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Research Article
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Plasma Cortisol Changes and Body Composition in
Stizostedion lucioperca Exposed to Handling Stress |
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Abbasi Fatemeh,
Ghafori Sanaz
and
Jamili Shahla
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ABSTRACT
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Stizostedion lucioperca aquaculture/stoking, remains
a restrained industry due to several factors such as the paucity of freshwater
resources and studies on the physiological responses of this species under
environmental changes. The fish were subjected to handling stress by holding
them out of the water in a hand-held dip net for 30 sec and netting the
fish from the rearing tanks and transferring them to a small confinement
tank. Sufficient aeration was supplied to the confinement tank to revert
additional stress from oxygen depletion. Then measured changes in plasma
cortisol levels and the growth ability (body composition) in Stizostedion lucioperca subjected to handling stress. Blood samples were collected from the fish after exposure to the handling stress. Crude protein (Nx6.25)
was determined according to the Kjeldahl method, moisture content was
determined by oven drying at 105 ± 2 °C to constant weight and
ash by heating in a muffle furnace at 550 °C to constant weight. Total
lipids were extracted according to the Bligh and Dyer method. The results
indicated that, handling stress significantly increased the plasma levels
of cortisol 59.04 ng mL-1 versus 40.83 ng mL-1 in
control group. Also the decrease of the level of protein and lipid concentrations
show a significant difference between treatment and control (p<0.05).
As protein and lipid decreased, moisture increased from 78.19% in control
to 80.40% in treatment groups. According to the results, there was no
significant change in ash content in control and treatment groups which
was about 9%. In other words, it could be emphasized that nutrition-related
behavior of Stizostedion lucioperca resulting from the activation
of the hypothalamic/inter-renal axis in response to stress despite of
different reactions bear resemblance to that of other fishes. Present
data indicate that cortisol appears to be adequate to assess stress in
Stizostedion lucioperca.
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INTRODUCTION
tress in teleosts is characterized by the immediate release
of catecholamines and cortisol; both hormones are concerned with energy
reallocation from anabolic activities such as growth toward activities
to restore homeostasis (Wendelaar Bonga, 1997). Fish are sensitive to
acute and chronic environmental changes and show a stress response (Wedemeyer
et al., 1990; Barton and Iwama, 1991; Iwama et al., 1995;
Wendelaar Bonga, 1997; Barton et al., 1997; Barton, 2000). In aquaculture,
fish are frequently exposed to stressful situations such as handling and
confinement (Van der Salm et al., 2006). Acute stresses from different
origins are reported to change the hematological parameters in different
species (Demers and Bayne, 1997; Jeney et al., 1997; Barcellos
et al., 1999; Kubokawa et al., 1999; Lohner et al.,
2001; Biswas et al., 2004; Geslin and Auperin, 2004; Wang et
al., 2004). Exposure of fish to stressors can elicit physiological
changes at multiple levels of animal organization, these alterations are
collectively known as the stress response. The initial response to stress
represents the perception of an altered state and is characterized by
a neuroendocrine response, which includes the release of stress hormones,
such as cortisol (Hosoya et al., 2007). Cortisol is the major corticosteroid
produced during stress-induced activation of the Hypothalamic-Pituitary-Interrenal
(HPI) axis and is considered a principal component of the primary stress
response (Donaldson, 1981) cortisol is a major stress-related hormone
and its plasma level increases in response to stress (Billard and Gillet,
1981; Pickering et al., 1982; Sumpter and Donaldson, 1986).
The purpose of this study was to investigate the effects
of an acute handling and confinement stress on circulating levels of cortisol
in Stizostedion lucioperca. In addition, determine protein, lipid,
moisture and ash concentration and physiological measurements were made
to verify and expand upon.
MATERIALS AND METHODS
Fish: There were two experimental groups: control and stressed
fishes (two replicates in each). Twenty eight specimens of Stizostedion
lucioperca were obtained from the volga river and transported to one
of the culture ponds of the Research Institute of Aquaculture of the Shahid
Rajaii, Rasht City, Iran, were used for the stress experiment. Their body
weights (means±SEM) were 750±47 g, respectively.
Experiment 1: Acute handling and confinement stress: After one
week of acclimation, a group of fourteen was subjected to acute stress
according to Biswas et al. (2004). After 48 h, Approximately 1
mL of blood was collected from the caudal vein using a heparinized syringe
equipped with a 25 G needle and maintained on ice until centrifugation.
Anaesthesia, measurement and blood withdrawal took less than 3 min for
five fish. Plasma was separated by 3000 rpm centrifugation at 4°C for
15 min and stored at -80°C for subsequent analysis of different parameters.
Radioimmunoassays: Concentrations of cortisol in the plasma samples
were determined by radioimmunoassays as described by Takahashi et al.
(1985) and v for cortisol. All samples were measured in duplicate determinations.
Proximate analysis of muscle tissue: Crude protein (Nx6.25) was
determined according to the Kjeldahl method (AOAC, 1996), moisture content
was determined by oven drying at 105±2°C to constant weight and ash by
heating in a muffle furnace at 550°C to constant weight. Total lipids
were extracted according to the Bligh and Dyer method (1959), evaporated,
weighted and then redissolved in chloroform methanol 9:1, v/v; finally
they were stored at a temperature of 0°C.
Statistical analyses: The data were subjected to one -way ANOVA
to test the effects of feeding frequency and dietary moisture content.
Where significant p<0.05 differences were found in the one-way ANOVA test.
RESULTS AND DISCUSSION
Plasma cortisol levels in fish exposed to acute handling and confinement
stress rapidly to maximum value and circulating levels of cortisol were
significantly (p<0.05) different between treatments at the time of control
(from 40.82 ng mL-1 in control group to 59.04 ng mL-1
in treatment group) (Fig. 1).
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Fig. 1: |
Changes in mean plasma cortisol
levels with the standard error (a vertical line on each column)
in Stizostedion lucioperca exposed to the acute stress
during and after 48 h **: Significantly different at p<0.05 |
Protein and lipid concentrations show significantly decrease
between treatment and control (p<0.05). Protein from 20.82% in control
to 18.74% in treatment and lipid from 39% in control to 22% in treatment
decreased. Moisture from 78.19% in control to 80.40% in treatment increased
(Fig. 2).
Characteristic cortisol elevations of fishes in response
to acute stressors tend to range within about 30 and 300 ng mL-1
(Wedemeyer et al., 1990; Barton and Iwama, 1991) but there are
notable exceptions. Barton et al. (1997) and Barton (2000) observed
that peak levels in cortisol following an acute handling stressor were
low in scaphirhynchid sturgeons (Scaphirhynchus sp.) and paddlefish
(Polyodon spathula). Their results suggest a trend toward lower
stress responses in those chondrosteans compared with teleosts.
Response differences to stressors are clearly evident among
closely related fish species and such differences appear to be consistent.
Barton (2000) showed that brown trout (Salmo trutta) exhibited
greater cortisol increases after brief handling and short-term confinement,
respectively, than did rainbow trout (Oncorhynchus mykiss). This
difference was also consistent with glucose responses between these two
species. Similarly, Barton (2000) found that lake trout (Salvelinus
namaycush) were more sensitive to a transport stressor than brook
trout (Salvelinus fontinalis), a closely related char species.
The acute handling and confinement stress used in these experiments evoked
a clear and similar stress response in Stizostedion lucioperca,
confirming and
expanding upon the results of Biron and Benfey (1994). This
includes rapid elevation of plasma cortisol levels, indicative of activation
of the HPI axis and increased plasma glucose levels, due to stress-induced
mobilization of energy reserves.
The present results in the stress experiment clearly show
that Stizostedion lucioperca responded to the acute stress caused
by capture, confinement by increasing plasma cortisol levels. Similar
results have been reported already in many teleost species including diploid
rainbow trout (Dick and Dixon, 1985; Angelidis et al., 1987; Pickering
et al., 1987), brown trout (Salmo trutta) (Pickering et
al., 1982, 1987), Atlantic salmon (S. salar) (Pickering et
al., 1987) and salmon (Fagerlund, 1967; Billard and Gillet, 1981;
Pickering et al., 1982; Pickering and Pottinger, 1987; Carragher
et al., 1989; Carragher and Sumpter, 1990; Foo and Lam, 1993; Chopin
et al., 1996; Jardine et al., 1996; Waring et al.,
1996; Clearwater and Pankhurst, 1997).
Present results show that Stizostedion lucioperca
responded to acute stress by increasing cortisol. As for the mechanism
that determines the maximal level of cortisol in plasma, we suspect the
presence of a negative feedback mechanism in the hypothalamus-pituitary-interrenal
axes. Suppression of ACTH secretion by elevated cortisol level might determine
the maximal level of cortisol as suggested by Pickering and Pottinger
(1987).
The cortisol in plasma in Stizostedion lucioperca
captured in the lake were measured to confirm whether the cultured Stizostedion
lucioperca used in the stress experiment are physiologically equivalent
to wild individuals. The values of the physiological parameters observed
at the beginning of the experiment in cultured fish were comparable to
the values observed in wild individuals captured. Accordingly, the initial
condition of the cultured fish can be regarded as equivalent to the wild
fish after ovulation but before spawning under the present experimental
conditions. This allows us to further generalize the results of the stress
experiment.
In the present study, transporting the fish, keeping the
fish in the pond and handling the fish for bleeding in all likelihood
caused stress and affected plasma hormone and body composition.
tudies of protein concentration of control group in autumn
and winter show that protein concentration is increased (from 20.82% in
control to 18.74% in treatment) which is due to growth of physiology and
tissue substitution against to moisture. Therefore structural role and
shaping body tissue by protein is important. In this species, increase
in cortisol level causes increased in lipid metabolism, lipid decomposed
and therefore a release of FFA. Increasing oxidation of lipid acids lead
to intensification of metabolite. Sadovy (1993), also has studied about
body composition in different phase of reproductive and found that spawning
decrease tissue reserve such as lipid. Steven et al. (2000) have
studied body composition biochemistry in the season changes as a stress
stimulus in fish. In this research we found that lipid in control group
39% is decreased to 22% in treatment group. That is statistically significant.
These results consist of data and the difference is due to cortisol role
as a lipitic factor in the treatment. As protein and lipid decreased,
moisture from 78.19% in control to 80.40% in treatment increased. Therefore
protein and lipid tissue substitution with moisture tissue increase. Moisture
group in autumn and winter shows that, in winter is decreased in control.
According to the results, there is no significant change in ash (control
and treatment) it is about 9%.
However primary and secondary physiological response to
season changing stress in Stizostedion lucioperca is completely
normal, but this species is as an active and hunter species tertiary response
(low active changing behavior).
In conclusion, present results have shown that Stizostedion
lucioperca exhibit the typical physiological responses to acute stress
induced by handling and brief confinement and acute stress increased the
plasma levels of cortisol in this species. Similarly, the acute stress
induced decrease in total protein level parallels the findings in other
species.
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