Cardiovascular
Research
ELSEVIER
Cardiovascular Research 3 I (I 996) X20-825
Decrease in Ca2+ sensitivity as a mechanism of hydrogen
peroxide-induced relaxation of rabbit aorta
Takafumi Iesaki a’* , Takao Okada b, Issei Shimada a, Hiroshi Yamaguchi a, Rikuo Ochi b
a Diuision
cj’cardiology,
Department
oj’Internu1
Medicine,
Juntmdo
University
School
ofMedicine,
b Depnrtmrnt
qfPhysiolo,qy,
Juntendo
Uniuersity
School of‘Mnrdicinr.
2-l-1,
Hoqo,
2-I-1,
Hongo,
Bunky,o-ku,
Tohqw
Bunkyo-ku,
Tokyo 113, Japan
113, Jupun
Received 12 July 1995; accepted 21 November 1995
Abstract
Objective: In vascular
strips, hydrogen peroxide (H,O,) relaxes cr,-adrenergic agonist-induced but not high-K+-induced
contractions.
The aim of this study was to explore H,O,-induced
changes in [Ca”+]; of vascular smooth muscle and to elucidate the mechanisms of
action of H,O,. Methods: Isolated rabbit aortic strips were isometrically contracted with high-K* (64.7 mM) or phenylephrine (PE, 0.3
PM). The effects of 300 PM H,O, on [Ca2+], of endothelium-denuded
vascular smooth muscle and tension were determined
simultaneously by the fura- method. Changes in [Ca2+], were expressed as percentages of high-K+-induced
values measured at the
beginning of the experiments. In another series of experiments, the relaxant effect of 300 /LM H,O, was examined in high-K+ (20
mM)-induced contraction in the presence of the protein kinase C activator, phorbol 12,13-dibutyrate (PDBu). Results: Hydrogen peroxide
caused a reversible rise in [Ca”],
of vascular smooth muscle under both resting conditions and in the precontracted state. During
high-K+-induced
contraction, H,O, further increased [Ca*+], by 26.6ts.e.m. 1.7>% accompanied by a small increase in tension of
6.5(1.9)% of high-K+-induced
tension. By contrast, during PE-induced contraction, although H,O, caused a comparable additional
increase in [Ca*+], (26.4(4.7)%), muscle tension fell by 28.9(2.2)0/o of the steady-state PE-induced tension. Hydrogen peroxide had a
relaxant effect on augumented high-K+-induced
contraction in which Ca2+ sensitivity of the contractile apparatus was elevated by PDBu.
Conclusions: In spite of its effect of increasing [Ca2+li of vascular smooth muscle, hydrogen peroxide causes relaxation of
endothelium-denuded,
PE-precontracted rabbit aorta. The mechanism is probably through suppression of agonist-induced augmentation of
Ca2+ sensitivity of the contractile apparatus.
Keywords: Free radicals; Reperfusion; Calcium, intracellular concentration; Fura-2; Vascular smooth muscle; Nitric oxide; Phenylephrine; Rabbit, arteries
1. Introduction
The cytotoxic
effects of oxgen-derived
free radicals
such as superoxide anion, hydroxyl radicals and hydrogen
peroxide (H202) have been widely investigated in relation
to their important roles in myocardial
[l-4] and coronary
vascular [5-lo]
reperfusion
injuries. We found that H,O,
reversively inhibits CY,-adrenergic agonist-induced
contractions of vascular smooth muscle, while it had no effect on
high-K+-induced
contractions
[I I]. These different effects
of H,O,
may result from the observation
that cr,-adren-
ergic agonists sensitize the contractile
apparatus to increase Ca*+ sensitivity whereas high-K+
does not. The
primary
aim of the present study was to elucidate the
mechanism of endothelium-independent
H,Oz-induced
relaxation of agonist-induced
contraction by simultaneously
examining
the changes in [Ca2+li
of vascular smooth
muscle and muscle tension. Hydrogen
peroxide
causes
relaxation
of rabbit aorta partly via endothelial
factors
[11,12], so we also aimed to confirm the endothelium-dependent factors responsible
for part of the H,O,-induced
relaxation.
* Corresponding author. Tel: ( + 8 I-3) 5802- 1056; Fax: ( + X1-3) 5689.
0627.
Time
0008.6363/96/$15.00
PII
SOOOS-6363(96)00022-3
0 1996 Elsevier
Science
B.V. All rights reserved
for primary
review 12 days
T. Irsuki
et al./Curdinousculur
2. Methods
2.1. Tissue preparation
tension
and measurement
of isometric
This investigation conformed to the Guide for the Care
and Use of Laboratory Animals published by the US
National Institutes of Health (NIH publication No. 85-23,
revised 1985) and was approved by our institutional animal experimentation committee.
Surgical procedures and measurement of isometric tension were the same as those described previously [l 11. In
brief, male Japanese white rabbits (weighing 2.8-3.6 kg)
were killed by administration of 25% urethane overdose
(lo-15
ml, intravenously),
and the descending thoracic
aorta was isolated. Surrounding fat and connective tissues
were removed, and transverse rings (2 mm wide) were cut
open to form strips. The endothelium of the strips was
denuded in some experiments by gently rubbing the intima1 surface with filter paper moistened with physiological
salt solution (PSS). The aortic strip was suspended vertically in a 5 ml glass chamber filled with PSS with the
following composition (mM): NaCl, 113.0; NaHCO,, 30.0;
MgSO,, 1.2; KH,PO,,
1.2; KCl, 4.7; CaCl,, 1.5; and
glucose, 5.5, oxygenated with a 95% 02/5% CO, gas
mixture. The pH was adjusted to 7.4, and its temperature
was maintained constant at 37°C. The strips were allowed
to equilibrate for 90-120 min under an initial preload of
1.75 g. After equilibration, the resting tension was reduced
to 1.5 g, and conditioning contractions were induced 3
times by applying 3M KC1 (final KC1 concentration of
64.7 mM) to the tissue chamber.
Either endothelium-denuded or endothelium-intact strips
were precontracted by high-K+ (20 mM) solution in which
NaCl in the PSS was replaced by an equimolar concentration of KCl. After stabilizing the tension at the high level,
the effects of H,O, (300 PM) on the precontracted strips
were examined. Phorbol 12,13-dibutyrate (PDBu, 30 nM),
a protein kinase C activator, which at this concentration is
reported to augment high-K+ contraction [ 131, was used to
study its effects on high-K+-induced
contraction and on
the subsequent administration of H,O,.
In another series of experiments, the effect of H,O, on
phenylephrine (PE, 0.3 PM)-precontracted
strips was examined using strips with intact endothelium. NG-monomethyl-L-arginine
(L-NMMA,
300 PM), a nitric oxide
(NO) synthase inhibitor, methylene blue (10 PM), a soluble guanylate cyclase inhibitor and tetraethylammonium
(TEA, 10 mM), a non-specific blocker of calcium-activated
potassium channels, were used in order to confirm the
endothelium-dependent
mechanism(s).
These concentrations were chosen because they have been previously used
in studies with isolated blood vessels (L-NMMA
[14],
methylene blue [15] and TEA [ 161). All were administered
15 min before application of PE.
The effects of pretreatment with these drugs were compared with those of control preparations excised from the
Research
31 (19961820-825
same animal. These experiments
time-matched manner.
821
were
2.2. Simultaneous measurement of [Ca’ ‘Ii
tension
performed
in a
and isometric
Vascular smooth muscle [Ca2+li and isometric tension
were measured simultaneously in strips which had been
mechanically denuded of endothelium [17] in order to
exclude the influence of the endothelium on [Ca2+ Ii values. Acetoxymethyl ester of fura- (fura-2-AM,
10 PM), a
fluorescent calcium indicator, together with 0.2% cremophor EL, a noncytotoxic detergent, was added to PSS
and sonicated. The strips were incubated with the fura-2AM-containing oxygenated PSS for 12-24 h in the dark at
20-25°C. After fura-2-AM loading, the strips were suspended horizontally in a temperature controlled (37°C) 10
ml tissue bath filled with normal (fura-2-free) PSS. One
end of the strip was connected to an isometric force
transducer to measure muscle tension and the other end to
a fixed hook. The strips were washed with normal PSS and
were equilibrated for 30 min under 1.5 g of resting tension.
A fluorometer (CAF 110, Japan Spectroscopic, Japan) with
a quartz-glass-bottomed
tissue bath was used. The strip
was illuminated with a xenon high-pressure (75W) lamp
alternating at 48 Hz with 340 and 380 nm of excitation
wavelength by means of a rotating filter wheel. The emitted fluorescence due to excitation at 340 nm (F,,,) and at
380 nm (F,,,) from the strip was measured through a 500
nm filter, and the ratio (F&F,,,)
was automatically
calculated. Only preparations in which F340 and F,,,
changed as a mirror image were used in the present study.
Changes in [Ca2+li were expressed as relative values;
[Ca2+li in the resting state was assigned O%, and those in
response to high K+ (64.7 mM), applied at the beginning
of each experiment, were taken as 100%. In strips precontracted by PE (0.3 PM) or high-K+ (64.7 mM), effects of
300 PM H,O, application on [Ca2+lj and muscle tension
were examined. In several experiments, H,O, was repeatedly administered under resting conditions both in normal
Ca2+-containing
PSS and in Ca*+-free solution. The
Ca2+-free solution was made by removing CaCl, from the
PSS and adding 0.5 mM EGTA.
2.3. Drugs
The following compounds were used; phenylephrine
hydrochloride,
hydrogen peroxide, and PDBu (Wako,
Japan); L-NMMA, TEA, and cremophor EL (Sigma, USA);
fura-2-AM and EGTA (Dojindo Laboratories, Japan). All
compounds, except PDBu, were dissolved in distilled water to make stock solutions and then diluted in the superfusing solution immediately before application. Phorbol
12,13-dibutyrate
was dissolved in dimethyl sulfoxide
(DMSO, Sigma, USA). The maximum concentration of
DMSO in the tissue chamber was 0.006%, which itself had
822
T. Iesaki er al./ Cardiovascular
no effect on the tension of the strips. Concentrations
all expressed as final concentrations.
were
Research
31 (1996)
820-825
A.
Cd+)
2.4. Statistical analysis
All values are expressed as mean(s.e.m.) and differences between them were analyzed using Student’s t-test
for unpaired samples.Differences at P < 0.05 were considered to be significant.
3. Results
p-w.
-
H,O,
HSOZ
I
_19
Sinill
6.
3.1. Effects of H202 on lCa2 ‘Ii and muscle tension in
high-K +- or PE-precontracted arteries
In endothelium-denudedstrips, tension development after application of either high-K+ (64.7 mM) or PE was
accompanied by an increase in [Ca2+li, expressed as a
percentage of the peak value measured during the first
high-K+ contraction. During high-K+-induced contraction
(Fig. 1, panel A, n = 4) administration of 300 PM H,O,
caused a further increase in [Ca2+Ji of 26.6(1.7)%, measured 10 min after application of H,O, , with a small
increase in tension of 6.5(1.9)% of high-K+-induced tension. During PE-induced contraction (Fig. 1, panel B,
n = 5) H,O, similarly caused an additional increase in
[Ca2+li of 26.4(4.7)%. However, in contrast to high-K+
contraction, H,O, decreasedmuscletension by 28.9(2.2)%,
expressedas a percentage of the steady-state PE-induced
tension.
Tension
Tension
-{
KCI
Fig. I. Effect of H,O,
on [Ca” 1, and muscle tension in high-K+or
phenylephrine-precontracted
arteries. The endothelium-denuded
strips
were precontracted
by either high-K+
(KCI, 64.7 mM, panel A) or
phenylephrine
(PE, 0.3 yM, panel B). Hydrogen
peroxide (300 PM) was
then administered.
Changes in [Ca’+ ], were expressed as F3.,,JF3s0
ratio. W = washout.
Cd-)
Tension
meem
T
--
Hz02
T!F
hii
1’S
Fig. 2. Effect of H,O,
on [Ca*+ 1, and muscle tension under resting
conditions.
Endothelium-denuded
strips were used. Hydrogen
peroxide
(300 FM) was administered
to the superfusing
solution under resting
conditions
in either Ca’+-containing
(1.5 mM, panel A) or Ca*+-free
(0
mM CaCl, plus 0.5 mM EGTA, panel B) solutions (black bars). After
washout of H,O,,
H,O, was administered
again. Changes in [Ca*+ I, are
expressed as the F340 /F,,,
ratio. W = washout.
3.2. Effects of H,O, on [Ca2 ‘I, and muscletension under
resting conditions
In normal PSS containing 1.5 mM Cazt (Fig. 2, panel
A, n = 5), administration of 300 PM H,O, to the endothelium-denudedmuscle for 15 min under resting conditions causeda gradual increasein [Ca*+], of 26.2(3.8)% of
the first high-K+-induced increasein [Ca”+];, with only a
very small increasein tension. In Ca2+-free solution (Fig.
2, panel B, n = 4) the H,O,-induced increase in [Ca2+li
was significantly (P < 0.05) depressed([Ca2+ji increased
by 10.5(2.8)%) and was further reduced by a second
challenge of H,O,, but the increase in [Ca2’li was still
evident. These changesof fluorescence induced by H,O,
were not due to its direct effect on fura-2, since 300 PM
H,O, did not affect the fluorescence of 100 PM fura-2containing solution over a pCa range of 8 to 6. This range
was chosen because [Ca2+Ii in vascular smooth muscle
cells hasbeen reported to range from several tens nanomolar to submicromolarlevels under physiological conditions
[18]. In addition, the pH of PSS was not altered by 300
PM H202.
3.3. Augmentation of high-K +-induced contraction by
PDBu and its relaxation by H202
The effects of PDBu pretreatment on high-K+-induced
contraction and on the action of subsequentapplication of
H,O, were examined in either the absenceor presenceof
T. Iesuki et al./Curdiouusculur
endothelium. In this series of experiments, the strips were
submaximally contracted with high-K+ (20 mM) at the
beginning of the experiments.
Fig. 3 shows typical experimental records using endothelium-denuded strips. Under control conditions (panel
A, n = S), a second challenge of high-K+
caused
107.2(1.6)% contraction of the first high-K+ contraction,
and subsequent administration of H,O, caused additional
contraction of l&4(2.6)% of the second high-K+ contraction. The effects of PDBu pretreatment are shown in panel
B (n = 8). Administration of 30 nM PDBu to the resting
muscle caused only a slight and gradual increase in tension. However, addition of PDBu significantly (P < 0.001)
augmented high-K+-induced
contraction to 165.1(5.2)%.
Subsequent H *O, administration caused relaxation; the
tension decreased by 21.9(1.1)%, expressed as a percentage of the augmented high-K+ contraction.
The same experiments were performed using strips with
intact endothelium (Fig. 4). In control conditions (panel A,
A,
Research
31 11996) 820-825
A.
823
control
v
H202
71
+
‘cv
i
KCI
t
KCI
t
KCI
t
KCI
t
control
W
Hz02 ‘f’
+
rd-----7
PDBu
t
KCI
t
KCI
B.
-1 L--l
t
KCI
Fig. 3. Effect of PDBu pretreatment
on high-K+-induced
contraction
and
the effect of subsequent administration
of H,O,
in the absence of
endothelium.
Endothelium-denuded
strips were first contracted by highK+ (KCl, 20 mM) followed
by washout with normal PSS. In control
conditions
(panel A), the snip was again contracted by high-K+
and
H,O, (300 PM) was administered.
In panel B, phorbol 12.13.dibutyrate
(PDBu, 30 nM) was applied to the superfusing solution for pretreatment
(black bar) after the first high-K+
contraction.
The strip was again
contracted by high-K+
in the presence of PDBu followed by H,O, (300
wM). W = washout.
I
‘3iGi”
Fig. 4. Effect of PDBu pretreatment
on high-K+-induced
contraction
and
the effect of subsequent
administration
of H,O,
in the presence of
endothelium.
Strips with intact endothelium
were first contracted
by
high-K+
(KC], 20 mM) followed
by washout with normal PSS. The
experimental
protocol was the same as shown in Fig. 3.
n = 7), a second challenge of high-K+ caused 107.4(1.4)%
contraction, and subsequent administration of H,O, caused
no significant change in tension. Pretreatment with PDBu
augmented high-K+-induced
contraction to 144.0(3.5)%,
and subsequent administration of H,O, caused relaxation
by 12.4(1.0)% (panel B, n= 6). Smaller changes in both
augmentation (P < 0.01) and relaxation (P < 0.001) in
strips with intact endothelium may reflect the action of
phorbol esters on endothelium [19].
3.4. Effects of L-NMMA,
ment on H,O,-mediated
arteries
methylene blue or TEA pretreatrelaxation of PE-precontracted
The effects of three different inhibitors on H,O,-mediated relaxation of PE-precontracted strips were examined in
the presence of intact endothelium. Relaxations in this
series of experiments were expressed as percentages of the
steady-state PE (0.3 PM)-induced
tension. Pretreatment
with 300 PM L-NMMA or 10 PM methylene blue significantly (P < 0.001 for both compounds) decreased the relaxation of PE-precontracted arteries induced by H,O,;
17.9(2.8)% and 43.2(4.4)% relaxation in the presence (n =
824
T. Iesuki
et al./Cardiouuscular
8) and absence (n = 6) of L-NMMA,
respectively, and
8.8( 1.3)% in the presence of methylene blue (n = 8) and
29.4(2.1)% in its absence (n = 4). Pretreatment with 10
mM TEA did not affect the relaxant effect of H,O, on
PE-precontracted strips: 34.9(2.9)% relaxation in its presence (n = 6) and 37.2(4.9)% in its absence (n = 6, NS).
4. Discussion
We found that H,O, depresses agonist-induced contraction though it resulted in an additional increase in [Ca2+li
of vascular smooth muscle. Hydrogen peroxide also counteracts the potentiating action of PDBu on high-K+-induced contraction.
We have previously reported that 300 PM H,O, causes
reversible relaxation of PE-induced contraction of vascular
smooth muscle through both endothelium-dependent and
-independent mechanisms, and that H,O, does not cause
relaxation of high-K+-induced contraction [ 11I. To investigate endothelium-independent mechanisms of H,O,-mediated relaxation, we measured [Ca*+], of vascular smooth
muscle
and muscle
tension
simultaneously
in
endothelium-denuded
rabbit aortic strips. Under resting
conditions, hydrogen peroxide increased [Ca2+li in both
Ca*+-containing
and Ca2+-free PSS. The increase was
smaller in the Ca’+-free PSS, and a second H,O, challenge in Ca2+-free PSS significantly depressed the increase
in [Ca2+li, indicating that Ca*+ release from internal
stores causes, at least in part, the increase in [Ca2+ Ii.
Hydrogen peroxide increases [Ca2+li in cultured vascular
smooth muscle cells [20,21], cultured endothelial cells
[22,23] and cultured renal tubular epithelial cells [24]. In
various tissues, the H,O,-induced
increase in [Ca’+],. ocinflux
from
extracellular fluid
curs not through Ca2’
[20,22,24], but through Cazf release from intracellular
stores [2 1,25,26]. Hydrogen peroxide also depresses Ca2+
uptake by canine cardiac SR [27].
The rise in [Ca2+li of vascular smooth muscle would be
expected to cause contraction. In the present study, Hz02
actually caused an additional increase in tension in endothelium-denuded, high-K+-precontracted
arteries (Figs.
1 and 3, panel A). High-K+-induced
contraction is generated both by Ca 2+ influx from extracellular space and by
Ca2+ release from internal stores without a significant
increase in Ca*+ sensitivity [2S]. On the other hand,
agonist-induced contraction is mediated by both increased
[Ca2+], and increased Ca2+ sensitivity probably resulting
from activation of protein kinase C‘ [28]. The fact that
hydrogen peroxide had a relaxant effect only on PE-induced contraction in spite of its increasing action on
[Ca*+]; indicates that Ca2+ sensitivity elevated in the
presence of PE is diminished by H,O,. We found that
high-K+-induced contraction was also depressed by H,O,
in the same manner as PE-induced contraction, when the
strips were pretreated with PDBu. Therefore, although the
Rrseurch
31 (1996)
520-825
exact mechanism of H,O,-mediated
suppression of Ca*+
sensitivity is not known, the present results indicate that
H,O, inhibits, at least partly, the protein kinase C-dependent mechanism of increase in Ca*+ sensitivity of vascular
smooth muscle, since phorbol esters are well-known
activators of protein kinase C [29].
In a previous study, we showed that endothelium-derived prostaglandins, hydroxyl radicals, which can be generated from H,O,, and lipid peroxidation of cell membranes do not make a major contribution to H,O,-mediated relaxation [ 111. Recently, hydrogen peroxide-induced
NO release has been reported in cultured bovine aortic
endothelial cells [23], in cultured human umblical vein
endothelial cells [30] and in the rabbit aorta[l2]. The
present results obtained by L-NMMA
or methylene blue
confirmed the findings that endothelium-derived NO contributes to the H,O,-mediated
relaxation [12]. Increase in
[Ca*‘],
induced by H,O,
in endothelial cells [22,23]
developed similarly with vascular smooth muscle cells
could result in stimulation of endothelial NO release [23].
The ineffectiveness of H,O, in reducing high-KS‘-induced tension suggests that endothelium-derived hyperpolarizing factor (EDHF) may contribute to H,O,-mediated
relaxation, since EDHF would be unable to hyperpolarize
sarcolemma which is strongly depolarized by high-K+.
One possible mechanism of EDHF-induced hyperpolarization of smooth muscle cell membrane is opening of calcium-activated potassium channels [3 1,321. However, TEA,
a non-selective blocker of calcium-activated
potassium
channels [16], failed to affect the H,O,-mediated
relaxation of PE-induced contraction, suggesting that H,O,
does not stimulate EDHF release. Therefore, NO is the
primary relaxant messenger released from the endothelium
in response to H,O,.
In the present study, H,O, caused no significant change
in tension of high-K+-precontracted
strips in the presence
of endothelium (Fig. 4, panel A), whereas H,O, caused an
additional increase in tension in the absence of endothelium (Fig. 3, panel A). In the presence of endothelium, the
contractile effect of H,O, due to a [Ca2+li increase in the
muscle could be cancelled by the relaxant effect of NO
released from the endothelium, although the NO release
could be smaller under conditions of high-K+-induced
membrane depolarization [33].
In conclusion, the net effect of H,O, is determined by
the balance of the following three effects: increase in
[Ca2+]; of vascular smooth muscle cells, suppression of
agonist-induced Ca2+ -sensitization of the contractile apparatus, and NO release from endothelial cells.
References
[l]
Bolli R, Jeroudi MO, Pate1 BS, et al. Direct evidence that oxygenderived free radicals contribute to postischemic
myocardial
dyslimction in the intact dog. Proc Nat1 Acad Sci USA 1989;86:4695-4699.
T. Iesuki
rt cd./
Currlioonsculur
[2] Hess ML, Manson NH. Molecular
oxygen: friend and foe. The role
of the oxygen free radical system in the calcium paradox, the oxygen
paradox
and ischemia/reperfusion
injury.
J Mol Cell Cardiol
1984; 16:969-985.
[3] McCord
JM. Oxygen-derived
free radicals in postischemic
tissue
injury. N Engl J Med 1985;3 12: l59- 163.
[4] Zweier JL. Measurement
of superoxide-derived
free radicals in the
reperfused heart. J Biol Chem 1988;263: l353- 1357.
[5] Hearse DJ, Maxwell
L, Saldanha C, Gavin JB. The myocardial
vasculature during ischemia and reperfusion:
a target for injury and
protection.
J Mol Cell Cardiol 1993;25:759~800.
[6] Jackson CV, Mickelson
JK, Pope TK, Rao PS, Lucchesi BR. 0,
free radical-mediated
myocardial
and vascular dysfunction.
Am J
Physiol 1986;25l:Hl225-H1231.
[7] Ku DD. Coronary
vascular reactivity
after acute myocardial
ischemia. Science 1982;218:576-578.
[8] VanBenthuysen
KM, McMurtry
IF, Horwitz
LD. Reperfusion
after
acute coronary
occlusion in dogs impairs endothelium-dependent
relaxation
to acetylcholine
and augments contractile
reactivity
in
vitro. J Clin Invest 1987;79:265-274,
[9] Mehta JL, Nichols WW, Donnelly WH, Lawson DL, Saldeen TGP.
Impaired canine coronary vasodilator
response to acetylcholine
and
bradykinin
after occlusion-reperfusion.
Circ Res 1989;64:43-54.
[ 101 Rubanyi GM, Vanhoutte PM. Oxygen-derived
free radicals, endothelium, and responsiveness
of vascular smooth muscle. Am J Physiol
1986;250:H815-H821.
[I l] Iesaki T, Okada T, Yamaguchi
H. Ochi R. Inhibition
of vasoactive
amine induced contractions
of vascular smooth muscle by hydrogen
peroxide in rabbit aorta. Cardiovasc Res 1994;28:963-968.
[l2] Zembowicz
A, Hatchett RJ, Jakubowski
AM, Gryglewski
RJ. Involvement
of nitric oxide in the endothelium-dependent
relaxation
induced by hydrogen
peroxide in the rabbit aorta. Br J Pharmacol
1993;110:151-158.
[ 131 Shima H, Blaustein
MP. Contrasting
effects of phorbol esters on
serotoninand vasopressin-evoked
contractions
in rat aorta and
small mesenteric artery. Circ Res 1992;70:978-990.
[14] Rees DD, Palmer RMJ, Schulz R, Hodson HF, Moncada S. Characterization of three inhibitors of endothelial
nitric oxide synthase in
vitro and in viva. Br J Pharmacol 199O;101:746-752.
[I51 Gruetter
CA, Gruetter
DY, Lyon JE, Kadowitz
PJ, Ignarro LJ.
Relationship
between cyclic guanosine 3’:5’-monophosphate
formation and relaxation
of coronary arterial smooth muscle by glyceryl
ttinitrate, nitroprusside,
nitrite and nitric oxide: effects of methylene
blue and methemoglobin.
J Pharmacol Exp Ther 1981;219:181-186.
[I61 Hutcheson IR, Griffith TM. Heterogeneous
populations of K+ chatnels mediate EDRF release to flow but not agonists in rabbit aorta.
Am J Physiol 1994;266:H590-H596.
[I71 Sato K, Ozaki H. Karaki H. Changes in cytosolic calcium level in
vascular smooth muscle strip measured simultaneously
with contraction using fluorescent
calcium indicator fura 2. J Pharmacol
Exp
Ther 1988;246:294-300.
[18] Somlyo AP, Himpens B. Cell calcium and its regulation in smooth
muscle. FASEB J 1989;3:2266-2276.
Heseurch
31 (1996)
820-825
825
[ 191 Stasek JE Jr, Patterson CE, Garcia JGN. Protein kinase C phosphorylates caldesmon and vimentin and enhances albumin permeability
across cultured bovine pulmonary
artery endothelial cell monolayers.
J Cell Physiol 1992; 153:62-75.
r20] Schachter M, Gallagher
KL, Pate] MK, Sever PS. Oxidants
and
vascular
smooth
muscle.
Biochem
Sot Trans 1989;17: 10961097(Abstract).
[21] Roveri
A, Coassin M, Maiorino
M, et al. Effect of hydrogen
peroxide
on calcium homeostasis
in smooth muscle cells. Arch
B&hem
Biophys 1992;297:265-270.
[22] Kimura M, Maeda K, Hayashi S. Cytosolic
calcium increase in
coronary
endothelial
cells after H,Oz
exposure and the inhibitory
effect of U785 l7F. Br J Pharmacol 1992; 107:488-493.
1231 Shimizu S, Saitoh Y, Yamamoto
T, Momose K. Stimulation
by
hydrogen peroxide of L-aginine
metabolism
to L-citrulline
coupled
with nitric oxide synthesis in cultured endothelial
cells. Res Commun Chem Path01 Pharmacol
1994;84:3 15-329.
[24] Ueda N, Shah SV. Role of intracellular
calcium in hydrogen peroxide-induced
renal tubular cell injury. Am J Physiol 1992;263:F214
F221.
[25] Schwartz-Sorensen
S, Christensen
F, Clausen T. The relationship
between the transport of glucose and cations across cell membranes
in isolated tissues. X. Effect of glucose transport stimuli on the
efflux of isotopically
labelled calcium and 3-o-methylglucose
from
soleus muscles and epididymal
fat pads of the rat. Biochim Biophys
Acta 1980;602:433-445.
[26] Boraso A, Williams
AJ. Modification
of the gating of the cardiac
sarcoplasmic
reticulum
Ca*+-release
channel by H,O,
and dithiothreitol. Am J Physiol 1994;267:HlOlO-HlOl6.
[27] Rowe GT, Manson NH, Caplan M, Hess ML. Hydrogen
peroxide
and hydroxyl
radical mediation of activated leukocyte depression of
cardiac sarcoplasmic
reticulum. Participation
of the cyclooxygenase
pathway. Circ Res 1983;53:584-591.
[28] Somlyo AP, Somlyo AV. Signal transduction
and regulation
in
smooth muscle. Nature 1994;372:23 l-236.
[29] Andrea JE, Walsh MP. Protein kinase C of smooth muscle. Hypertension 1992;20:585-595.
[30] Madge LA, Baydoun
AR. HaO, modulates basal and histaminestimulated nitric oxide release in human umbilical vein endothelial
cells: stimulation
of NO synthase by H,O,
is inhibited by L-NAME
and removal of extracellular
calcium. J Physiol (Land) 1994;475.
P:65P-66P(Abstract).
[3 I] Garland CJ, Plane F, Kemp BK, Cocks TM. Endothelium-dependent
hyperpolarization:
a role in the control of vascular tone. Trends
Pharmacol Sci 1995; 16:23-30.
[32] Hecker M, Bara AT. Bauersachs J, Busse R. Characterization
of
endothelium-derived
hyperpolarizing
factor as a cytochrome
P450derived arachidonic
acid metabolite
in mammals. J Physiol (Land)
1994;481:407-414.
[33] Lickhoff
A, Busse R. Calcium influx into endothelial
cells and
formation of endothelium-derived
relaxing factor is controlled by the
membrane potential. Pflligers Arch 1990;416:305-3
1 I.