Effects of endurance training and acute exhaustive exercise on antioxidant
defense mechanisms in rat heart
Mustafa Gula,⁎, Berna Demircanb, Seyithan Taysie, Nuray Oztasanf, Kenan Gumustekina,
Erdinc Siktarc, M. Fevzi Polatd, Sedat Akara, Fatih Akcayb, Senol Danea
aDepartment of Physiology, Faculty of Medicine, Ataturk University, 25240, Erzurum, Turkey
bDepartment of Biochemistry, Faculty of Medicine, Ataturk University, Erzurum, Turkey
cCollege of Physical Education and Sports, Ataturk University, Erzurum, Turkey
dBiotechnology Application and Research Center, Ataturk University, Erzurum, Turkey
eDepartment of Biochemistry, Nenehatun Obstetrics and Gynecology Hospital, Erzurum, Turkey
fDepartment of Physiology, Faculty of Medicine, Afyon Kocatepe University, Afyon, Turkey
Received 17 July 2005; received in revised form 28 November 2005; accepted 30 November 2005
We investigated whether 8-week treadmill training strengthens antioxidant enzymes and decreases lipid peroxidation in rat heart. The effects of
acute exhaustive exercise were also investigated. Male rats (Rattus norvegicus, Sprague-Dawley strain) were divided into trained and untrained
groups. Both groups were further divided equally into two groups where the rats were studied at rest and immediately after exhaustive exercise.
Endurance training consisted of treadmill running 1.5 h day−1, 5 days week−1for 8 weeks. For acute exhaustive exercise, graded treadmill running
was conducted. Malondialdehyde level in heart tissue was not affected by acute exhaustive exercise in untrained and trained rats. The activities of
glutathione peroxidase and glutathione reductase enzymes decreased by both acute exercise and training. Glutathione S-transferase and catalase
activities were not affected. Total and non-enzymatic superoxide scavenger activities were not affected either. Superoxide dismutase activity
decreased by acute exercise in untrained rats; however, this decrease was not observed in trained rats. Our results suggested that rat heart has
sufficient antioxidant enzyme capacity to cope with exercise-induced oxidative stress, and adaptive changes in antioxidant enzymes due to
endurance training are limited.
© 2005 Elsevier Inc. All rights reserved.
Keywords: Heart; Acute exhaustive exercise; Endurance training; Malondialdehyde; Superoxide dismutase; Glutathione peroxidase; Glutathione reductase;
Glutathione S-transferase; Catalase; Superoxide scavenger activity
Cells have developed different antioxidant systems and
various antioxidant enzymes to defend themselves against free
radical attacks. Superoxide dismutase (SOD), the first line of
defense against oxygen-derived free radicals, catalyses the
dismutation of the superoxide anion into hydrogen peroxide.
H2O2can be transformed into H2O and O2by catalase (CAT),
which is present in peroxisomes of eukaryotic cells (Michiels et
al., 1994). Glutathione-dependent antioxidant system consisting
of reduced glutathione (GSH) and an array of functionally
related enzymes plays a fundamental role in cellular defense
against reactive free radicals and other oxidant species (Sen,
1997; Gul et al., 2000). Of these enzymes, glutathione
peroxidase (GPx) is a selenoprotein that reduces hydroper-
oxides as well as H2O2while oxidizing glutathione. A number
of potentially toxic electrophilic xenobiotics (such as certain
carcinogens, bromobenzene, chlorobenzene) are conjugated to
the nucleophilic glutathione by glutathione S-transferases
(GSTs) present in high amounts in cell cytosol. GST can also
catalyze reactions reducing peroxides like GPx. Reduction of
oxidized glutathione (GSSG) to GSH is mediated by the widely
distributed enzyme GSSG reductase (GRD) that uses NADPH
as the reductant (Sen, 1997; Gul et al., 2000).
Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245
⁎Corresponding author. Tel.: +90 442 2361212/2010; fax: +90 442 2360968.
E-mail address: firstname.lastname@example.org (M. Gul).
1095-6433/$ - see front matter © 2005 Elsevier Inc. All rights reserved.
Exhausting (Khanna et al., 1999; Gul et al., 2003; Oztasan et
al., 2004) or moderate (Alessio, 1993; Gul et al., 2001) exercise,
in rats may increase ROS production exceeding the capacity of
antioxidant defences. Oxidative stress is the imbalance of pro-
and anti-oxidants in favor of the former. Exercise-induced
oxidative stress was also reported in thoroughbred racehorses
after a 1000 m race at maximum velocity (White et al., 2001).
Increased oxidative stress can be harmful to all cellular
macromolecules, such as lipids, proteins and DNA (Halliwell
and Gutteridge, 1984). In contrast, beneficial effects of
endurance training on antioxidant defense mechanisms in
various tissues have been reported by swim (Kanter et al.,
1985; Venditti and Di Meo, 1996) and treadmill-trained normal
(Sen et al., 1992; Oztasan et al., 2004) and diabetic (Gul et al.,
2002) rats, and also in horses (Avellini et al., 1999) and men
(Dane et al., in press).
Heart muscle has a high oxygen uptake at resting conditions.
from the blood by heart muscle increases further during heavy
physical exercise (Wilmore and Costill, 1999; Gul and
Hänninen, 2002). Although it is essential for aerobic metabo-
lism, increased oxygen metabolism can sometimes lead to
increased oxidative stress in the heart during physical exercise
(Frankiewicz-Jozko et al., 1996; Venditti and Di Meo, 1996;
Asami et al., 1998; Gul et al., 2003). The primary source of
reactive oxygen species in heart is mitochondria. High oxygen
flux in the heart mitochondria may favor a higher rate of leakage
of free radicals (Ji, 1994). In addition, xanthine oxidase in
cytosole contributes to H2O2production in the heart especially
during ischemia–reperfusion injury (Bindoli et al., 1988). Heart
is equipped with all the major antioxidant enzymes, i.e. SOD,
CAT and GPx, as well as adequate levels of GRD and GST (Ji,
1994). As recently reviewed (Atalay and Sen, 1999), regular
physical exercise may beneficially influence cardiac antioxidant
defenses and promote overall cardiac function. However,
chronic exercise has dual effects: on the one hand, it results in
oxidant formation and oxidative stress; on the other hand,
perhaps as a consequence, it may also induce antioxidant
enzymes to minimize the effects of oxidative stress due to
m min−1with 10° inclination, for 60 min day−1) in Fisher 344
rats (Husain, 2003) and lower levels of lipid peroxidation after
ischemia and reperfusion in left ventricles of female Sprague-
et al., 1998) have been reported. On the other hand, unchanged
(Venditti and Di Meo, 1996), after 12-week treadmill training
(45-min duration at 25 m min−1with a 0% slope, 5 days a week)
in normal Wistar rats (Moran et al., 2004) and after an 8-week
treadmill training in diabetic Wistar rats (Gul et al., unpublished
results), and mice (Reddy Avula and Fernandes, 1999). Even
increased lipid peroxidation in heart tissue in female Sprague-
Dawley rats exercised at 1.6 km h−1, 2 h day−1, 5 days
week−1for 8 weeks (Liu et al., 2000) has been reported.
Alterations in the antioxidant enzymes due to endurance
training are also not consistent. Increased GPx activity in heart
tissue by endurance training has been reported in rats (Lew and
Quintanilha, 1991; Reddy Avula and Fernandes, 1999; Ramires
and Ji, 2001; Husain, 2003). Sprint training on a treadmill for
6 weeks increased GPx activity in heart of rats (Atalay et al.,
1996). However, unchanged heart tissue GPx activity by
training was reported in male Wistar (Moran et al., 2004) and
Sprague-Dawley (Ji et al., 1992) rats. In contrast, decreased
GPx activity in heart tissue in streptozotocin-induced diabetic
male Wistar rats by treadmill training for 8 weeks (Gul et al.,
2003) has also been reported. The situation is similar for
other antioxidant enzymes (Johnson, 2002). Whether endur-
ance training has beneficial effects on antioxidant defense
mechanisms and decreases lipid peroxidation in heart is not
yet clear. Therefore, the aim of this study was to investigate
whether an 8-week treadmill training strengthens antioxidant
enzymes and decreases lipid peroxidation in heart in male
rats. In addition, the effects of acute exhaustive exercise on
oxidative stress in heart tissue in untrained and trained rats
were also investigated. As recently reported, an 8-week
endurance training resulted in attenuated exercise-induced
oxidative stress in erythrocytes (Oztasan et al., 2004), but did
not affect the ventricular weights (Oztasan et al., in press) in
these rats. The results of this study may have implications in
2. Materials and methods
2.1. Animals and groups
12-week-old 56 male rats (Rattus norvegicus, Sprague-
Dawley strain) fed with standard laboratory chow and water
were used. Animal experimentations were approved by the
Ethical Committee of the Ataturk University and carried out in
an ethically proper way by following the guidelines provided.
2.2. Training and acute exhaustive exercise
Male rats were equally divided into trained (TR, TE) and
untrained groups (UR, UE) at random. Both groups were further
divided into two groups of 14 where the rats were studied at rest
(TR and UR) and immediately after exhaustive exercise (TE,
UE). After familiarizing the rats to the treadmill (MAY TME
9805 Treadmill Exerciser, Commat Iletisim Ltd., Ankara,
Turkey), endurance training began with gradual increases in
training speed and time such that rats were running 2.1 km h−1
at the fourth week. Training continued 1.5 h day−1, 5 days
week−1for 8 weeks. During the eighth week of training
program, the UE subgroup was also accustomed to treadmill
running 1.0–1.2 km h−1, 15 min day−1, for 5 days before
sample collection. This regimen was used to ensure that
untrained rats could also tolerate the acute exhaustive exercise
without having a significant training effect (Sen et al., 1992).
Two rats in the training group, which could not run well,
were excluded from the study. At the end of the training period,
half of the rats were randomly selected into the acute exercise
group. In acute exhaustive exercise, running speed was 1.2
km h−1(10% uphill gradient) for the first 10 min, after that,
240 M. Gul et al. / Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245
the speed was increased gradually to 2.1 km h−1at 95th min,
and kept constant until the rats were exhausted. The loss of
the righting reflex when the rats were turned on their backs
was the criterion of exhaustion.
2.3. Sample collection and tissue homogenization for heart
At the end of the experiment, the animals were anesthetized
with ketamine–HCl (Ketalar, 20 mg kg−1, i.p.). Heart was taken
out of the body after thoracotomy, auricles and vessels were
removed, rinsed in saline and stored in −80 °C.
A piece of heart tissue (approximately 300 mg) was
homogenized by an OMNI TH International, model TH 220
(Warrenton, VA, USA) homogenizer in 20 mM Tris–HCl, pH
7.4 (1:10 w/v) on ice for 30 s in the first speed level. Then, the
homogenate was centrifuged at 10000×g for 15 min at 4 °C.
The supernatant was stored at −80 °C in aliquots. Activities of
the antioxidant enzymes, GPx, GST, GRD, SOD, and CAT, and
TSSA (total, enzymatic plus non-enzymatic, superoxide radical
scavenger activity), NSSA (non-enzymatic superoxide radical
scavenger activity) and malondialdehyde (MDA) levels were
spectrophotometrically determined from these supernatants.
2.4. Biochemical measurements
2.4.1. TSSA, NSSA and SOD activity
TSSA and NSSA assays, as indicators of tissue antioxidant
capacity, were performed in the samples before and after adding
trichloroacetic acid (TCA, 20%), as described (Durak et al.,
1998). First, TSSA is measured. In this method, xanthine–
xanthine oxidase complex produces superoxide radicals that
react with nitroblue tetrazolium (NBT) to form a farmazone
compound. TSSA activity is measured at 560 nm by detecting
inhibition of this reaction. By using a blank reaction in which all
reagents are present except the supernatant sample and by
determining the absorbance of the sample and blank, TSSA
activity is calculated. Second, NSSA activity is measured in
TCA-treated fractions prepared by treating part of the sample
with final concentration of 20% (w/v) TCA solution (to remove
all enzymes and proteins), and centrifuging at 5000×g for
30 min. After the elimination of proteins by this procedure,
NSSA activity assay is performed in the supernatant
fraction. SOD activity is calculated as the difference
between TSSA and NSSA (Durak et al., 1998).
2.4.2. GPx, GST, GRD and CAT activities
GPx activity was measured by coupled spectrophotometric
assay at 340 nm from the oxidation of NADPH in the presence
of H2O2as substrate (Paglia and Valentina, 1967). GSTactivity
of the supernatant was measured by using 1-chloro-2,4-
dinitrobenzene (CDNB) and GSH as described (Habig et al.,
1974). GRD activity was determined by coupled spectropho-
tometric registration at 340 nm, using GSSG as substrate and
NADPH at 37 °C (Carlberg and Mannervik, 1985). CAT
activity was determined by the method of Aebi measuring the
rate of decay of H2O2absorbance at 240 nm (Aebi, 1984).
Protein concentration of the supernatant was measured by the
method of Bradford (1976).
2.5. Measurement of MDA level
MDA, an important indicator of lipid peroxidation, was
determined by spectrophotometry of the pink-colored product
of the thiobarbituric acid-reactive substances complex (Ohkawa
et al., 1979). Total thiobarbituric acid-reactive substances
(TBARS) were expressed as MDA.
2.6. Statistical analyses
One-way ANOVA with post-hoc LSD (Least significant
difference) test was used to compare group means. Unpaired t
tests were used to compare endurance times at the end of the
study and body weights both in the beginning and at the end of
8 weeks of treadmill training. Paired t test was also used to
compare the body masses before and after 8 weeks of training. P
values less than 0.05 were considered significant.
Malondialdehyde level in heart tissue was not affected
by acute exhaustive exercise in untrained and trained rats
The activities of GPx (Fig. 1) and GRD (Table 1) enzymes in
heart tissue decreased by both acute exercise and training. GST
and CAT activities in heart tissue were not affected by either
acute exercise or endurance training (Table 1). SOD activity was
decreased by acute exercise in untrained rats; however, this
decrease was not observed in trained rats (Fig. 2). Total
(enzymatic plus non-enzymatic) superoxide scavenger activity
Effects of endurancetraining andacute exhaustive exercise onlipid peroxidation
and antioxidant enzyme activities in heart tissue of untrained and trained rats
GroupsAt rest After acute exhaustive
26.30±2.93 29.37±4.15 27.60±2.51 23.79±3.04
17.63±1.22 14.34±1.55 14.02±1.31 13.95±0.95
9.27±0.707.69±0.80 8.22±0.83 7.91±0.73
8.27±0.64 7.01±0.73 7.33±0.737.21±0.68
Results given are means±S.E.M. One-way ANOVAwith post-hoc LSD test was
used to compare group means. MDA=malondialdehyde; GST=glutathione S-
transferase; GRD=glutathione reductase; TSSA=total (enzymatic plus non-
enzymatic) superoxide radical scavenger activity; NSSA=non-enzymatic
superoxide radical scavenger activity; CAT=catalase.
⁎ Untrained rest vs. all other groups; pb0.05, one-way ANOVAwith post-hoc
241 M. Gul et al. / Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245
and non-enzymatic superoxide scavenger activity were not
affected by either acute exercise or endurance training (Table 1).
Treadmill training increased the endurance time in trained
rats compared with sedentary rats (Oztasan et al., 2004).
Endurance times (mean±SD) of untrained and trained rats were
115±9 min and 170±26 min, respectively (pb0.001). There
was no difference between the increase in body weights (mean
±SD) in untrained (48±17 g) and trained (52±25) groups
during the study (Oztasan et al., 2004).
In line with our finding, increased endurance time in rats was
reported after treadmill training (2 h day−1, 5 days week−1for 5
weeks, at a speed of 30 m min−1at the fifth week, 15° uphill)
(Terblanche et al., 2001). As recently noted (Oztasan et al., in
press), there was no difference between ventricular weights of
untrained and trained groups in our study. No change in
ventricular or heart mass in rats (Fuller and Nutter, 1981;
Crisman et al., 1985; Senturk et al., 2001) and rabbits (Such et
al., 2002) was also reported in various endurance training
studies in line with our finding. Despite unaltered ventricular
mass, increased endurance may mean that ventricular function
should have improved by endurance training to increase stroke
volume in these studies.
4.1. Lipid peroxidation
MDA level in heart tissue was not affected by acute
exhaustive exercise in untrained rats. In agreement with our
finding, unchanged MDA level in heart tissue in female rats run
on the treadmill at 1.6 km h−1until exhaustion (Liu et al.,
2000), and unchanged TBARS levels in male diabetic rats after
exhaustive exercise (Gul et al., unpublished results) were found,
although heart tissue conjugated dienes levels were increased in
these diabetic rats (Gul et al., 2003). In contrast, after acute
exercise in heart tissue, increased MDA levels were found in the
old (25 months old) rats but not in young (8 months old) female
Fischer rats (Bejma et al., 2000), while increased TBARS levels
were found in both young and old male Wistar rats (Navarro-
Arevalo et al., 1999) and increased TBARS levels were found in
untrained rats after running until exhaustion (Frankiewicz-
Jozko et al., 1996). Increased MDA levels in heart tissue were
also reported after swimming to exhaustion (Rajguru et al.,
1994) and also after 30 min of swimming in rats (Turgut et
MDA level in heart tissue was not affected by acute
exhaustive exercise in 8-week treadmill-trained rats, either, in
our study. Parallel to our finding, unaltered TBARS levels were
found in Sprague-Dawley rats exercised to exhaustion by
swimming after 9 weeks of swim training (Benderitter et al.,
1996). While conjugated dienes levels were increased (Gul et
al., 2003), TBARS levels were not affected by acute exhaustive
exercise in heart tissue of endurance trained streptozotocin-
induced diabetic rats (Gul et al., unpublished results). In
contrast, increased TBARS levels in heart tissue after running
until exhaustion in trained rats (Frankiewicz-Jozko et al., 1996),
and increased MDA level in heart tissue after swimming to
exhaustion in swim-trained rats (Venditti and Di Meo, 1996)
have been reported.
MDA level in heart tissue was not affected by an 8-week
treadmill training in rats. Similarly, in heart tissue, unchanged
MDA level after a 10-week swim training (Venditti and Di Meo,
1996), unchanged TBARS after 12-week treadmill training
(45-min duration at 25 m min−1with a 0% slope, 5 days
week−1) in normal Wistar rats (Moran et al., 2004) and after
an 8-week treadmill training in diabetic Wistar rats (Gul et al.,
unpublished results), and unchanged lipid peroxide levels after
an 8-week treadmill training in mice (Reddy Avula and
Fernandes, 1999) have been reported. On the contrary,
decreased lipid peroxidation after training has also been
found. In heart tissue, decreased TBARS concentration in
trained rats (Frankiewicz-Jozko et al., 1996), decreased MDA
and protein carbonyls levels in Fisher 344 rats (Husain, 2003)
after an 8-week treadmill training (with a speed of 20 m min−1
with 10° inclination, for 60 min day−1) have been reported.
Decreased lipid peroxidation products, malondialdehyde and
after acute exhaustive exercise
SOD activity (U/mg protein)
Fig. 2. Superoxide dismutase (SOD) activity in the heart tissue of untrained
and trained rats at rest and after acute exhaustive exercise. The bars stand for
means±S.E.M. *Untrained rest vs. untrained and trained acute exercise
groups; pb0.05, one-way ANOVA with post-hoc LSD test.
GPx activity (U/mg protein)
after acute exhaustive exercise
Fig. 1. Glutathione peroxidase (GPx) activity in the heart tissue of untrained
and trained rats at rest and after acute exhaustive exercise. The bars stand for
means±S.E.M. *Untrained rest vs. all other groups; pb0.05, one-way
ANOVA with post-hoc LSD test.
242M. Gul et al. / Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245
lipofuscin in the left and right ventricles have been reported in
male rats after for 4 weeks of swim training (Ravi Kiran et al.,
2004). In addition, Powers et al. have found lower levels of
lipid peroxidation after ischemia and reperfusion in left
ventricles of female Sprague-Dawley rats after a 10-week
endurance exercise training (Powers et al., 1998). However,
increased MDA level in heart tissue was also found in female
Sprague-Dawley rats exercised at 1.6 km h−1, 2 h day−1, 5
days week−1for 8 weeks (Liu et al., 2000). Unaltered lipid
peroxidation after acute exercise or 8-week endurance training
found in our study suggests that heart tissue should have a
strong enzymatic and non-enzymatic antioxidant defense
against exercise-induced oxidative stress.
4.2. Antioxidant enzymes
The activities of GPx (Fig. 1) and GRD (Table 1) enzymes in
heart tissue were decreased by both acute exercise and training.
In agreement with decreased GPx activity in heart tissue after
acute exhaustive exercise in sedentary rats, decreased GPx
activity in heart tissue was found in streptozotocin-induced
Sprague-Dawley rats (Ji et al., 1992), or increased GPx activity
in heart tissue in Fischer 344 rats exercised acutely to 100% of
maximal oxygen consumption (Somani et al., 1995). Again,
in agreement with decreased GPx activity in heart tissue in
streptozotocin-induced diabetic male Wistar rats treadmill-
trained at 1.8 km h−1, 1.5 h day−1, 5 days week−1for 8 weeks
or 24 weeks (Moran et al., 2004), and in male Sprague-Dawley
ratsafter treadmill training (Jietal.,1992).Incontrast, increased
GPx activity in heart tissue was reported in Fisher 344 rats
trained for 6.5 (Husain and Somani, 1997), 8 (Husain, 2003) or
10 weeks (Somani et al., 1995), in female Sprague-Dawley rats
treadmill-trained (2 h day−1, 5 days week−1, at a speed of 26 m
and (2 h day−1, 5 days week−1, at a speed of 26 m min−1, up to
15% gradient, for 10 weeks) (Lew and Quintanilha, 1991) or in
female mice treadmill-trained daily from 45 to 50 min, at 1 km
h−1, 6 days week−1for a total of 8 weeks (Reddy Avula and
The activity of GRD (Table 1) in heart tissue decreased by
both acute exercise and training. However, unchanged GR
activity has been reported after acute exhaustive exercise in
sedentary and treadmill-trained streptozotocin-induced diabetic
male Wistar rats (Gul et al., 2003). Increased GR activity was
found after training in Sprague-Dawley rats (Lew and
Quintanilha, 1991; Ramires and Ji, 2001) for 10 weeks.
Unchanged GR activity has also been observed in male Wistar
rats, treadmill-trained for 12 or 24 weeks (Moran et al., 2004)
and in streptozotocin-induced diabetic male Wistar rats
treadmill-trained for 8 weeks (Gul et al., 2003).
GST and CAT activities in heart tissue were not affected by
either acute exercise or endurance training (Table 1). Parallel
our finding, unchanged glutathione S-transferase activity in
heart tissue after acute exercise, unaltered GST activity was
reported in sedentary and treadmill-trained streptozotocin-
induced diabetic male Wistar rats (Gul et al., 2003). Unaltered
GST activity was also reported in treadmill-trained male
Sprague-Dawley rats (Ji et al., 1992) and female mice (Reddy
Avula and Fernandes, 1999). In contrast to unchanged CAT
activity in heart tissue after acute exercise, increased CAT
activity was found after acute treadmill exercise in male Fisher
344 rats (Somani et al., 1995), in male Sprague-Dawley rats (Ji
et al., 1992), and also after 1 h swimming in Sprague-Dawley
rats (Terblanche, 2000). In agreement with unchanged CAT
activity in heart tissue after treadmill training, unchanged CAT
activity was reported in female Fischer 344 rats (Powers et al.,
1993) and male Sprague-Dawley rats (Ji et al., 1992) after
treadmill training. On the contrary, increased CAT activity was
observed in Fisher 344 rats trained for 8 weeks (Husain, 2003)
or 10 weeks (Somani et al., 1995), and also in Swiss White male
mice (Mus musculus) after swim training for 21 weeks (Kanter
et al., 1985).
SOD activity was decreased by acute exercise in untrained
rats; however, this decrease was not observed in trained rats
(Fig. 2). Total (enzymatic plus non-enzymatic) superoxide
scavenger activity and non-enzymatic superoxide scavenger
activity were not affected by either acute exercise or
endurance training (Table 1). However, unchanged (Ji et al.,
1992) or increased SOD activities have been reported in male
Wistar rats (Navarro-Arevalo et al., 1999), in Fisher 344 rats
(Somani et al., 1995), and in female Sprague-Dawley rats
(Ramires and Ji, 2001) after acute exercise. In line with our
finding of unchanged SOD activity after training, unchanged
SOD activities have been found after a 12-week treadmill
training in male Wistar rats (Ji et al., 1992; Moran et al.,
2004). On the other hand, increased SOD activities have been
reported by treadmill training of 6.5 weeks in male (Husain
and Somani, 1997), 8 weeks in male (Husain, 2003) and 10
weeks in female (Powers et al., 1993; Somani et al., 1995)
Fisher 344 rats, increased total and mitochondrial SOD
activities in male Wistar rats by 24-week treadmill training
(Moran et al., 2004), and in female Sprague-Dawley rats
treadmill-trained for 10 weeks (Ramires and Ji, 2001), and in
female mice treadmill-trained for 8 weeks (Reddy Avula and
Fernandes, 1999). Swim training for 4 weeks has also
increased superoxide dismutase (Mn-SOD) activity in the
left and right ventricles in male rats (Ravi Kiran et al., 2004).
TSSA and NSSA were not affected by both acute exhaustive
exercise and endurance training, which, to the best of our
knowledge, were first noted by this study.
Decreases in GPx and GRD enzyme activities would have
made the heart susceptible to exercise-induced oxidative
stress. However, we did not see an increase in lipid pero-
xidation due to acute exhaustive exercise or training. Since we
did not see much improvement in the antioxidant enzymes
(catalase, GST, SOD) in this study, it is possible that other
enzymatic and non-enzymatic antioxidants such as glutathi-
one, thioredoxin, vitamin E, etc., might have taken part in the
protection of the heart against exercise-induced oxidative
243 M. Gul et al. / Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245
stress in our study. Controversial results as oxidative markers
and antioxidants in response to acute exercise or training
reported in literature might arise from differences in the
models employed (low vs. high training intensity, running vs.
swimming, male vs. female, and rats vs. mice, etc.) and also
from the different oxidative stress parameters and methodo-
logies used for their determinations.
Although both acute exhaustive exercise and endurance
training decreased some antioxidant enzyme activities, they
could not induce oxidative stress in the heart tissue of the rats.
The decrease in SOD activity in heart observed in untrained rats
due to acute exercise was prevented by endurance training in
trained rats, revealing its potential role in myocardial antiox-
idant defense. Our results suggested that rat heart has sufficient
reserve of antioxidant enzyme capacity to cope with oxidative
stress induced by acute exhaustive exercise, and adaptive
changes in antioxidant enzymes due to endurance training are
This study has been supported by The Research Foundation
of the Ataturk University, Erzurum, Turkey (2001/53). The
authors thank Delali Camgoz who took care of the animals.
Aebi, H., 1984. Catalase in vitro. Methods Enzymol. 105, 121–126.
Alessio, H.M., 1993. Exercise-induced oxidative stress. Med. Sci. Sports Exerc.
Asami, S., Hirano, T., Yamaguchi, R., Tsurudome, Y., Itoh, H., Kasai, H., 1998.
Effects of forced and spontaneous exercise on 8-hydroxydeoxyguanosine
levels in rat organs. Biochem. Biophys. Res. Commun. 243, 678–682.
Atalay, M., Sen, C.K., 1999. Physical exercise and antioxidant defenses in the
heart. Ann. N.Y. Acad. Sci. 874, 169–177.
Atalay, M., Seene, T., Hanninen, O., Sen, C.K., 1996. Skeletal muscle and heart
antioxidantdefences in response to sprinttraining. Acta Physiol. Scand.158,
Avellini, L., Chiaradia, E., Gaiti, A., 1999. Effect of exercise training, selenium
and vitamin E on some free radical scavengers in horses (Equus caballus).
Comp. Biochem. Physiol. B 123, 147–154.
Bejma, J., Ramires, P., Ji, L.L., 2000. Free radical generation and oxidative
stress with ageing and exercise: differential effects in the myocardium and
liver. Acta Physiol. Scand. 169, 343–351.
Benderitter, M., Hadj-Saad, F., Lhuissier, M., Maupoil, V., Guilland, J.C.,
Rochette, L., 1996. Effects of exhaustive exercise and vitamin B6 deficiency
on free radical oxidative process in male trained rats. Free Radical Biol.
Med. 21, 541–549.
Bindoli, A., Cavallini, L., Rigobello, M.P., Coassin, M., Di Lisa, F., 1988.
Modification of the xanthine-converting enzyme of perfused rat heart during
ischemia and oxidative stress. Free Radical Biol. Med. 4, 163–167.
Bradford, M.M., 1976. A rapid and sensitive method for the quantitation of
microgram quantities of protein utilizing the principle of protein–dye
binding. Anal. Biochem. 72, 248–254.
Carlberg, I., Mannervik, B., 1985. Glutathione reductase. Methods Enzymol.
Crisman, R.P., Rittman, B., Tomanek, R.J., 1985. Exercise-induced myocardial
capillary growth in the spontaneously hypertensive rat. Microvasc. Res. 30,
Dane, S., Taysi, S., Gul, M., Akcay, F., Gunal, A., in press. Acute exercise
induced oxidative stress is prevented in erythrocytes of male long distance
athletes. Biol. Sport.
Durak, I., Canbolat, O., Kacmaz, M., Ozgen, G., Ozturk, H.S., 1998.
Antioxidant interferences in superoxide dismutase activity methods using
superoxide radical as substrate. Clin. Chem. Lab. Med. 36, 407–408.
Frankiewicz-Jozko, A., Faff, J., Sieradzan-Gabelska, B., 1996. Changes in
concentrations of tissue free radical marker and serum creatine kinase during
the post-exercise period in rats. Eur. J. Appl. Physiol. 74, 470–474.
Fuller,E.O.,Nutter,D.O.,1981.Endurancetrainingin therat: II. Performance of
isolated and intact heart. J. Appl. Physiol. 51, 941–947.
Gul, M., Hänninen, O., 2002. Physiological basis of exercise. In: Hänninen, O.,
Atalay, M. (Eds.), Physiology and Maintenance. In Encyclopedia of Life
SupportSystems(EOLSS),DevelopedUndertheAuspices ofthe UNESCO.
Eolss Publishers Co. Ltd., Oxford.
Gul, M., Kutay, F.Z., Temocin, S., Hanninen, O., 2000. Cellular and clinical
implications of glutathione. Indian J. Exp. Biol. 38, 625–634.
Gul, M., Oztasan, N., Taysi, S., Gumustekin, K., Akar, S., Bakan, N., Dane, S.,
2001. Short-term swimming exercise as an oxidative stress model in rat.
Hacet. J. Sport Sci. 12, 26–32 (in Turkish).
Gul, M., Laaksonen, D.E., Atalay, M., Vider, L., Hanninen, O., 2002. Effects of
endurance training on tissue glutathione homeostasis and lipid peroxidation
in streptozotocin-induced diabetic rats. Scand. J. Med. Sci. Sports 12,
Gul, M., Atalay, M., Hänninen, O., 2003. Endurance training and glutathione-
dependent antioxidant defense mechanism in heart of the diabetic rats.
J. Sports Sci. Med. 2, 52–61.
Habig, W.H., Pabst, M.J., Jakoby, W.B., 1974. Glutathione S-transferases. The
first enzymatic step in mercapturic acid formation. J. Biol. Chem. 249,
Halliwell, B., Gutteridge, J.M.C., 1984. Lipid peroxidation, oxygen radicals,
cell damage and antioxidant therapy. Lancet 1, 1396–1397.
Husain, K., 2003. Interaction of physical training and chronic nitroglycerin
treatment on bloodpressure, nitric oxide, and oxidants/antioxidants in the rat
heart. Pharmacol. Res. 48, 253–261.
Husain, K., Somani, S.M., 1997. Response of cardiac antioxidant system to
alcohol and exercise training in the rat. Alcohol 14, 301–307.
Ji, L.L., 1994. Exercise-induced oxidative stress in the heart. In: Sen, C.K.,
Packer, L., Hänninen, O. (Eds.), Exercise and Oxygen Toxicity. Elsevier
Science, B.V., Amsterdam, pp. 249–267.
Ji, L.L., Stratman, F.W., Lardy, H.A., 1992. Antioxidant enzyme response to
selenium deficiency in rat myocardium. J. Am. Coll. Nutr. 11, 79–86.
Johnson, P., 2002. Antioxidant enzyme expression in health and disease: effects
of exercise and hypertension. Comp. Biochem. Physiol. C Toxicol.
Pharmacol. 133, 493–505.
Kanter, M.M., Hamlin, R.L., Unverferth, D.V., Davis, H.W., Merola, A.J., 1985.
Effect of exercise training on antioxidant enzymes and cardiotoxicity of
doxorubicin. J. Appl. Physiol. 59, 1298–1303.
Khanna, S., Atalay, M., Laaksonen, D.E., Gul, M., Roy, S., Sen, C.K., 1999.
Alpha-lipoic acid supplementation: tissue glutathione homeostasis at rest
and after exercise. J. Appl. Physiol. 86, 1191–1196.
Lew, H., Quintanilha, A., 1991. Effects of endurance training and exercise on
tissue antioxidative capacity and acetaminophen detoxification. Eur. J. Drug
Metab. Pharmacokinet 16, 59–68.
Liu, J., Yeo, H.C., Overvik-Douki, E., Hagen, T., Doniger, S.J., Chu, D.W.,
Brooks, G.A., Ames, B.N., 2000. Chronically and acutely exercised rats:
biomarkers of oxidative stress and endogenous antioxidants. J. Appl.
Physiol. 89, 21–28.
Michiels, C., Raes, M., Toussaint, O., Remacle, J., 1994. Importance of Se-
glutathione peroxidase, catalase, and Cu/Zn-SOD for cell survival against
oxidative stress. Free Radical Biol. Med. 17, 235–248.
Moran, M., Delgado, J., Gonzalez, B., Manso, R., Megias, A., 2004. Responses
of rat myocardial antioxidant defences and heat shock protein HSP72
induced by 12 and 24-week treadmill training. Acta Physiol. Scand. 180,
Navarro-Arevalo, A., Canavate, C., Sanchez-del-Pino, M.J., 1999. Myocardial
and skeletal muscle aging and changes in oxidative stress in relationship to
rigorous exercise training. Mech. Ageing Dev. 108, 207–217.
244 M. Gul et al. / Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245
Ohkawa, H., Ohishi, N., Yagi, K., 1979. Assay for lipid peroxides in animal Download full-text
tissues by thiobarbituric acid reaction. Anal. Biochem. 95, 351–358.
Oztasan, N., Taysi, S., Gumustekin, K., Altinkaynak, K., Aktas, O., Timur, H.,
Siktar, E., Keles, S., Akar, S., Akcay, F., Dane, S., Gul, M., 2004. Endurance
training attenuates exercise-induced oxidative stress in erythrocytes in rat.
Eur. J. Appl. Physiol. 91, 622–627.
Oztasan, N., Timur, H., Siktar, E., Gumustekin, K., Akar, S., Dane, S., Gul, M.,
in press. Effects of endurance training on gonadal fat pad and ventricular
mass in rat. Biol. Sport.
Paglia, D.E., Valentina, W.N., 1967. Studies on the quantitative and qualitative
characterization of erythrocyte glutathione peroxidase. J. Lab. Clin. Med.
Powers, S.K., Criswell, D., Lawler, J., Martin, D., Lieu, F.K., Ji, L.L., Herb, R.
A., 1993. Rigorous exercise training increases superoxide dismutase activity
in ventricular myocardium. Am. J. Physiol. 265, H2094–H2098.
Powers, S.K., Demirel, H.A., Vincent, H.K., Coombes, J.S., Naito, H.,
Hamilton, K.L., Shanely, R.A., Jessup, J., 1998. Exercise training improves
myocardial tolerance to in vivo ischemia–reperfusion in the rat. Am. J.
Physiol. 275, R1468–R1477.
Rajguru, S.U., Yeargans, G.S., Seidler, N.W., 1994. Exercise causes oxidative
damage to rat skeletal muscle microsomes while increasing cellular
sulfhydryls. Life Sci. 54, 149–157.
Ramires, P.R., Ji, L.L., 2001. Glutathione supplementation and training
increases myocardial resistance to ischemia–reperfusion in vivo. Am. J.
Physiol.: Heart Circ. Physiol. 281, H679–H688.
Ravi Kiran, T., Subramanyam, M.V., Asha Devi, S., 2004. Swim exercise
training and adaptations in the antioxidant defense system of myocardium of
old rats: relationship to swim intensity and duration. Comp. Biochem.
Physiol. B 137, 187–196.
Reddy Avula, C.P., Fernandes, G., 1999. Modulation of antioxidant enzymes
and lipid peroxidation in salivary gland and other tissues in mice by
moderate treadmill exercise. Aging (Milano) 11, 246–252.
Sen, C.K., 1995. Oxidants and antioxidants in exercise. J. Appl. Physiol. 79,
Sen, C.K., 1997. Nutritional biochemistry of cellular glutathione. J. Nutr.
Biochem. 8, 660–672.
Sen, C.K., Marin, E., Kretzschmar, M., Hanninen,O., 1992. Skeletal muscle and
liver glutathione homeostasis in response to training, exercise, and
immobilization. J. Appl. Physiol. 73, 1265–1272.
Senturk, U.K., Gunduz, F., Kuru, O., Aktekin, M.R., Kipmen, D., Yalcin, O.,
Bor-Kucukatay, M., Yesilkaya, A., Baskurt, O.K., 2001. Exercise-induced
oxidativestress affects erythrocytes in sedentary rats but not exercise-trained
rats. J. Appl. Physiol. 91, 1999–2004.
Somani, S.M., Frank, S., Rybak, L.P., 1995. Responses of antioxidant system to
acute and trained exercise in rat heart subcellular fractions. Pharmacol.
Biochem. Behav. 51, 627–634.
Such, L., Rodriguez, A., Alberola, A., Lopez, L., Ruiz, R., Artal, L., Pons, I.,
Pons, M.L., Garcia, C., Chorro, F.J., 2002. Intrinsic changes on automatism,
conduction, and refractoriness by exercise in isolated rabbit heart. J. Appl.
Physiol. 92, 225–229.
Terblanche, S.E., 2000. The effects of exhaustive exercise on the activity levels
of catalase in various tissues of male and female rats. Cell Biol. Int. 23,
Terblanche, S.E., Gohil, K., Packer, L., Henderson, S., Brooks, G.A., 2001. The
effects of endurance training and exhaustive exercise on mitochondrial
enzymes in tissues of the rat (Rattus norvegicus). Comp. Biochem. Physiol.
A 128, 889–896.
Tiidus, P.M., 1998. Radical species in inflammation and overtraining. Can. J.
Physiol. Pharm. 76, 533–538.
Turgut, G., Demir, S., Genc, O., Karabulut, I., Akalin, N., 2003. The effect of
swimming exercise on lipid peroxidation in the rat brain, liver and heart.
Acta Physiol. Pharmacol. Bulg. 27, 43–45.
Venditti, P., Di Meo, S., 1996. Antioxidants, tissue damage, and endurance in
trained and untrained young male rats. Arch. Biochem. Biophys. 331,
White, A., Estrada, M., Walker, K., Wisnia, P., Filgueira, G., Valdes, F.,
Araneda, O.,Behn,C.,Martinez,R.,2001.Role ofexerciseandascorbateon
plasma antioxidant capacity in thoroughbred race horses. Comp. Biochem.
Physiol. A 128, 99–104.
Wilmore, J.H., Costill, D.L., 1999. Physiology of Sport and Exercise. Human
Kinetix, Second ed. Campaign IL.
245 M. Gul et al. / Comparative Biochemistry and Physiology, Part A 143 (2006) 239–245