. Szécsi
Department of cardiac surgery, Faculty of Medicine,
University of Szeged, H-6701 Szeged, PO Box 427, Hungary
–100 mV (3.6±1.4 and 3.7±1.4 pA/pF, n=4). The present
results indicate that in undiseased human ventricular
myocytes ET-1 inhibits both ICa-L and IKr; however, the
degree of suppression of the two currents is different.
Keywords Action potentials · Calcium currents ·
Cardiac cells · Endothelins · Human myocytes ·
Potassium currents
Introduction
Endothelins are recently described vasoactive peptides released from vascular and endocardial endothelium. They
consist of a group of three isopeptides, of which endothelin-1 (ET-1) is an extremely potent vasoconstrictor [11,
35]. Endothelin receptors have been detected in various
cardiac membranes [8, 26] and increasing evidence suggests that ET-1 may be an important factor in cardiovascular regulation under physiological [7, 11, 17, 26] and
various pathological [1, 24, 25, 32, 33] conditions.
There is controversy about the effects of ET-1 on cardiac cells, and they strongly depend on the experimental conditions used as well as the origin and developmental state
of the preparation studied. ET-1 has a positive inotropic action in several mammalian cardiac preparations [6, 18, 21,
27]; however, according to other investigators, the effects
of ET-1 are rather similar to those of acetylcholine. ET-1
causes a hyperpolarization, shortens action potentials and
reduces heart rate through inhibition of L-type Ca current
(ICa-L) and activation of K currents [4, 19, 28, 34, 37]. Furthermore, ET-1 inhibits ATP-dependent K current (IK(ATP))
in ischemic myocardium [20] and also the protein-kinaseA-dependent Cl– current [15, 16] in rabbit and guinea pig
myocytes. These effects of ET-1 are accompanied by a fall
in the intracellular cAMP level [12, 34], which is sensitive
to pertussis toxin [28]. Since the inhibitory actions of ET-1
are greatly enhanced following isoproterenol treatment,
the general picture emerged that the main mechanism of
action of ET-1 is to inhibit adenylate cyclase via G-proteincoupled ETA receptors [9, 15, 16, 34].
145
The aim of the present work was to study the effects
of ET-1 on calcium and potassium currents in ventricular
myocytes isolated from undiseased human hearts. Due to
the lack of relevant human data on endothelin and the diversity of results obtained from other mammalian cells,
the present results may help to elucidate how ET-1 exerts
its effects on the human heart.
Materials and methods
Cell isolation
Human ventricular cells were prepared from five donor hearts.
The hearts were obtained from general organ donor patients (four
males and one female, with an average age of 43.7 years) whose
semilunar valves were used for transplantation. Before explantation of their hearts, the patients did not receive any medication
other than dobutamine, furosemide and plasma expanders. The
myocytes were isolated using a recently developed enzymatic dissociation procedure. After explantation and valve removal, the
hearts were transported to the laboratory in cold (4°C) cardioplegic solution. Part of the left ventricular wall was excised together
with its arterial branch and was mounted on a modified Langendorff apparatus, where it was perfused through the left anterior descending coronary artery according to the following sequence: (1)
modified Tyrode solution (containing, in mmol/l: NaCl, 135; KCl,
4.7; KH2PO4, 1.2; MgSO4, 1.2; HEPES, 10; NaHCO3, 4.4; glucose 10; pH=7.2) for 10 min; (2) Ca2+-free modified Tyrode for
10 min; (3) Ca2+-free modified Tyrode containing collagenase
(660 mg/l, type I, Sigma), elastase (45 mg/l, type III, Sigma), taurine (50 mmol/l) and bovine albumin (2 g/l, fraction V, fatty acid
free, Sigma) for 45 min; (4) after this step of enzymatic digestion
the solution was supplemented with protease (120 mg/l, type XIV,
Sigma) for a further 40–60 min. Portions of the left ventricular
wall, which was clearly digested by the enzymes, were cut into
small pieces and either stored in KB medium [14], or equilibrated
for 15 min in modified Tyrode containing 1.25 mmol/l CaCl2 and
50 mmol/l taurine. Single myocytes were obtained from the tissue
chunks after gentle agitation. The solutions were oxygenated
(100% O2) and the temperature was maintained at 37°C throughout the whole isolation procedure. The cells were allowed to sediment for 10 min, then the supernatant was decanted and replaced
by fresh solution. This procedure was repeated three times. The
cells in KB medium were stored at 4°C, while those stored in
modified Tyrode solution were maintained at 12–14°C before use.
Recording of action potentials
Thin trabecular muscle bundles from the left ventricle were dissected before cell digestion. These preparations were individually
mounted in a Plexiglass chamber allowing continuous superfusion
with Krebs solution, containing (in mmol/l): NaCl, 115; KCl, 5.6;
CaCl2, 2.5; MgCl2, 1; NaH2PO4, 1; NaHCO3, 26; glucose 10, pH
7.4±0.05 when gassed with a mixture of 95% O2 and 5% CO2.
Transmembrane potentials were recorded at 37°C using conventional glass microelectrodes, filled with 3 mol/l KCl, and whose
tip resistance was 10–20 MΩ. These electrodes were connected to
the input of an Axoclamp-2B amplifier (Axon Instruments). The
preparations were continuously paced at 1.2 Hz through a pair of
platinum electrodes using 1-ms-wide rectangular current pulses
with 120% threshold amplitude. Action potentials were digitized
at 100 kHz using a Digidata 1200 A/D card (Axon Instruments)
and stored for later analysis.
Voltage clamp
Experiments were performed on Ca2+-tolerant cells, which were
rod shaped and maintained clear cross-striations following expo-
sure to oxygenated Tyrode solution containing (in mmol/l): NaCl,
140; KCl, 5.4, CaCl2, 2.5; MgCl2, 1.2; Na2HPO4, 0.35; HEPES, 5;
glucose, 10 at pH 7.4. This solution was supplemented with either
3 mmol/l 4-aminopyridine or 0.25 mmol/l CdCl2, when measuring
calcium or potassium currents, respectively. Suction pipettes, fabricated from borosilicate glass (Clark), had tip resistances of
2–3 MΩ after filling with pipette solution. When measuring potassium currents, the solution composition was (in mmol/l): Kaspartate, 100; KCl, 20; Mg-ATP, 5; HEPES, 10; K4BAPTA, 5;
glucose, 5. When measuring calcium current the pipette solution
contained (in mmol/l): KCl, 110; KOH, 40; K-ATP, 3; HEPES, 10;
EGTA, 10; tetraethylammonium chloride (TEACl), 20; glucose, 5;
GTP, 0.25. The pH of both solutions was adjusted to 7.2 using
KOH. Membrane currents were recorded at 37°C using an Axopatch-1D amplifier (Axon Instruments) in the whole-cell configuration of the patch-clamp technique [10]. After establishing a
high-resistance (1–10 GΩ) seal by gentle suction, the cell membrane beneath the tip of the electrode was disrupted by further suction or by applying 1.5-V electrical pulses for 1–5 ms. The series
resistance was typically 4–8 MΩ before compensation (usually
50–80%). Experiments were discarded when the series resistance
was high or substantially increasing during the measurement. Current densities are expressed by current amplitudes normalized to
cell capacitance measured using 10-mV hyperpolarizing pulses
applied from –10 mV. Outputs of the clamp amplifier were digitized at 100 kHz using an A/D converter (Digidata-1200, Axon Instruments) under software control (pClamp 6.0, Axon Instruments). Data were stored on video tape for later analysis. The experimental protocol for each measurement is described where pertinent in the Results.
Results are expressed as mean ±SEM values. Student’s t-test
for paired data was used to determine statistical significance.
Changes were considered significant when P was less than 0.05.
The entire investigation conforms with the principles outlined in
the Declaration of Helsinki. The experimental protocol applied to
human hearts was also approved by the local ethics committee
(No. 51–57/1997OEJ).
Results
Calcium current
ICa-L was measured at a rate of 0.2 Hz using depolarizing
voltage pulses of 400 ms duration clamped from the holding potential of –40 mV to test potentials increasing from
–35 mV up to +60 mV in 5-mV steps. Families of ICa-L
were recorded before and 5 min after ET-1 treatment.
Superfusion with 8 nmol/l ET-1 significantly decreased
ICa-L at all membrane potentials studied (Fig. 1A). At
+5 mV, ICa-L was reduced by ET-1 from 10.2±0.6 to
6.8±0.8 pA/pF (66.7±8.9% of control, P<0.05, n=5). This
reduction of peak current was accompanied by a lengthening of inactivation. Inactivation of ICa-L was fitted as a
sum of two exponentials, with estimated time constants of
12.8±1.1 and 97.4±11 ms at +5 mV in Tyrode solution.
Both time constants were increased by ET-1: to 17.2±1.2
and 122±9 ms, respectively (Fig. 1B). Lengthening of the
faster component was statistically significant (P<0.05,
n=5), while that of the slower component was not. The
voltage dependence of steady-state activation and inactivation was not altered by ET-1. The effect of ET-1 on
peak ICa-L developed rapidly (within 2 min) and was not
reversible within a 7-min period of superfusion with ET-1free Tyrode solution (Fig. 1C). ET-1 did not affect the
voltage dependence of activation of ICa-L: half-activation
146
Fig. 1A–E Effect of 8 nmol/l endothelin-1 (ET-1) on L-type calcium current (ICa-L) in human ventricular myocytes (n=5). A Current–voltage (I–V) relationship obtained for peak calcium current
in control conditions (squares) and in the presence of 8 nmol/l
(triangles). The I–V curve was obtained by plotting peak current
densities against test pulse potential. The holding potential was
–40 mV. B Superimposed records of ICa-L measured at +5 mV before and after ET-1 treatment. C Time course of development and
lack of reversibility of ET-1’s effect on peak ICa-L. D Voltage dependence of steady-state activation of ICa-L. Peak calcium current
was divided by driving force for ICa-L, and the calcium conductance (GCa-L) calculated at each membrane potential was normalized to that obtained at a test pulse potential of +25 mV. The
results were fitted to a two-state Boltzmann model (solid curves).
E Steady-state inactivation of ICa-L determined using a pairedpulse protocol. Test pulses to +5 mV were preceded by a set of
prepulses clamped to various voltages between –55 and +15 mV.
Peak currents measured after these prepulses were normalized to
the peak current measured after the –55-mV prepulse and plotted
against the respective prepulse potential. Solid curves were obtained by fitting data to a two-state Boltzmann model. Symbols
and bars represent means ±SEM values
voltages and slope factors, obtained by fitting the activation curves to a two-state Boltzmann model, were –4.7±0.7
versus –4.9±0.7 mV, and 3.7±0.3 versus 4.1±0.8 mV in
absence and presence of ET-1, respectively, (n=5, N.S.,
Fig. 1D). The steady-state inactivation of ICa-L was determined using a paired-pulse protocol. Test pulses to +5 mV
were preceded by a set of prepulses clamped to various
voltages between –55 and +15 mV. Peak currents measured after these prepulses were normalized to the peak
current measured after the –55-mV prepulse and plotted
against the respective prepulse potential. As shown in
Fig. 1E, steady-state inactivation curves, obtained in the
absence and presence of ET-1, were almost identical (midpoint potentials and slope factors were –19.3±1.2 versus
–21.6±0.7 mV, and 3.45±0.27 versus 3.83±0.63 mV, respectively, n=5, N.S.).
The effect of ET-1 on ICa-L was also studied in two
myocytes pretreated with 50 nmol/l isoproterenol (not
shown). Isoproterenol is known to increase ICa-L in various mammalian cardiac preparations. In our two cells
isoproterenol increased peak ICa-L from 1.27 to 2.03 nA
and from 1.03 to 1.5 nA (to 160% and 146% of their
control values). Application of 8 nmol/l ET-1 in the presence of isoproterenol reduced peak ICa-L to 1.66 nA and
1.36 nA, respectively (reduction of 18.3% and 9.4%).
Again, the effect of ET-1 was practically irreversible,
since removal of ET-1 caused no change in ICa-L despite
the continuous presence of isoproterenol.
Potassium currents
IKr was activated using depolarizing voltage pulses of
1000 ms duration clamped from a holding potential of
–40 mV to test potentials ranging between –20 and
+50 mV. The decaying tail current recorded at –40 mV after the end of the test pulse was assessed as IKr. The
current was fully abolished by 5 µmol/l E-4031, the selective blocker of IKr. ET-1 significantly decreased the amplitude of the tail current at each membrane potential studied (Fig. 2). Following depolarization to +40 mV, ET-1
decreased IKr tails from 0.31±0.02 to 0.06±0.02 pA/pF
(20.3% of control, P<0.05, n=4).
Transient outward K current (Ito) was measured at a
rate of 0.2 Hz using depolarizing voltage pulses of
400 ms duration clamped from a holding potential of
–80 mV to test potentials ranging between –10 and
+60 mV (Fig. 3A). ET-1 failed to change the peak amplitude of Ito (9.3±4.6 and 9.0±4.4 pA/pF at +50 mV before
and after ET-1, respectively, n=4, N.S). The steady-state
current–voltage relationship of the membrane was studied between –130 and 0 mV at the end of the 400-ms de-
147
Fig. 2 Effect of ET-1 (8 nmol/l) on the rapid component of the delayed rectifier potassium current, IKr, in human ventricular myocytes. The current was activated by a 1000-ms-long depolarization, arising from the holding potential of –40 mV, to test potentials ranging between –20 and +50 mV. The amplitude of IKr was
measured as the amplitude of the tail current obtained upon repolarization to –40 mV. In the experiment displayed on the left ET-1
almost fully abolished IKr. On the right average tail current amplitudes (n=4) are plotted against test potentials before (diamonds)
and 5 min after application of ET-1 (squares)
Fig. 4 Effect of ET-1 and isoproterenol on action potential configuration in a left ventricular trabecula, recorded using a conventional microelectrode. After taking control records 8 nmol/l ET-1
was applied for 15 min, then the preparation was exposed to
50 nmol/l isoproterenol for an additional 15 min in the continuous
presence of ET-1
polarization. At these potentials the current is most likely
IK1. As Fig. 3B shows, no change of this current (IK1)
was noted after superfusion with 8 nmol/l ET-1. Applying hyperpolarization to –100 mV, the IK1 current densities were 3.6±1.4 and 3.7±1.4 pA/pF in control conditions and in the presence of ET-1, respectively (n=4,
N.S.).
Action potential characteristics
Fig. 3 Effect of ET-1 (8 nmol/l) on transient outward current, Ito
(A), and the steady-state current–voltage relationship most likely
representing the inward rectifier potassium current, IK1 (B), in human ventricular myocytes. Test pulses of 400 ms duration were
applied to test potentials ranging between –130 and +60 mV, the
holding potential was –80 mV. Left panels show families of Ito and
IK1 currents recorded before and 5 min after application of ET-1.
Peak amplitudes of Ito are plotted against test pulse potential
(upper right panel). The steady-state current–voltage relationship
of the membrane was studied between –130 and 0 mV at the end
of the 400-ms depolarization (lower right panel). The negative
branch of the I–V curve represents IK1. Symbols (squares for control, triangles for ET-1) and bars represent arithmetic means
±SEM, obtained from four cells
In multicellular left ventricular muscle preparations
(n=3) paced at 1.2 Hz, ET-1 caused a moderate change in
action potential morphology. Action potential duration,
measured at 50% of repolarization (APD50), was slightly
increased (from 146±6 to 160±3 ms) by 8 nmol/l ET-1.
This effect was reverted by isoproterenol. Application of
isoproterenol (50 nmol/l) in the presence of ET-1 shortened APD50 from 160±3 to 144±11 ms, a value close to
the control level (Fig. 4). The corresponding changes in
APD90 were less pronounced (APD90 values of 213±7,
221±3 and 209±7 ms were obtained in control, after application of ET-1, and in the presence of ET-1 plus isoproterenol, respectively). ET-1 had no effect on the resting potential (–81.8±3.3 versus –83.2±1.9 mV), action
potential amplitude (106.7±1.4 versus 107.8±4.6 mV)
and maximum rate of depolarization (215±33 versus
204±21 V/s), measured before and after superfusion with
ET-1, respectively.
Discussion
The major result of the present work is to show that in
undiseased human ventricular cells ET-1 suppresses both
ICa-L and IKr without changing Ito and IK1. Considering
the inhibitory action of ET-1 on ICa-L in human cells, our
results are similar to those obtained in studies of atrial
148
[4, 28] and ventricular [22, 34] myocytes of rabbits and
guinea pigs. Suppression of ICa-L by ET-1 was also similar in human and canine ventricular cells (our unpublished results). Our results with ET-1 on human ICa-L,
however, are markedly different from those of Delpech
et al. [5] obtained in rat atrial cells. In that study
50 nmol/l ET-1 effectively reversed the isoproterenolinduced increase in ICa-L, leaving basal ICa-L unaltered. In
addition, ET-1 was shown to enhance ICa-L in embryonic
human and chick ventricular cells [2]. These results suggest that ET-1-dependent control of ICa-L may be very
different in atrial and ventricular myocytes, as well as in
embryonic and adult cardiac tissues.
Regarding potassium currents, the effects of ET-1 differ greatly between species. IK is inhibited by ET-1 in rat
myocytes [3], but is enhanced in guinea pig cells [23],
whereas in canine ventricular myocytes IK is not affected
by ET-1 (our unpublished results). This latter result is
strongly different from the marked suppression of IKr observed in the present study of human cells. As in canine
cells, ET-1 failed to modify IK in embryonic human and
chick myocytes [2]. Although ET-1 depresses IKs in
guinea pig ventricular myocytes studied under conventional whole-cell voltage-clamp conditions, this effect
failed to develop when perforated patch-clamp techniques were used [31]. In contrast to guinea pig, in
which IKs is a strong repolarizing current [29], little IKs
could be demonstrated in human ventricular cells [13].
Therefore, no attempt was made in the present study to
investigate the effect of ET-1 on IKs. In contrast to the
present results obtained from human ventricular cells,
ET-1 increases IK1 in atrial myocytes of rat, rabbit and
guinea pig [19, 28], but suppresses the acetylcholineactivated K current in rabbit atrial cells [30]. As in humans, neither IK1 nor Ito is influenced by ET-1 in canine
ventricular myocytes (unpublished results).
In the present study 8 nmol/l ET-1 had no effect on
human action potential configuration, except to moderately lengthen APD. This result is similar to those obtained in studies of canine and rat ventricular cells [3,
36]. In contrast, APD was shortened by ET-1 in guinea
pig and rabbit atrial myocytes [28]. The ET-1-induced
lengthening of APD is consistent with inhibition of IKr
and ICa-L, considering that suppression of IKr is more
prominent than reduction of ICa-L. The comparison of data obtained in multicellular and single cell preparations,
however, may be hampered by differences in experimental conditions; for example, using different bathing solutions or the presence of EGTA in the pipette solution in
voltage-clamp studies.
Although it was not studied intensively in the present
study, ET-1 clearly antagonized the effects of isoproterenol on APD as well as on ICa-L in human ventricular
cells. From this point of view it is worth noting that the
inhibitory effect of ET-1 on ICa-L was less pronounced in
the presence than in the absence of isoproterenol. Although these experiments may not be conclusive because
of their limited number (n=2), these results are in contrast with those of Delpech et al. [5] obtained from rat
atrial cells. An increased sensitivity to ET-1 might be expected following isoproterenol treatment if the action of
ET-1 were mediated mainly by decreasing intracellular
cAMP levels [9, 15, 16, 34], as was observed in rabbit
myocytes [28], but not in guinea pig cells [4, 5]. Our results suggest that this is not the case in human ventricular myocardium. It is possible, therefore, that the electrophysiological effects of endothelins may involve multiple mechanisms of action in human cardiac cells; however, further detailed studies are required to elucidate this
point. In conclusion, the marked interspecies differences
found with ET-1 indicate that the actions of this peptide
in humans cannot be deduced from data obtained in studies of other mammalian species.
Acknowledgements This study was supported by grants from
the Hungarian National Research Foundation (OTKA T-026577,
T-032558) and the Hungarian Ministry of Education (FKFP0243/2000, FKFP-1025/1997) and from the Hungarian Academy
of Sciences. Further support was obtained from Bolyai Fellowship.
References
1. Arendt RM, Wilbert-Lampen U, Heucke L, Schmoeckel M,
Suhler K, Richter WO (1991) Increased endothelin plasma
concentration in patients with coronary artery disease or hyperlipoproteinemia without coronary events. Res Exp Med
(Berl) 193:225–230
2. Bkaily G, Wang S, Bui M, Menard D (1995) ET-1 stimulates
Ca2+ currents in cardiac cells. J Cardiovasc Pharmacol 26:
S293–S296
3. Damron DS, Van Wagoner DR, Moravec CS, Bond M (1993)
Arachidonic acid and endothelin potentiate Ca transients in rat
cardiac myocytes via inhibition of distinct K channels. J Biol
Chem 268:27335–27344
4. Delpech N, Soustre H, Potreau D (1995) Antagonism of betaadrenergic stimulation of L-type Ca current by endothelin in
guinea-pig atrial cells. Eur J Pharmacol 285:217–220
5. Delpech N, Soustre H, Potreau D (1997) Endothelin-1 inhibits
L-type Ca2+ current enhanced by isoprenaline in rat atrial
myocytes. J Cardiovasc Pharmacol 29:136–143
6. Eid H, O’Niel M, Kraemer B, Reus M, Kelly R, Smith TW
(1989) Endothelin, a potent positive inotrope in isolated rat
ventricular myocytes. Circulation 80S-II:193
7. Evans HG, Lewis MJ, Shah AM (1994) Modulation of myocardial relaxation by basal release of endothelin from endocardial endothelium. Cardiovasc Res 28:1694–1699
8. Gu X-H, Casley D, Nayler W (1989) Specific high-affinity
binding sites for 125I-labelled porcine endothelin in rat cardiac
membranes. Eur J Pharmacol 167:281–290
9. Haber E, Lee M-E (1994) Endothelin to the rescue? Nature
370:252–253
10. Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ
(1981) Improved patch-clamp techniques for high-resolution
current recording from cells and cell-free membrane patches.
Pflügers Arch 391:85–100
11. Haynes WG, Webb DJ (1993) The endothelin family of peptides: local hormones with diverse roles in health and disease?
Clin Sci 84:485–500
12. Hilal-Dandan R, Urasawa K, Brunton LL (1992) Endothelin
inhibits adenylate cyclase and stimulates phosphoinositide hydrolysis in adult cardiac myocytes. J Biol Chem 267:10620–
10624
13. Iost N, Virág L, Opincariu M, Szécsi J, Varró A, Papp JG
(1998) Delayed rectifier potassium current in undiseased human ventricular myocytes. Cardiovasc Res 40:508–515
149
14. Isenberg G, Klöckner U (1982) Isolated bovine ventricular
myocytes. Characterization of the action potential. Pflügers
Arch 395:19–29
15. James AF, Xie LH, Fujitani Y, Hayashi S, Horie M (1994) Inhibition of the cardiac protein kinase A-dependent chloride
conductance by endothelin-1. Nature 370:297–300
16. James AF, Xie LH, Horie M (1994) The effects of endothelin1 on the PKA-dependent chloride current in the heart. Jpn J
Physiol 44:S227–S230
17. Karwatowska-Prokopczuk E, Wennmalm A (1990) Effects of
endothelin on coronary flow, mechanical performance, oxygen
uptake, and formation of purines and on outflow of protacyclin in the isolated rabbit heart. Circ Res 66:46–54
18. Kelly RA, Eid H, Kramer BK, O’Neill M, Liane BT, Reers M,
Smith TW (1990) Endothelin enhances the contractile responsiveness of adult rat ventricular myocytes to calcium by a pertussis sensitive pathway. J Clin Invest 86:1164–1171
19. Kim D (1991) Endothelin activation of an inwardly rectifying
K current in atrial cells. Circ Res 69:250–255
20. Kobayashi S, Nakaya H, Takizawa T, Hara Y, Kimura S, Saito
T, Masuda Y (1996) Endothelin-1 partially inhibits ATPsensitive K current in guinea pig ventricular cells. J Cardiovasc Pharmacol 27:12–19
21. Kramer BK, Smith T, Kelly RA (1991) Endothelin and increased contractility in adult rat ventricular myocytes. Role of
intracellular alkalosis induced by activation of the protein kinase C-dependent Na+/H+ exchanger. Circ Res 68:269–279
22. Lauer MR, Gunn MD, Clusin WT (1992) Endothelin activates
voltage-dependent Ca current by a G protein-dependent mechanism in rabbit cardiac myocytes. J Physiol (Lond) 448:729–
747
23. Lu T, Huang Y, Jiang W (1995) The electrophysiological effects of endothelin. A patch clamp study in guinea pig ventricular myocytes. Chin Med J 108:618–625
24. McMurray JJM, Ray SG, Addullah I, Dargie HJ, Morton JJ
(1992) Plasma endothelin in chronic heart failure. Circulation
85:1374–1379
25. Miyauchi T, Yanagisawa M, Tomizawa T, Sugishita Y, Suzuki
N, Fujino M, Ajisaka R, Masaki T (1989) Increased plasma
concentrations of endothelin-1 and big endothelin-1 in acute
myocardial infarction. Lancet 2:53–54
26. Moody CJ, Dashwood MR, Sykes RM, Chester M, Jones SM,
Yacoub MH, Harding SE (1990) Functional and autoradio-
27.
28.
29.
30.
31.
32.
33.
34.
35.
36.
37.
graphic evidence for endothelin-1 receptors on human and rat
cardiac myocytes. Circ Res 67:764–769
Moravecz CS, Reynolds EE, Stewart RW, Bond M (1989) Endothelin is a positive inotropic agent in human and rat heart in
vivo. Biochem Biophys Res Commun 159:14–19
Ono K, Tsujimoto G, Sakamoto A, Eto K, Masaki T, Ozaki Y,
Satake M (1994) Endothelin-A receptor mediates cardiac inhibition by regulating calcium and potassium currents. Nature
370:301–304
Sanguinetti MC, Jurkiewicz NK (1990) Two components of
cardiac delayed rectifier K+ current. Differential sensitivity to
block by class III antiarrhythmic agents. J Gen Physiol 96:
195–215
Spiers P, Kelso EJ, McDermott BJ, Scholfield CN, Silke B
(1996) Endothelin-1 mediated inhibition of the acetylcholineactivated potassium current from rabbit isolated atrial cardiomyocytes. Br J Pharmacol 119:1427–1437
Washizuka T, Horie M, Watanuki M, Sasayama S (1997) Endothelin-1 inhibits the slow component of cardiac delayed rectifier K+ currents via a pertussis toxin-sensitive mechanism.
Circ Res 81:211–218.
Watanabe T, Suzuki N, Shimamoto N, Fujino M, Imada A
(1991) Contribution of endogenous endothelin to the extension
of myocardial infarct size in rats. Circ Res 69:370–377
Wieczorek I, Haynes WG, Webb DJ, Ludma CA, Fox AD
(1994) Raised plasma endothelin in ubstable angina and non-Q
wave myocardial infarctioin: relatiion to cardiovascular outcome. Br Heart J 72:436–441
Xie LH, Horie M, James AF, Watanuki M, Sasayama S (1996)
Endothelin-1 inhibits L-type Ca currents enhanced by isoproterenol in guinea-pig ventricular myocytes. Pflügers Arch
431:533–539
Yanagisawa M, Kurihara H, Kimura S, Tomobe Y, Kobayasi
M, Mitsui Y, Yazaki Y, Goto K, Masaki T (1988) A novel potent vasoconstrictor peptide produced by vascular endothelial
cells. Nature 332:411–415
Yorikane R, Koike H, Miyake S (1991) Electrophysiological
effects of endothelin-1 on canine myocardial cells. J Cardiovasc Pharmacol 17:S159–S162
Zhang Z, Li YL, Ho SY (1996) Effects of endothelin on the
electrical activity of sino-atrial pacemaker cells of rabbit.
Sheng Li Hsueh Pao 48:53–58