Circulation 1990 Bolli 723 38

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ElResearch Advances Series

Mechanism of Myocardial "Stunning"
Roberto Bolli, MD
P ostischemic ventricular dysfunction, or myocardial "stunning,"' has recently become the
focus of considerable attention among
researchers and clinicians. There are two fundamental reasons for the explosive growth of interest in this
phenomenon. First, it is now recognized that spontaneous reperfusion after coronary spasm or thrombosis is common in patients with coronary artery disease. This implies that postischemic myocardial
stunning is likely to be an important component of
the natural history of this disorder. Second, with the
advent of interventional recanalization, an increasingly large number of patients are subjected to coronary reperfusion in an effort to preserve left ventricular function. The occurrence of myocardial
stunning in these patients could significantly delay
the benefits of reperfusion therapy.
During the past decade, the efforts of experimental
investigators have produced considerable progress in
our understanding of the mechanism responsible for
postischemic contractile dysfunction. This is clearly
the central problem because it will not be possible to
effectively prevent myocardial stunning unless its
pathogenesis is elucidated. The purpose of this article is to critically review the various mechanisms
proposed for postischemic dysfunction and their
pathophysiological and clinical implications. The
major unresolved issues and areas for future research
will be identified, and an attempt will be made to
integrate different theories into a unifying pathogenetic paradigm.
Definition of Myocardial Stunning
One cannot overemphasize the importance of a
clear definition of myocardial stunning because this
term is sometimes loosely applied to situations in
which the persistence of contractile abnormalities
in postischemic tissue is due to other causes (e.g.,
myocellular death). Postischemic dysfunction, or
myocardial stunning, is the mechanical dysfunction
that persists after reperfusion despite the absence of
From the Section of Cardiology, Department of Medicine,
Baylor College of Medicine, Houston, Tex.
Supported in part by National Institutes of Health (NIH) grants
HL-43151 and HL-36277 and NIH SCOR grant HL-42267. R.B.
was a recipient of the Physician-Scientist Award, American College of Chest Physicians.
Address for correspondence: Roberto Bolli, MD, Section of
Cardiology, Baylor College of Medicine, 6535 Fannin, MS F-905,
Houston, TX 77030.
Received February 7, 1990; revision accepted May 8, 1990.
irreversible damage. The essential point of this definition is that postischemic dysfunction, no matter
how severe or prolonged, is a fully reversible abnormality. Diagnosis of myocardial stunning should not
be made unless reasonable assurance can be provided that the tissue in question is still entirely
viable.
In accordance with this definition, myocardial
stunning is a relatively mild, sublethal injury that must
be kept quite distinct from myocardial infarction. It is
unknown whether these two conditions share a common mechanism. Consequently, this review will discuss only results obtained in experimental preparations in which mechanical dysfunction is likely to be
due to stunning rather than to cell death. The
mechanism of lethal ischemia-reperfusion injury has
been reviewed previously2-7 and is beyond the scope
of this article.
Classification of Myocardial Stunning
Experimentally, the concept of myocardial stunning encompasses a wide variety of settings with
major pathophysiological differences. A classification
is in order because different mechanisms have been
described in different experimental conditions. The
available observations can be grouped into the fol-
lowing categories (Table 1).

Myocardial Stunning After One Completely Reversible
Ischemic Episode
In the dog, a coronary occlusion lasting less than 20
minutes does not result in any myocardial necrosis2,8
but nevertheless produces prolonged abnormalities
of contractile performance.8-14 This is the classic
model of myocardial stunning, the one in which the
phenomenon was originally described,9 and the one
most commonly used in experimental investigations.
Myocardial Stunning After Multiple Completely

Reversible Ischemic Episodes
Repeated brief (5 or 10 minutes) coronary occlusions have a cumulative effect on systolic function
and result in prolonged contractile impairment
despite absence of irreversible damage.15-19 The largest decrease in contractility occurs after the first
occlusion; with subsequent occlusions, the additional
decrements in mechanical performance become progressively smaller.20 (Whether repetitive ischemia
also has a cumulative effect on the time to complete
recovery, however, remains unclear.) This model of
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Circulation Vol 82, No 3, September 1990
TABLE 1. Classification of Myocardial Stunning and Evidence for Various Mechanisms Proposed in Experimental Animals
Evidence for pathogenetic role of:
Reduced
Sarcoplasmic
Damage of
reticulum
Calcium
calcium
Oxygen
collagen
radicals
overload
Experimental setting
dysfunction
matrix
sensitivity
Stunning due to decreased blood flow
Regional ischemia
One completely reversible ischemic
++
?
episode
Multiple completely reversible
+
+
?
ischemic episodes
+
One partly irreversible ischemic
?
?
?
episode (subendocardial infarction)
Global ischemia
+
+
+
Isolated heart in vitro
?
+
?
?
?
?
Cardioplegic arrest in vivo
Stunning due to increased 02 demands
Exercise-induced ischemia
?
+, Published studies support this mechanism; + +, published studies from multiple laboratories consistently support this mechanism, and
evidence is also available in conscious animal preparations; -, published studies do not support this mechanism; , published studies are
conflicting; ?, no data are available.
myocardial stunning differs from the single 10- or

15-minute occlusion model in that the mechanical
dysfunction develops gradually and is associated with
a considerably greater total ischemic burden (50-60
versus 10-15 minutes).
Myocardial Stunning After One Partly Irreversible
Ischemic Episode (Subendocardial Infarction)
In the dog, when reperfusion is instituted after a
period of coronary occlusion of more than 20 minutes
but less than 3 hours, the subendocardial portion of
the region at risk is generally found to be infarcted,
whereas variable quantities of subepicardial tissue
remain viable.2 The recovery of function in this
subepicardial region salvaged by reperfusion is, however, delayed for days or weeks.8,21-25 It is particularly
difficult to evaluate the effect of therapy on this form
of postischemic dysfunction because the reperfused
region contains a complex admixture of necrotic
subendocardium and stunned subepicardium, and
the relative proportions of these two components are
highly variable. Other confounding factors include
the tethering of surviving myocytes by dead, nonfunctional tissue; the expansion of the infarcted region;
and the progressive replacement of necrotic myocardium by scar.
Myocardial Stunning After Global Ischemia in
Isolated Hearts
The occurrence of cell death in these preparations
depends on species, temperature, duration of ischemia, and perfusate composition. Under selected
conditions, isolated hearts reperfused after transient
ischemia exhibit complete normalization of phosphocreatine content and intracellular pH,26-34 indicating that viability is generally preserved. Although
in these models the reversibility of the contractile
abnormalities cannot be verified, the metabolic
recovery suggests that the injury is mostly nonlethal.

Accordingly, despite the numerous obvious differences from ischemia in vivo, under selected circumstances isolated heart preparations can mimic myocardial stunning. In other cases, however, these
preparations may be associated with significant cell
death,35-44 and their relevance to myocardial stunning is questionable.
Myocardial Stunning After Global Ischemia During

Cardioplegic Arrest In Vivo
Despite the use of hypothermic cardioplegia, global
ischemia in intact animals is usually followed by prolonged contractile abnormalities.45-48 The reversibility
of these derangements has not been documented, but
under carefully controlled conditions they are likely to
be due mostly to stunning.
Myocardial Stunning After Exercise-Induced Ischemia
Stresses such as exercise provoke myocardial ischemia and dysfunction in animals with a flow-limiting
coronary stenosis. With cessation of exercise, these
contractile abnormalities persist.49 Thus, myocardial
stunning can also occur after high-flow ischemia, in
which the primary problem is an increase in oxygen
requirements rather than a decrease in supply.
Because of the many significant pathophysiological
differences among these situations, one cannot
assume that observations made in one setting necessarily apply to the others. An important, unresolved
issue is whether all forms of stunning share a common pathogenesis.
Factors That Determine the Severity of
Myocardial Stunning
In conscious dogs undergoing a 15-minute coronary occlusion, there is a sensitive coupling between
the degree of myocardial dysfunction after reperfu-
Bolli Mechanism of Stunned Myocardium

TABLE 2. Mechanisms Proposed for Myocardial Stunning
Most likely mechanisms

Generation of oxygen-derived free radicals
Excitation-contraction uncoupling due to sarcoplasmic
reticulum dysfunction
Calcium overload
Other proposed mechanisms
Insufficient energy production by mitochondria
Impaired energy use by myofibrils
Impairment of sympathetic neural responsiveness
Impairment of myocardial perfusion
Damage of the extracellular collagen matrix
Decreased sensitivity of myofilaments to calcium
sion and the magnitude of blood flow reduction

during the preceding period of ischemia, whereby
even small differences in ischemic perfusion are
associated with large differences in postischemic
recovery.12 (In open-chest dogs, this correlation is
weaker,50 probably because of the confounding
effects of anesthesia and trauma.) Furthermore, the
severity of stunning is greater in the inner layers of
the left ventricular wall, which are the most severely
ischemic, than in the outer layers.'4'51 Another important factor is the duration of flow deprivation:
The longer the ischemic period, the greater the
ensuing mechanical abnormalities.9'52
In conclusion, the severity of postischemic dysfunction is determined primarily by the severity and
duration of the antecedent ischemia. This concept
has two important implications. First, whatever may
be the precise mechanism responsible for stunning,
such mechanism must be initiated and modulated by
perturbations associated with ischemia. Although
stunning appears to be a form of "reperfusion injury"
(see below), it is ischemia that "primes" the myocardium for the development of such injury. Second, any
intervention that improves perfusion during ischemia
would be expected to attenuate stunning after reflow.
Reducing the severity of ischemia is probably the most
effective way to reduce postischemic dysfunction.
725
thesize enough high-energy phosphates to sustain

contractile function, possibly because of loss of adenine nucleotide precursors. Numerous subsequent
observations, however, have refuted this theory.
First, no correlation is observed between myocardial
ATP levels and recovery of contractility in several
models of postischemic dysfunction.26'27,56-59 Second,
the content of phosphocreatine in the stunned myocardium is normal or supranormal ("phosphocreatine overshoot"),8'26'27'3053'55'57'60 implying that the
phosphorylation ability of the mitochondria is intact.
Third, the stunned myocardium responds to inotropic stimuli with a marked and sustained increase in
contractility, such that function exceeds the levels
measured at baseline.17,30,61-66 This striking inotropic
response occurs without a decrease in ATP or phosphocreatine stores,30,63 indicating that energy production is not inherently impaired and can keep pace
with high metabolic demands. Fourth, manipulations
that increase ATP levels in the stunned myocardium
fail to increase mechanical function.33,67
In summary, the available evidence suggests that a
defect in the rate of resynthesis of ATP by the
mitochondria is not the primary cause of myocardial
stunning.
Impaired Energy Use by Myofibrils

Myocardial stunning is associated with a decrease
in myofibrillar creatine kinase activity68 and a reduction of the substrate (free ADP) used by the myofibrillar creatine kinase to produce ATP at the contraction site.30,68 Both of these abnormalities might
Insufficient Energy Production by Mitochondria

ATP levels in the stunned myocardium are
depressed and recover slowly,8,2453-55 with a time
disrupt normal energy use and thus have been postulated to contribute to postischemic dysfunction.68
This hypothesis, however, appears implausible
because it does not explain the considerable contractile and metabolic reserve of the stunned myocardium.17'30'61-66 Indeed, the reversal of postischemic dysfunction by inotropic stimuli implies that the
residual activity of myofibrillar creatine kinase is still
sufficient to run the myofibrillar ATPase reaction at
normal or supranormal rates.
Impairment of Sympathetic Neural Responsiveness
After 25 minutes of coronary occlusion, the reperfused myocardium loses its inotropic responsiveness
to sympathetic nervous stimulation, suggesting injury
to the sympathetic-neural axis.69 However, no such
defect is observed after 15 minutes of coronary
occlusion.70 Furthermore, as noted above, myocardial stunning occurs in isolated (and thereby denervated) hearts. Although functional sympathetic denervation may contribute to postischemic dysfunction
in certain situations in which contractility is strongly
dependent on sympathetic drive, it seems unlikely
that this abnormality plays a major role in most
experimental preparations of myocardial stunning.
course similar to that of contractile function.8'24

Thus, in the early 1980s, the hypothesis was
advanced",54'55 that postischemic dysfunction may
result from an inability of the myocardium to resyn-
Impairment of Myocardial Perfusion

When myocardial stunning is produced by a 15minute coronary occlusion, postreperfusion subepi-
Mechanism of Myocardial Stunning

Thus, in very general terms, the abnormalities of the
postischemic myocardium are governed by the abnormalities occurring during ischemia. But what is the
specific sequence of events whereby transient ischemia leads to prolonged depression of contractility?
Several different hypotheses, which are not necessarily exclusive, have been proposed (Table 2) and
are reviewed below.
726
cardial blood flow is normal, whereas subendocardial

flow is slightly decreased (-20%)."-14,71-73 The
mechanism and significance of this phenomenon are
unclear.72,73 Although it is possible, as speculated by
Heyndrickx et al,1' that the reduced subendocardial
perfusion may contribute to the impaired mechanical
performance, two considerations suggest that it is not
a major causative factor. First, there is no correlation
between subendocardial blood flow and wall thickening in the postischemic myocardium.72,73 Second, the
loss of systolic function (dyskinesis or akinesis) is out
of proportion to the slight (-20%) decrease in
subendocardial perfusion.72'73
When myocardial stunning is produced by
repeated 5-minute coronary occlusions, blood flow to
the postischemic myocardium, measured by radioactive microspheres, is generally normal.15-17,19 However, microspectrophotometry reveals increased veinto-vein heterogeneity of oxygen saturations, with an
excess of vessels demonstrating very low oxygen
saturations, suggesting that the stunned myocardium
contains discrete microregions of either increased
oxygen extraction or reduced flow.19 A nonuniform
reduction of flow (microregions of ischemia intermixed with microregions of hyperemia) has also been
invoked to explain the enhancement of function that
is observed in stunned myocardium when perfusion is
augmented.16 However, the findings that myocardium
stunned by repeated 5-minute coronary occlusions
has normal flow reserve'6 and intact microvascular
patency'9 exclude nonuniform ischemia secondary to
anatomical obstruction. Furthermore, the hypothesis
of persistent ischemia as a cause of dysfunction does
not explain the normal or near-normal contractile
reserve of the postischemic myocardium.17'65,66
In summary, impaired perfusion (either uniform or
nonuniform) is unlikely to be the primary pathogenetic mechanism in postischemic contractile failure.
Damage of Extracellular Collagen Matrix
Using a model of myocardial stunning produced by
12 5-minute coronary occlusions separated by 10
minutes of reperfusion, Eng's group has demonstrated that postischemic dysfunction is associated
with severe ultrastructural damage of the extracellular collagen matrix74 and a modest (-10%) loss of
collagen.75 These findings have led to the hypothesis
that myocardial stunning may be due to a structural
defect, namely, disruption of the mechanical coupling function provided by the extracellular collagen
network.74 To fully evaluate this hypothesis, it will be
important to explore the time course of the collagen
changes and its relation to the time course of postischemic dysfunction. Because in conscious dogs
undergoing one brief (c15 minutes) coronary occlusion most of the contractile recovery takes place in
the first 3-4 hours of reperfusion9'12,13,52 (an interval
too short for significant resynthesis or remodeling of
the collagenous components), it seems improbable
that structural damage to collagen plays a pathogenetic role in this model of myocardial stunning.
Indeed, a recent study76 demonstrated that collagen

is disrupted by 12 5-minute coronary occlusions but
not by one 15-minute occlusion followed by reperfusion.
In summary, collagen damage may contribute to
myocardial stunning after multiple ischemic episodes
but is unlikely to be a causative factor after a single
completely reversible ischemic episode (Table 1).
One possible explanation for this difference is that in
the former setting, repetitive ischemia results in
repetitive systolic bulging for a total duration of more
than 15 minutes; this may cause greater mechanical
stress and, consequently, greater damage of the
extracellular collagen network.
Decreased Sensitivity of Myofilaments to Calcium

Using isolated ferret hearts subjected to 15 minutes of global ischemia at 370 C, Kusuoka et a127
observed that the stunned myocardium exhibits
decreased responsiveness to calcium, as manifested
by a decrease in the maximal calcium-activated force
and a decrease in the myocardial sensitivity to extracellular calcium. They speculated that the reduced
sensitivity to extracellular calcium could in turn be
due to either a decrease in the intracellular free Ca21
concentration ([Ca]i) transient or a decrease in the
sensitivity of myofilaments to calcium.27 The former
possibility seems improbable because two recent
studies with different techniques32'77 indicate that the
calcium transient is (paradoxically) increased in the
stunned myocardium after 1532 or 20 minutes77 of
global ischemia at 370 C in isolated ferret hearts.
Accordingly, it has been proposed32'77 that the fundamental mechanism for postischemic dysfunction is
not insufficient availability of free cytosolic calcium
during systole but instead is reduced sensitivity of the
contractile apparatus to calcium. One problem with
this hypothesis derived from in vitro studies is that it
does not explain two observations made in vivo. First,
when the stunned myocardium is challenged with
inotropic stimuli, it exhibits a normal or near-normal
contractile reserve.1765,66 Second, the apparent sensitivity of the stunned myocardium to intracoronary
calcium is not decreased.65 If the primary problem
was a reduced sensitivity of myofilaments to calcium,
then the contractile response to exogenous calcium
or to other inotropic agents (which act by raising
[Ca]i) should be reduced.
In summary, studies in vitro suggest that myocardial stunning is a result of reduced calcium sensitivity
rather than reduced calcium availability, but studies
in vivo are not readily reconcilable with this interpretation (Table 1).
Calcium Overload
Several lines of evidence point to a pathogenetic
role of calcium overload in myocardial stunning.
First, when isolated ferret hearts subjected to 15
minutes of global normothermic ischemia are reperfused with solutions containing low concentrations of
calcium, the postischemic contractile abnormalities
are significantly attenuated.27 The results of this

experiment, in which no intervention is applied during ischemia, indicate that calcium entry upon reperfusion is an important mechanism of myocardial
stunning. A decrease in the severity of stunning is
also observed in hearts pretreated with ryanodine, an
inhibitor of cellular calcium overload.29 Second,
exposure of isolated ferret hearts to a transient
calcium overload in the absence of ischemia produces
mechanical and metabolic abnormalities similar to
myocardial stunning.78 Third, transient intracellular
acidosis during early reperfusion can prevent myocardial stunning in isolated ferret hearts subjected to
15 minutes of global ischemia31 and open-chest dogs
undergoing 15 minutes of coronary occlusion.79
Because acidosis antagonizes not only the influx of
calcium into cells (by inhibiting the Na4-Ca21
exchange and the slow calcium channels) but also the
intracellular binding of calcium,31 these results provide indirect evidence that a transient calcium overload contributes to postischemic dysfunction. Fourth,
recent studies28'80'81 have shown that [Ca]i increases
between 10 and 20 minutes of global ischemia in
isolated hearts. This is the same duration of ischemia
that results in myocardial stunning. In these models,
[Ca]i returns to normal values within a few minutes
after reperfusion,28,7780 but appears to remain transiently elevated during very early reflow.77'82
The mechanism for this rise in [Ca]i during ischemia remains uncertain. It may be secondary to
decreased calcium uptake by the sarcoplasmic
reticulum.83 It may also be mediated through the
Na+-Ca2' exchange, secondary to a rise in intracellular Na+ during ischemia due to 1) metabolic inhibition of the Na+,K+-ATPase, and 2) acidosis and
consequent Na+-H+ exchange.84-86 Because the Na+Ca2' exchange is inhibited by acidosis and intracellular pH in the stunned myocardium normalizes
quickly after reflow 27,30'31'33'60 entry of calcium via
the Na4-Ca2' exchange may be greatly increased
upon reperfusion.84,86-88 Further research will be
necessary to identify the precise circumstances leading to calcium overload. The mechanisms by which a
transient calcium overload induces prolonged contractile dysfunction are also unclear. Increased cytosolic calcium can activate phospholipases and other
degradative enzymes.6,34 In addition, elevation of
cytosolic calcium could trigger production of oxygen
radicals via xanthine oxidase89 (see below).
The calcium overload theory is not incompatible
with the fact that exogenous calcium ameliorates
function in the stunned myocardium.27,65 The increase in [Ca]i is postulated to be a brief phenomenon immediately after reflow, following which there
may be either a normalization of [Ca]i transients
or a relative calcium deficiency (see "ExcitationContraction Uncoupling Due to Sarcoplasmic Reticulum Dysfunction"). The early excessive [Ca]i levels
could damage intracellular organelles concerned
with contraction and thereby produce prolonged
mechanical dysfunction.
727
In summary, considerable evidence suggests that a

transient calcium overload during early reperfusion
contributes to the pathogenesis of myocardial stunning after global ischemia in vitro. The role of
calcium overload in in vivo models of myocardial
stunning remains to be defined (Table 1) and represents an important area for future investigations.
Effect of calcium channel blockers. Calcium channel blockers have been used to gain insights into the
role of altered calcium homeostasis in vivo, but
unfortunately the results are difficult to interpret.
Verapamil,59 diltiazem,90 nifedipine,91 nitrendipine,92 and amlodipine93 have been shown to
improve recovery of function in regionally stunned
myocardium in intact animals. However, it is
unclear whether these beneficial effects reflected a
direct protective action of the drugs or were mediated by favorable modifications of afterload, preload, heart rate, and regional myocardial blood
flow, all of which could change the systolic properties of the stunned myocardium.16,59 A direct protective action was demonstrated by a recent study94
in which nifedipine enhanced the functional recovery of the stunned myocardium independently of
any effect on systemic hemodynamics or regional
myocardial blood flow. However, interpretation of
these results is problematic because the beneficial
effects of nifedipine were noted even when treatment was started 30 minutes after reperfusion,94
whereas myocardial stunning has been suggested to
result from a transient calcium overload immediately after reflow.27 It is unlikely that nifedipine
acted in a specific way to decrease [Ca]i because
administration of calcium or inotropic agents
(which raise [Ca]i) improves contractile function in
the stunned myocardium.17,30,61-66 The mechanism
of action of nifedipine in this intriguing study94
remains to be elucidated.
Excitation-Contraction Uncoupling Due to

Sarcoplasmic Reticulum Dysfunction
Krause et a195 investigated this mechanism in a
canine model of postischemic dysfunction produced
by eight to 12 5-minute occlusions separated by
10-minute reflow periods. Sarcoplasmic reticulum
isolated from the stunned myocardium demonstrated
a decrease in the ability to transport calcium, concomitant with a reduction in the activity of the
Ca2+,Mg2+-ATPase.95 These researchers proposed
that a decrease in the amount of calcium stored in
the sarcoplasmic reticulum as a result of a reduction
in the calcium pump activity could diminish contractile protein activation through attenuated calcium
release during systole.95
The concept of inadequate delivery of calcium to
the contractile proteins, secondary to sarcoplasmic
reticulum dysfunction, is an attractive hypothesis. The
postulated relative calcium deficiency would be consistent with the observation that exogenous administration of calcium can return contractile function of
stunned myocardium to preischemic levels.65 It would
728
also be consistent with the notion that inotropic agents

(which increase [Ca]i) can reverse myocardial
stunning.17'30'61-66 However, since this hypothesis
implies that the amplitude of the [Ca]i transient is
decreased, it is not easily reconcilable with in vitro
data32'77 (see above). Considerably more research is
necessary to determine whether similar dysfunction of
the sarcoplasmic reticulum occurs in other models of
myocardial stunning (Table 1) and whether it is associated with decreased [Ca]i transients in vivo. In
addition, the factors that cause injury to the sarcoplasmic reticulum remain to be elucidated.
Generation of Oxygen-Derived Free Radicals
Effect of antioxidants on myocardial stunning after a
brief coronary occlusion. Approximately 7 years ago, a
number of investigators, including ourselves, postulated that myocardial stunning is caused in part by
the generation of reactive oxygen metabolites, such
as superoxide anion (02 ), hydrogen peroxide (H202),
and hydroxyl radical (.OH), and began a series of
experiments designed to test this hypothesis. We used
an open-chest canine preparation in which the left
anterior descending coronary artery is occluded for 15
minutes and then reperfused.96 In the first study,97 we
found that administration of superoxide dismutase
(SOD) (an enzyme that catalyzes the dismutation of
*02 to 02 and H202) and catalase (an enzyme that
reduces H202 to 02 and H20) significantly enhanced
recovery of function after reperfusion. Attenuation of
myocardial stunning by SOD and catalase was also
observed by other investigators50'57'98 using similar
experimental preparations. In a more recent study99
we found that neither SOD nor catalase alone significantly improved recovery of function in the stunned
myocardium; however, when the two enzymes were
combined, contractile recovery was significantly greater
than that observed in controls or in dogs receiving
either enzyme alone. These results suggest that both
*02- and H202 contribute to the cellular damage
responsible for myocardial stunning and that combined
administration of SOD and catalase is more likely to be
effective against stunning than separate administration.
The inability of SOD alone to mitigate postischemic
dysfunction has also been observed in a pig model.100
Although both *2- and H202 appear to contribute to stunning, it remains uncertain whether they
do so by direct cytotoxicity or via formation of other
species. *2 and H202 can interact through the
metal-catalyzed Haber-Weiss reaction to generate
the highly reactive *OH radical.10' Consequently,
accumulation of *2 and H202 could produce
postischemic dysfunction, at least in part, indirectly
through generation of OH. To elucidate this problem, we evaluated dimethylthiourea, an -OH scavenger that is more effective than traditional *OH
scavengers102 and does not react with *2 or H202
in vitro.'03 We observed that dimethylthiourea produced a significant and sustained improvement in
the function of the stunned myocardium.103 These

results were further corroborated by studies with
N-2-mercaptopropionyl glycine (MPG), a free radical scavenger that readily enters the intracellular
space and is active orally. MPG was found to be a
powerful scavenger of OH with no effect on *2 or
H202 in vitro104 and to effectively attenuate myocardial stunning in vivo.104105 Taken together, these
results suggest that the OH radical (or one of its
reactive products) is a mediator of postischemic
dysfunction and that the beneficial effects of SOD
and catalase previously demonstrated5057'97-99 are
due in part to prevention of OH generation. This
conclusion is also consistent with the finding that
the iron chelator desferrioxamine attenuates postischemic dysfunction.106"07 Because iron catalyzes
the formation of OH (through the Haber-Weiss or
Fenton mechanism) as well as the propagation of
.OH-initiated lipid peroxidation,10' the protective
effects of desferrioxamine are compatible with the
view that OH is a mediator of myocardial stunning.
The effectiveness of antioxidants that scavenge
.OH or prevent its generation should not be construed as evidence that all of the damage initiated by
oxygen is mediated via OH. It is true that H202 is a
relatively nonreactive species and its toxicity may be
due in large part to its reduction to OH.101 However,
there is evidence that *2 can directly cause cellular
toxicity in vitro (Reference 108; reviewed in Reference 109). Furthermore, because catalase alone prevents formation of OH by removing H202, the fact
that SOD has to be added to catalase to produce
significant attenuation of stunning in vivo99 implies
the existence of a component of damage specifically
due to *02- It appears, therefore, that all three
species (2- , H202, and *OH) are important in
postischemic dysfunction (O2- and *OH as mediators
of injury and H202 as a precursor of OH).99
Although the studies discussed above50,57,97-99,103-107

consistently support the oxyradical hypothesis, their
significance is limited by the fact that they were all
performed in open-chest animals. Thus, artifacts due
to the combined effects of anesthesia, hypothermia,
surgical trauma, volume and ionic imbalances, nonphysiological conditions, and associated neurohumoral perturbations, as well as other potentially
confounding variables, cannot be excluded. It is
therefore essential that the oxyradical hypothesis be
tested in conscious animal preparations. In a recent
experiment,"10 we observed that the combined
administration of SOD and catalase significantly
enhanced recovery of function after a 15-minute
coronary occlusion in conscious, unsedated dogs and
that this effect was sustained for 24 hours after
reflow. The accelerated recovery was not followed by
any subsequent deterioration, indicating that postischemic depression of contractility is not a useful
"protective" response to injury.
In summary, numerous investigations from several
independent laboratories5057,97- 99,103-107, 10 uniformly
suggest that oxygen metabolites play a significant role
729
in the genesis of myocardial stunning after a 15minute period of ischemia, in both open-chest and
conscious animals. The relative contributions of the
various metabolites have not been definitively established. It is clear that a portion of the damage is
mediated by OH; however, the available evidence
suggests that myocardial stunning cannot be simplistically ascribed to a single oxygen species and that all
of the three initial metabolites of oxygen (02 , H202
and *OH) contribute to the cellular injury responsible
for postischemic dysfunction. Iron also appears to
play a role in stunning, presumably by catalyzing the
formation of OH.
Direct evidence for the oxyradical hypothesis. A
major limitation of the studies reviewed heretofore50,57,97-99,103-107,110 is that all of the evidence provided by these investigations is indirect and therefore
inconclusive. Clearly, to definitively confirm a causative role of oxygen metabolites in postischemic
dysfunction, it is necessary to directly demonstrate
and quantitate free radical generation in the stunned
myocardium in the presence and absence of antioxidant interventions.
Production of free radicals has been directly
demonstrated in isolated rabbit or rat hearts undergoing global ischemia and reperfusion,42,4311-113
but because of the numerous important differences
in experimental conditions, results obtained in vitro
cannot necessarily be extrapolated to the intact
animal. We used the spin trap a-phenyl N-tert-butyl
nitrone (PBN) and electron paramagnetic resonance (EPR) spectroscopy to detect and measure
production of free radicals in our in vivo model of
postischemic dysfunction (15-minute coronary
occlusion in open-chest dogs).14 After infusion of
PBN, EPR signals characteristic of PBN radical
adducts were detected in the venous blood draining
from the ischemic-reperfused vascular bed.15 The
EPR signals were consistent with a mixture of
different secondary lipid radicals, such as alkyl and
alkoxy radicals. The myocardial production of radicals began during coronary occlusion but increased
dramatically after reperfusion, peaking at 2-4 minutes. After this initial burst, production of radicals
abated but did not cease, persisting for as long as 3
hours after reflow.1" There was a linear, positive
relation between the magnitude of adduct production and the magnitude of ischemic flow reduction,
indicating that the intensity of free radical generation after reflow is determined by the severity of the
antecedent ischemia"15-the greater the degree of
hypoperfusion, the greater the subsequent production of free radicals and, by inference, the severity
of reperfusion injury. These findings imply that
interventions that improve perfusion during ischemia will attenuate free radical reactions after
cals are derived from univalent reduction of oxygen.

Furthermore, the inhibition of free radical production was associated with inhibition of myocardial
stunning.17 More recently, we observed that MPG or
desferrioxamine administered just before reperfusion
markedly attenuated myocardial stunning and the
associated production of PBN adducts04,114; however, the same agents given 1 minute after reperfusion did not attenuate myocardial stunning or initial
PBN adduct production. Thus, three different antioxidant interventions (SOD plus catalase, MPG, and
desferrioxamine) reduced postischemic dysfunction
at the same doses and under the same experimental
conditions in which they reduced formation of PBN
adducts. The correlation between the two effects
suggests that the production of free radicals in the
stunned myocardium plays a causal role in the
depression of contractility.
In summary, the results of the studies that have
measured free radicals in the stunned myocardium104114-117 provide direct evidence supporting a
pathogenetic role of oxygen metabolites. Specifically,
these studies indicate that 1) free radicals are produced in the stunned myocardium in the intact dog
after reversible regional ischemia, 2) the univalent
pathway of reduction of oxygen is the source of the
radicals, and 3) inhibition of free radical reactions
results in enhanced recovery of contractility (i.e., the
radical reactions are necessary for postischemic dysfunction to occur).
Effect of oxygen radicals on cardiac function. The
ability of oxygen metabolites to depress myocardial
function has been directly demonstrated in vitro and
in vivo. Exposure of isolated rabbit interventricular
septa,18 isolated rat"19 or rabbit'08 papillary muscles,
and isolated rat120-122 or rabbit'23"124 hearts to free
radical-generating solutions or pure H202 has uniformly resulted in decreased mechanical function
and ATP levels (i.e., in changes similar to those
observed in the stunned myocardium). In most of the
studies in which an attempt was made to discern the
relative roles of different oxygen species,118,119,122'123
it was found that the deleterious effects could be
prevented by catalase or by *OH scavengers but not
by SOD, suggesting that H202 or its byproduct, *OH,
was the oxygen metabolite responsible for the
observed depression of contractile function. In one
study,108 however, SOD was protective, whereas catalase had no effect, implying that the major negative
inotropic agent was *02 . These in vitro observations
have been recently expanded by Przyklenk et al,'25
who reported that infusion of xanthine oxidase plus
purine plus iron-loaded transferrin, administered
through a coronary vein in open-chest dogs, resulted
in the development of significant wall motion
reflow. Generation of free radicals in in vivo models

of regional myocardial stunning has also been
reported by Leiboff et al.1"6
In a subsequent study,17 we found that SOD plus
catalase suppressed the production of free radicals in
the stunned myocardium, indicating that these radi-
abnormalities.
In summary, it is clear that reactive oxygen species
depress myocardial contractility both in vitro and in
vivo. The precise role of each species remains to be
defined. There is considerable evidence that the
detrimental effects of *2- and H202 on cardiac
730
function are mediated in part by generation of *OH.

This is in accordance with the results of in vivo
studies,103-'07 which suggest an important role of *OH
as a mediator of stunning. However, there is also
evidence for a direct negative inotropic action of
*02 , which is also congruent with observations in
vivo.99
Mechanism of oxyradical-mediated contractile dysfunction. The precise mechanism whereby oxygen
metabolites depress contractile function remains
speculative and represents one of the major unresolved issues pertaining to the pathogenesis of myocardial stunning. Free radicals are reactive species
that have no specific target and can attack virtually all
cellular components. In theory, every abnormality
thus far described in the stunned myocardium (see
above) could be caused by oxyradicals. At least two
key cellular components, proteins and lipids, are
likely to be involved in free radical-initiated reactions in the postischemic myocardium. Activated
oxygen species can denaturate proteins and inactivate enzymes.126 Furthermore, they can produce peroxidation of the polyunsaturated fatty acids contained in cellular membranes,127 which would impair
selective membrane permeability and interfere with
the function of various cellular organelles.
Evidence for the occurrence of lipid peroxidation
in the stunned myocardium is provided by two recent
studies. Romaschin et al128 observed increased myocardial concentration of hydroxy conjugated dienes
(which are products of free fatty acid oxidation)
during and after a 45-minute period of global normothermic ischemia in open-chest dogs. The maximal concentration of conjugated dienes was measured at 5 minutes of reflow. Importantly, the tissue
examined after reperfusion was dysfunctional but not
necrotic, thus representing stunned myocardium.
Weisel et al129 subsequently reported that in patients
undergoing cardioplegic arrest during coronary
artery bypass surgery, there was myocardial release
of conjugated dienes in the coronary sinus blood at 3
and 60 minutes of reperfusion, which was associated
with a decrease in the myocardial concentration of
the antioxidant a-tocopherol.
Dysfunction of the sarcoplasmic reticulum may be
an important mechanism whereby oxyradicals mediate the contractile abnormalities observed after
reversible ischemia. As mentioned above, both calcium uptake and Ca2+,Mg2+-ATPase activity have
been found to be significantly depressed in the
stunned myocardium.95 A similar decrease in calcium
uptake and Ca2',Mg2+-ATPase activity is observed in
isolated sarcoplasmic reticulum after exposure to
oxygen radicals,127,130 and oxyradical scavengers preserve sarcoplasmic reticulum function.'27,130
Other studies suggest that the sarcolemma may
be a critical target of free radical-mediated damage. Oxyradicals have been shown to interfere with
calcium transport and calcium-stimulated ATPase
activity in the sarcolemma.'31132 Oxygen radicals
have also been shown to interfere with the Na+-
Ca 2+ exchange133 and to inhibit the Na+,K+-ATPase

activity.134 Furthermore, the sarcolemmal Na+,K+ATPase activity is impaired in reperfused myocardium, and this impairment is prevented by
antioxidants.135 Impairment of the Na+,K+-ATPase
activity would result in Na+ overload, with consequent activation of the Na+-Ca2+ exchange activity.87'88 All of these observations imply that excessive production of oxyradicals could result in
increased transarcolemmal calcium influx and cellular calcium overload.
It is important to point out that the foregoing
postulated mechanisms involve alterations in calcium
homeostasis and thus would reconcile the oxyradical
and calcium-overload hypotheses of stunning into
one pathogenetic mechanism.
Sources of oxygen radicals in the stunned myocardium. Among the various potential mechanisms of
oxyradical generation in the stunned myocardium,
only two have been explored thus far, namely, the
enzyme xanthine oxidase and the activated neutrophil. We found that the xanthine oxidase inhibitor,
allopurinol, produced a marked improvement in the
functional recovery of the stunned myocardium.136
Furthermore, allopurinol inhibited the increased cardiac production of urate observed in control dogs
during ischemia and early reperfusion, indicating
effective inhibition of xanthine oxidoreductase.136
Attenuation of stunning has also been demonstrated
with oxypurinol after 15137 and 90 minutes138 of
ischemia and reperfusion. These data suggest that
xanthine oxidase is one of the sources of the oxygen
radicals that contribute to postischemic dysfunction
in the dog. Whether this concept applies to humans
remains uncertain because the myocardial content of
xanthine oxidase is species dependent. Reports
regarding the myocardial content of the enzyme in
the human heart are conflicting.139-143
In contrast to these earlier reports,136-138 in a
recent study,'44 oxypurinol and amflutizole, two xanthine oxidase inhibitors, failed to mitigate stunning
after 15 minutes of ischemia in dogs. Whether the
discrepancy is due to different degrees of inhibition
of xanthine oxidoreductase or to other reasons is
unclear.
Leukocytes are another potential source of oxygen
metabolites.145,146 However, several investigators
have found that myocardial stunning is not attenuated by depletion of neutrophils with antiserum71'147
or filtration,148 by administration of nafazatrom,
which inhibits production of leukotrienes,149 by
administration of antibodies to the adhesionpromoting Mol glycoprotein,150 and by administration of dextran, which inhibits leukocyte adherence
to the endothelium.151 Recent studies have shown
that the neutrophil content of the stunned myocardium after a 12-minute period of ischemia is
decreased152 and that myeloperoxidase activity (a
marker of neutrophils) in myocardium stunned by a
15-minute occlusion is not different from nonischemic myocardium.150 Furthermore, myocardial stun-
ning occurs in isolated heart preparations that are

devoid of circulating neutrophils. Taken together,
these results71'147-'52 suggest that granulocytes do not
play a major role in the genesis of postischemic
dysfunction after brief, reversible ischemia, although
they appear to be major mediators of irreversible
tissue injury and vascular obstruction after prolonged
periods of ischemia.145"146153
In contrast to the aforementioned findings, however,
others have reported that depletion of granulocytes
by Leukopak filtration prevents154 or alleviates155
postischemic dysfunction after a 15-minute coronary
occlusion. The reasons for the differences between
these studies'54"155 and the aforementioned
investigations71"147-151 are unclear and warrant additional research.156 One possibility to be explored,
raised by Engler and Covell,154 is that the relation
between circulating neutrophil counts and myocardial damage may be nonlinear.
In summary, in the canine model of myocardial
stunning produced by one reversible ischemic episode, xanthine oxidase may be a source of free
radicals (although the data are not entirely consistent), whereas the role of neutrophils remains controversial. Of course, many other processes could
lead to formation of 2- and H202 during myocardial ischemia and reperfusion, including activation of
the arachidonate cascade, autoxidation of catecholamines and other compounds, ischemia-induced
damage of the electron transport chain in the mitochondria, and accumulation of reducing equivalents.127 It will be difficult to selectively assess the
role of these potentially important mechanisms in a
complex system such as the intact animal. Identification of the sources of oxyradicals in the stunned
myocardium represents another major unresolved
issue.
Time course of free radical-induced damage in the
stunned myocardium. When exactly in the ischemiareperfusion sequence that leads to stunning does the
crucial free radical-induced damage take place? In a
recent study,104 we observed that infusion of the
antioxidant MPG attenuated postischemic dysfunction to a similar extent regardless of whether the
infusion was started before ischemia or 1 minute
before reperfusion; however, infusion started 1
minute after reflow was ineffective, suggesting that
the critical radical-mediated injury occurs in the first
few moments of reperfusion. We have subsequently
obtained similar results with desferrioxamine.14 Furthermore, the spin trap PBN enhances contractile
recovery even when the infusion is commenced 20
seconds before reflow; the magnitude of the protective effect is similar to that observed when the
infusion is commenced before ischemia.1" Captopril,
an angiotensin converting enzyme inhibitor with
alleged *2 scavenging properties, produces similar
attenuation of postischemic dysfunction when the
administration is started before ischemia or 2 minutes before reperfusion.157 The conclusion that a
substantial portion of the cellular damage responsi-
731
ble for stunning occurs immediately after reflow is

further corroborated by direct measurements of free
radicals104"'15"17 and lipid peroxidation products128 in
the stunned myocardium, both of which have shown
a burst in the initial moments after restoration of
flow. If free radical production is inhibited during
this initial burst, postischemic dysfunction is mitigated; however, if free radical production is inhibited
after the first 5 minutes of reperfusion (i.e., after the
initial burst), no functional improvement is
observed.04 These observations suggest that the free
radicals important in causing myocardial stunning
are those produced immediately after reflow.
In summary, myocardial stunning appears to be, at
least in part, a form of oxyradical-mediated "reperfusion injury." This concept may have significant
therapeutic implications because it suggests that
antioxidant therapies begun after the onset of ischemia could still be effective in preventing postischemic dysfunction; however, a delay in the implementation of such therapies until after reperfusion would
result in loss of efficacy.
Role of oxygen radicals in other forms of myocardial
stunning (Table 1). Studies reviewed thus far used one
brief (.15 minutes) coronary occlusion. In recent
unpublished studies in our laboratory, we found that
the *OH scavenger MPG markedly improves contractile recovery after 10 5-minute coronary occlusions
separated by 10-minute reflow periods in open-chest
dogs. Thus, oxyradicals also appear to contribute to
the genesis of myocardial stunning after multiple
brief ischemic episodes. Furthermore, numerous
studies from the surgical literature suggest that oxygen radicals also contribute to postischemic dysfunction after global ischemia in in vivo models of cardioplegic arrest,45-48 and there is evidence that
oxidative stress contributes to postoperative dysfunction in patients undergoing cardiac surgery.158
Finally, antioxidants consistently alleviate mechanical dysfunction after global ischemia in isolated
hearts,35-4144 but the relevance of these in vitro
preparations to myocardial stunning is uncertain, as
discussed above.
Data regarding the role of oxygen radicals in
myocardial stunning after a prolonged coronary
occlusion are conflicting. Three studies failed to
detect improvement in functional recovery with SOD
and catalase after coronary occlusions lasting 1 hour,'59
90 minutes,160 or 2 hours16' in open-chest'6' or conscious dogs.'59 160Wel62 also observed that SOD fails
to enhance recovery of contractility after a 2-hour
coronary occlusion in anesthetized dogs. These
results suggest that short-term administration of
antioxidant enzymes is not effective in mitigating
myocardial stunning associated with subendocardial
infarction. It may be that the postischemic dysfunction that follows a prolonged, partly irreversible ischemic insult differs from the dysfunction
that follows a short, completely reversible insult.
However, other studies138"63 have shown that the
cell-permeant antioxidants oxypurinol and N-
732
acetylcysteine attenuate myocardial stunning independently of infarct size limitation in closed-chest

dogs subjected to 90 minutes of coronary occlusion
and 24 hours of reflow.
Initial observations164 indicate that SOD and catalase do not alleviate exercise-induced stunning.
In summary, there is strong evidence that oxyradicals contribute to postischemic dysfunction after
global ischemia (in vitro as well as in vivo) and after
multiple episodes of regional ischemia. They do not
appear to contribute to exercise-induced postischemic dysfunction. The role of oxygen radicals in
myocardial stunning after a prolonged, partly irreversible ischemic insult remains uncertain and represents a major unresolved problem. Elucidation of this
issue will be difficult because the dysfunction is due
in part to the presence of infarction and in part to the
presence of stunning a situation that complicates
the evaluation of therapy, as discussed above.
-
Integration of Different Hypotheses

Myocardial stunning is probably a multifactorial
process that involves complex sequences of cellular
perturbations and the interaction of multiple pathogenetic mechanisms. Our understanding of this phenomenon is still fragmentary, and none of the theories discussed herein (Table 2) explains the entire
cascade of events that culminates in postischemic
contractile failure. For example, the oxyradical
hypothesis does not explain how reactive oxygen
species cause mechanical dysfunction, and the sarcoplasmic reticulum theory does not explain why this
organelle is damaged. Integration of the various
hypotheses is complicated by the fact that for the
most part, each hypothesis has been developed in a
different experimental preparation (Table 1).
Nevertheless, it is important to emphasize that
these hypotheses are not mutually exclusive and in
fact may represent different parts of the same pathophysiological sequence. There is considerable evidence to suggest a link between generation of oxygen
radicals and impaired calcium homeostasis in the
setting of reperfusion injury. For example, the damage associated with the "calcium paradox" resembles
that associated with the "oxygen paradox" and probably has a similar pathogenetic mechanism.165 Furthermore, as discussed above, oxyradicals generated
upon reperfusion can cause dysfunction of the sarcoplasmic reticulum127,130 and alter calcium flux across
the sarcolemma.131-135 The result of these actions
would be excitation-contraction uncoupling and cellular calcium overload.127131 In this regard, it has
recently been demonstrated that reoxygenation of
cultured myocytes results in Ca'+ overload, which can
be greatly attenuated by antioxidant enzymes.85 Oxygen radicals could also damage the proteins of the
contractile machinery in a manner that reduces their
responsiveness to calcium. On the other hand, calcium overload may exaggerate oxyradical production
by promoting the conversion of xanthine dehydroge-
nase to xanthine oxidase, which appears to be mediated by a calcium-dependent protease.89

A unifying hypothesis for the pathogenesis of
myocardial stunning is proposed in Figure 1 (detailed
description of the postulated mechanisms is provided
in the figure legend). This paradigm is largely speculative but nevertheless based on the evidence available at this time and discussed in this review. According to this conceptual scheme (Figure 1), oxyradical
generation, calcium overload, and sarcoplasmic reticulum dysfunction can be viewed as different aspects
of the same pathogenetic mechanism, thereby reconciling the three major current hypotheses of myocardial stunning.
In summary, the pathogenesis of myocardial stunning has not been definitively determined. Thus far,
researchers have focused on the identification of
individual mechanisms of injury. Three abnormalities
(oxyradical generation, calcium overload, and excitation-contraction uncoupling) have emerged as likely
contributing factors (although in different experimental preparations), and in vitro evidence suggests
that they are interrelated. It is now essential to clarify
the precise interactions among these three factors in
vivo and the role that each of them plays in the
various experimental settings of myocardial stunning.
Clinical Implications
There are numerous clinical settings in which the
myocardium is subjected to transient ischemia,
including unstable or variant angina, acute myocardial infarction with early reperfusion (either spontaneous or induced by thrombolytic therapy), openheart surgery with cardioplegic arrest, and cardiac
transplantation. Recent observations suggest that
these situations may be associated with myocardial
stunning, which may be an important factor precipitating left ventricular failure with its attendant morbidity and mortality.1,166 Furthermore, in patients in
whom silent episodes of ischemia recur at short
intervals in the same territory, the myocardium might
remain reversibly depressed for prolonged periods of
time or even chronically.1 Elucidation of the mechanism responsible for postischemic dysfunction is
indispensable for developing effective clinical therapies. The evidence discussed herein implies that
myocardial stunning could be effectively attenuated
by pharmacological manipulations targeted at specific pathogenetic mechanisms, for example, by
antioxidant agents given before reperfusion. Compared with standard treatments for cardiac failure
such as inotropic stimulation, antioxidants represent
a novel approach; by interfering with the mechanism
of cell injury, they might prevent rather than simply
temporarily correct postischemic dysfunction. This
approach also appears desirable because it is uncertain whether prolonged inotropic stimulation of
stunned myocardium has deleterious effects.
733
REVERSIBLE ISCHEMIA / REPERFUSION
Ja+
IN
rload
over
,
? t Reducing equivalents
Xanthine oxidase ?
* *-? tMitochondrial "univalent leak"
eutrophil activation ?
"
? Autoxidative processes
Arachidonate cascade ? -SOD
_
*0 2
H2 2
I
FeFI1
*OH
Enzyme inactivation
.u
KhhO
Lipid peroxidation
Damage of:|
/"
<eofjj t
? SARCOPLASMIC
*****K
? OTHER S TRUCTURES
or COLLAGEN
MATRIX)
? SARCOLEMMA
RETICULUM
(e.g., CONTRACTIIILE
PROTEINSS
'Ca+" overload
+ f+
f
Excitation-contraction uncoupling-
I Sensitivity
to Ca++
Altered Cat+ homeostasis
..~
L.IMECHANICAL DYSFUNCTION
Loss of
mechanical
coupling
m
FIGURE 1. Proposed pathogenesis of postischemic myocardial dysfunction. This proposal integrates and reconciles different
mechanisms into a unifyingpathogenetic hypothesis. Transient reversible ischemia followed by reperfusion could result in increased
production of superoxide radicals (02 ) through several mechanisms, including 1) increased activity of xanthine oxidase, 2)
activation of neutrophils, 3) activation of arachidonate cascade, 4) accumulation ofreducing equivalents during oxygen deprivation,
5) derangements of intramitochondrial electron transport system resulting in increased univalent reduction of oxygen, and 6)
autoxidation of catecholamines and other substances. Superoxide dismutase (SOD) dismutates '02- to hydrogen peroxide (H202);
in the presence of catalytic iron, '02- and 11202 interreact in a Haber- Weiss reaction to generate the hydroxyl radical (.OH). H202
can also generate *OH in the absence of '02 through a Fenton reaction provided that other substances (such as ascorbate) reduce
Fe (III) to Fe (II). '02- and OH attack proteins and polyunsaturated fatty acids, causing enzyme inactivation and lipid
peroxidation, respectively. In reversible ischemia, the intensity of this damage is not sufficient to cause cell death but is sufficient to
produce dysfunction ofkey cellular organelles. Postulated targets offree radical damage include 1) the sarcolemma, with consequent
loss of selective permeability, impairment of calcium-stimulated ATPase activity and calcium transport out of the cell, and
impairnent of Na+K+-ATPase activity (the net result of these perturbations would be increased transarcolemmal calcium influx
and cellular calcium overload); 2) the sarcoplasmic reticulum, with consequent impairment of calcium-stimulated A TPase activity
and calcium transport (this would result in impaired calcium homeostasis- specifically, decreased calcium sequestration, which
would contribute to increase free cytosolic calcium, and decreased calcium release during systole, which would cause
excitation-contraction uncoupling); and 3) possibly other structures, such as the extracellular collagen matrix (with consequent loss
of mechanical coupling) or the contractile proteins (with consequent decreased sensitivity to calcium). At the same time, reversible
ischemia-reperfusion could cause cellular Na+ overload due to 1) inhibition of sarcolemmal Na+,K+-ATPase and 2) acidosis and
Na+-H+ exchange. This could further exaggerate calcium overload by increased Na +-Ca2' exchange. An increase in free cytosolic
calcium would activate phospholipases and other degradative enzymes and further exacerbate the injury to the aforementioned key
subcellular structures (sarcolemma, sarcoplasmic reticulum, and contractile proteins). Thus, calcium overload could serve to
amplify the damage initiated by oxygen radicals. In addition, calcium overload could in itself impair contractile performance and
contribute to mechanical dysfunction. It is also possible that the increase in free cytosolic calcium could increase oxyradical
production bypromoting conversion ofxanthine dehydrogenase to xanthine oxidase. The ultimate consequence of this complex series
ofperturbations is a reversible depression of contractility.
Summary
Among the numerous mechanisms proposed for
myocardial stunning, three appear to be more plausible: 1) generation of oxygen radicals, 2) calcium
overload, and 3) excitation-contraction uncoupling.
First, the evidence for a pathogenetic role of
oxygen-derived free radicals in myocardial stunning
is overwhelming. In the setting of a single 15-minute
coronary occlusion, mitigation of stunning by antioxidants has been reproducibly observed by several
independent laboratories. Similar protection has
been recently demonstrated in the conscious animal,
that is, in the most physiological experimental preparation available. Furthermore, generation of free
radicals in the stunned myocardium has been directly
demonstrated by spin trapping techniques, and atten-
734
uation of free radical generation has been repeatedly

shown to result in attenuation of contractile dysfunction. Numerous observations suggest that oxyradicals
also contribute to stunning in other settings: after
global ischemia in vitro, after global ischemia during
cardioplegic arrest in vivo, and after multiple brief
episodes of regional ischemia in vivo. Compelling
evidence indicates that the critical free radical damage occurs in the initial moments of reflow, so that
myocardial stunning can be viewed as a sublethal
form of oxyradical-mediated "reperfusion injury."
Second, there is also considerable evidence that a
transient calcium overload during early reperfusion
contributes to postischemic dysfunction in vitro; however, the importance of this mechanism in vivo
remains to be defined.
Third, inadequate release of calcium by the sarcoplasmic reticulum, with consequent excitationcontraction uncoupling, may occur after multiple
brief episodes of regional ischemia, but its role in
other forms of postischemic dysfunction has not been
explored.
It is probable that multiple mechanisms contribute
to the pathogenesis of myocardial stunning. The
three hypotheses outlined above are not mutually
exclusive and in fact may represent different steps of
the same pathophysiological cascade. Thus, generation of oxyradicals may cause sarcoplasmic reticulum
dysfunction, and both of these processes may lead to
calcium overload, which in turn could exacerbate the
damage initiated by oxygen species. The concepts
discussed in this review should provide not only a
conceptual framework for further investigation of the
pathophysiology of reversible ischemia-reperfusion
injury but also a rationale for developing clinically
applicable interventions designed to prevent postischemic ventricular dysfunction.
Acknowledgment
The excellent secretarial assistance of William
LaCour is gratefully acknowledged.
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R Bolli
Circulation. 1990;82:723-738
doi: 10.1161/01.CIR.82.3.723
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Circulation 1990 Bolli 723 38

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Circulation 1990 Bolli 723 38

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723

ElResearch Advances Series

lowing categories (Table 1).

Myocardial Stunning After Multiple Completely

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Circulation Vol 82, No 3, September 1990

myocardial stunning differs from the single 10- or

recovery suggests that the injury is mostly nonlethal.

Myocardial Stunning After Global Ischemia During

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Bolli Mechanism of Stunned Myocardium

Most likely mechanisms

sion and the magnitude of blood flow reduction

thesize enough high-energy phosphates to sustain

Impaired Energy Use by Myofibrils

Insufficient Energy Production by Mitochondria

course similar to that of contractile function.8'24

Impairment of Myocardial Perfusion

Mechanism of Myocardial Stunning

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Circulation Vol 82, No 3, September 1990

cardial blood flow is normal, whereas subendocardial

Indeed, a recent study76 demonstrated that collagen

Decreased Sensitivity of Myofilaments to Calcium

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Bolli Mechanism of Stunned Myocardium

are significantly attenuated.27 The results of this

In summary, considerable evidence suggests that a

Excitation-Contraction Uncoupling Due to

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Circulation Vol 82, No 3, September 1990

also be consistent with the notion that inotropic agents

the function of the stunned myocardium.103 These

Although the studies discussed above50,57,97-99,103-107

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Bolli Mechanism of Stunned Myocardium

cals are derived from univalent reduction of oxygen.

reflow. Generation of free radicals in in vivo models

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Circulation Vol 82, No 3, September 1990

function are mediated in part by generation of *OH.

Ca 2+ exchange133 and to inhibit the Na+,K+-ATPase

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Bolli Mechanism of Stunned Myocardium

ning occurs in isolated heart preparations that are

ble for stunning occurs immediately after reflow is

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Circulation Vol 82, No 3, September 1990

acetylcysteine attenuate myocardial stunning independently of infarct size limitation in closed-chest

Integration of Different Hypotheses

nase to xanthine oxidase, which appears to be mediated by a calcium-dependent protease.89

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Bolli Mechanism of Stunned Myocardium

REVERSIBLE ISCHEMIA / REPERFUSION

Altered Cat+ homeostasis

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Circulation Vol 82, No 3, September 1990

uation of free radical generation has been repeatedly

ischemic myocardium: Effects of reperfusion of arterial

size, myocardial function, biochemistry, ultrastructure and

return of function. JAm Coll Cardiol 1985;6:864-870

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Bolli Mechanism of Stunned Myocardium

reperfusion injury by oxygen free radical scavengers. J Thorac