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First published online August 17, 2006
Journal of Experimental Biology 209, 3429-3439 (2006)
Published by The Company of Biologists 2006
doi: 10.1242/jeb.02347
Associations between tissue fatty acid composition and physiological traits of performance and metabolism in the seabass (Dicentrarchus labrax)
1 CNRS-Ifremer UMR 10, Centre de Recherche sur les Écosystèmes
Marins et Aquacoles, Place du Séminaire, BP 5, 17137 L'Houmeau,
France
2 Unité mixte Nutrition, Aquaculture, Génomique
Inra-Ifremer-Bordeaux 1, Laboratoire Adaptation Reproduction Nutrition des
Poissons, Ifremer, Centre de Brest, BP 70, 29280 Plouzané,
France
* Author for correspondence (e-mail: Aurelien.Chatelier{at}crhl.ulaval.ca)
Accepted 24 May 2006
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) and net cardiac scope, and a higher
active metabolic rate (AMR) and aerobic scope (AS) than in the FO group.
Analysis of tissue FA composition revealed that the fish fed the CO and PO
diets had accumulated significantly higher levels of OA and LA in their heart
and muscle than the fish from the FO group, which had significantly higher
levels of highly unsaturated FA of the n-3 series, such as EPA and DHA
(20:5n-3 and 22:6n-3, respectively). Principal components analysis revealed
significant positive associations between tissue OA and LA content and
Ucrit, maximum Q, the increase in Q during
exercise, AMR and aerobic scope. There was a negative association between
these physiological traits and tissue content of EPA. Therefore, diet
composition is an environmental factor that can generate significant
phenotypic diversity in major physiological traits of performance and
metabolism in the seabass, with increased intake of FAs such as OA and LA
leading to improved cardiorespiratory performance.
Key words: seabass, Dicentrarchus labrax, swimming, metabolism, cardiovascular performance, fatty acid, diet
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;
Webb, 1998;
Videler, 1993), the energetics
of activity metabolism (e.g. Beamish,
1978; Jones and Randall,
1978; Webb, 1993),
and the use of locomotor capacity as a gauge of fish stress level and/or water
quality (e.g. Jain et al.,
1998; McKenzie et al.,
2003a). Logic would dictate that the ability to swim should factor
into the Darwinian fitness of many fish species and it is, therefore, widely
assumed that swimming performance will be critical to the success of
individual fish. Intra-specific diversities in performance, and the potential
sources of such diversity have, however, received little attention (reviewed
in Kolok, 1999;
Plaut, 2001;
Nelson et al., 2002) despite
the fact that this is the raw material upon which natural selection might act.
Clearly, individual physiological diversity may have genetic or environmental
origins but, at present, we have little understanding of the relative
contributions from each of these sources.
One environmental factor that is emerging as a significant source of
physiological diversity in fish is diet quality, and particularly the relative
intake and subsequent tissue accumulation of certain fatty acids (FAs)
(Tocher, 2003;
McKenzie, 2005). Fishes
accumulate FAs from their diet, storing them as neutral lipids
(triacylglycerols) and inserting them into membranes as polar
phosphoglycerides (Sargent et al.,
1999; Tocher,
2003). McKenzie et al.
(McKenzie et al., 1998) found
a direct positive relationship between the sustained aerobic exercise
performance of Atlantic salmon (Salmo salar) and their muscle levels
of oleic acid (OA; 18:1n-9) and linoleic acid (LA; 18:2n-6). On the other
hand, highly unsaturated FA of the n-3 series (n-3HUFA), namely EPA (20:5n-3)
and DHA (22:6n-3), have also been reported to have beneficial effects upon the
exercise performance of Atlantic salmon
(Wagner et al., 2004). Indeed,
the same n-3HUFA have a number of effects upon the cardiorespiratory
physiology of fish (McKenzie,
2001; McKenzie,
2005) that might impinge upon aerobic exercise performance. When
compared with individuals fed a diet rich in saturated fatty acids (SFA, such
as 14:0 and 16:0), whole-animal metabolic oxygen demand (standard and routine
metabolic rates) was significantly lower in Adriatic sturgeon (Acipenser
naccarii) and European eels (Anguilla anguilla) fed a diet rich
in n-3 HUFA (McKenzie, 2001).
These relative effects of SFA and n-3HUFA might influence aerobic swimming
ability if changes in standard and routine metabolic rates influence metabolic
scope for aerobic activities (Fry,
1971). Furthermore, in vitro experiments have
demonstrated that hearts from the sturgeon fed the diet enriched in SFA were
unable to maintain performance when oxygen supply was reduced, unlike hearts
from sturgeon fed the n-3HUFA (Agnisola et
al., 1996). The myocardium is a predominantly aerobic muscle, and
cardiac performance is believed to be one of the major factors underlying
aerobic exercise capacity in active teleosts
(Farrell, 1997;
Farrell, 2002;
Claireaux et al., 2005;
Clark et al., 2005). The
mechanisms for these effects of tissue FA are not known, although they
presumably derive from the manner in which different neutral triacylglycerols
are used as oxidative fuels, and/or from changes in cell function and
metabolism consequent to changes in the phosphoglyceride composition of
membranes (McKenzie, 2001;
McKenzie, 2005).
The objective of the current study was, therefore, to investigate how the
relative tissue levels of OA, LA, SFA and n-3HUFA influenced traits of
exercise performance, respiratory metabolism and in vivo cardiac
performance in the European seabass (Dicentrarchus labrax). Seabass
are active predatory teleosts that feed primarily upon smaller fish and
crustaceans, which they capture by pursuit
(Pickett and Pawson, 1994).
Young stages are subject to predation by pelagic fishes and several species of
bird (Pickett and Pawson,
1994). The species performs substantial migrations and is
profoundly euryhaline (Chatelier et al.,
2005), colonising environments ranging from offshore oceanic
waters to inshore, brackish and freshwaters in estuaries and coastal lagoons
(Pickett and Pawson, 1994).
Thus, exercise ability should be a correlate, and a predictor, of the fitness
of an individual seabass in its natural environment. The availability of
particular FAs (e.g. n-3HUFA and LA) can display large spatial and seasonal
fluctuations between estuarine and coastal marine foodwebs
(Galois et al., 1996), so the
different environments colonised by the seabass can be expected to provide
different diets.
Three groups of fish were fed for 4 months with one of three diets in which
lipids were provided as either canola oil (CO), palm oil (PO), or cod liver
oil, a fish oil (FO), to supply different amounts of dietary OA, LA, SFA and
n-3HUFA. Exercise respirometry and cardiac flow probes were then employed to
measure a suite of physiological traits of metabolism and performance. The
dietary oils provided a complex mixture of FA alongside specific ones of
interest. Therefore, tissue levels of FA and the suite of traits were all
measured upon the same fish, such that principal components analysis (PCA)
could be performed to highlight associations between physiological traits and
specific FA in the tissues. We investigated the hypothesis that fish with
tissues rich in OA and LA (from CO and PO) would exhibit improved exercise
performance by comparison with those with tissues rich in n-3HUFA (from FO).
We also investigated the hypothesis that an improved exercise performance in
seabass with tissues rich in OA and LA would be linked to greater aerobic
metabolic scope and in vivo cardiac performance. Furthermore, we
investigated the hypothesis that fish with tissues rich in SFA (from PO) would
have a higher metabolic rate than the fish with tissues rich in n-3HUFA (from
FO). Tissue polar and neutral lipid fractions were analysed separately, to
provide some insight into whether any associations between physiological
traits and tissue FA levels might derive from changes to membrane polar lipid
composition, or from the quality of neutral lipids as oxidative fuels
(McKenzie, 2001;
McKenzie, 2005).
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Experimental diets
The experimental diets were prepared at the Ifremer Centre de Nutrition des
Poissons (Brest, France) as 4.5 mm pellets. The basal composition of these
diets was identical. The addition of CO as dietary lipid provided a FA mixture
dominated by OA, LA and 
-linolenic acid (18:3n-3). Palm oil (PO)
provided a dietary FA mixture dominated by OA and LA but also by palmitic acid
(16:0), a SFA. The fish oil (FO) provided a mixture of many FA but, in
particular, relatively high quantities of EPA and DHA, n-3HUFA that were only
present at low levels in the two vegetable oils. The FA composition of the
three diets is shown in Table
1.
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Experimental animals and feeding
European seabass (Dicentrarchus labrax L.) with a mean mass
(±s.d.) 200±10 g and length 26.56±0.28 cm were obtained
from a commercial supplier on Ile de Ré (Charente Maritime, France).
They were maintained at CREMA in 1 m3 fibreglass tanks (water
volume approximately 400 liters) under a natural photoperiod. Tanks were
provided with biofiltered seawater (SW) at a temperature of 20°C and
salinity of 30
. A total of 72 fish were slightly anaesthetised, fitted
with a subcutaneous passive integrated transponder (PIT) tag for individual
identification, then allocated randomly to one of 6 experimental groups (2
replicates per diet, 12 individuals per replicate). The fish were then allowed
1 month of acclimation to the prevailing holding conditions, during which they
were fed a commercial diet (Bar D Perform Natura 4.5; Sica du Gouessant,
Lamballe, France). They were then fasted for 2 weeks prior to the beginning of
the feeding protocol, at which point they accepted the novel feeds eagerly.
Fish were then fed by hand daily to satiation, with their designated
experimental diet. After 4 months the fish had approximately doubled in mass.
Daily growth and condition factor were calculated as follows:
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where Mf is final mass in kg, Mi is initial mass in kg, n is the number of feeding days and BL is body length in cm.
Surgical preparation
At this stage of the experiment, bass with a mean mass and fork length
FL of 395±10 g and 30.58±0.28 cm, respectively, were
anaesthetised with tricaine methane sulphonate (MS-222) at a concentration of
0.1 g l-1, and transferred to an operating table where their gills
were irrigated with aerated water containing 0.05 g l-1 MS-222. A
2S-type Transonic (EMKA, Paris, France) ultrasound flow probe (resolution 0.1
ml min-1; absolute accuracy ±15%) was placed around the
ventral aorta, as described (Axelsson et
al., 2002). Following surgery, the animals were allowed 48 h
recovery in opaque PVC chambers provided with a flow of water.
Exercise and cardiac performance
Swimming respirometry was performed using an automated Brett-type
swim-tunnel respirometer designed to exercise fish in a non-turbulent water
flow with a uniform velocity profile
(McKenzie et al., 2001). Fish
were transferred individually to the respirometer and allowed to recover for
at least 12 h (overnight) at a current speed of 0.5 BL
s-1. At this low current speed bass rested on the bottom and
maintained position by gentle sculling of their pectoral fins and occasional
tail flicks. The following day, fish were exposed to progressive increments in
swimming speed at 1, 2, 3, 3.5 and 4 BL s-1, every 30 min,
until fatigue. Fish were considered to be fatigued when they were unable to
remove themselves from the posterior screen of the swimming chamber despite
gentle encouragement by sudden increases in current velocity. Measurements of
O2 uptake
(
, in
mg kg-1 h-1) were collected automatically at each
swimming speed with the custom-designed data-acquisition system described
elsewhere (McKenzie et al.,
2001) and custom-made software (G. Guillou, CREMA). Water
O2 saturation in the sealed respirometer was measured with an
Orbisphere clarke-type polarographic oxygen electrode and associated meter
(Orbisphere Laboratory, Geneva, Switzerland). The measurements of
during
exercise were used to derive the notional metabolic rate of the immobile fish
(IMR) (McKenzie et al.,
2003b). The maximum metabolic rate of activity (AMR) was
identified during swimming (this occurred at speeds approaching critical
swimming speed Ucrit) and used to calculate net aerobic
scope (AS) relative to IMR (McKenzie et
al., 2003b). Ucrit was calculated in
BL s-1 (Brett,
1964).
At each swimming speed, cardiac output
() was measured in ml min-1
kg-1 with the signal from the flowprobe displayed on the Transonic
amplifier and acquired by a PC with the custom-designed labview software
described elsewhere (Axelsson et al.,
2002). The signal was used to calculate heart rate
fH in beats min-1 and, together with the data
for , used to calculate cardiac stroke
volume VSH, in ml beat-1 kg-1.
Cardiac scope during exercise was calculated as
max-routine.
max always occurred at
swimming speeds approaching Ucrit,
routine was taken as the
lowest measured when the fish was
swimming very gently at a speed of 5 cm s-1, prior to the exercise
protocol. fH,routine and VSH,routine
were derived from the measures of
routine. Increase in
fH and VSH during exercise was
calculated, respectively, as
fH,max-fH,routine and
VSH,max-VSH,routine.
After experiments, animals were rapidly removed from the respirometer and killed with a blow to the head. The ventricle and a piece of white muscle were taken and stored at -80°C until fatty acid analysis.
Tissue fatty acid analysis
Total lipid extraction and measurement
Lipids were extracted from tissues following a procedure derived from that
of Folch et al. (Folch et al.,
1957). A double step extraction was carried out on rehydrated
samples by grinding them in chloroform:methanol mixtures (1:2 then 2:1, v/v)
with an all-glass Potter homogeniser. For each sample, the two homogenates
were filtered on GF/F pre-combusted filters (Whatman, Brentford, UK) and
pooled in a conical glass centrifuge tube. Following the addition of a 1% NaCl
solution, the crude extract separated into two phases. After decantation, the
lower phase containing lipids was recovered by suction and stored in
Teflon-capped glass tubes at -20°C until analysed.
Total lipids were measured with a Chromarod SIII-Iatroscan TH-10 system
(TLC-FID, Iatron Laboratories, Tokyo, Japan) connected to a CR3A integrator
(Shimadzu, Kyoto, Japan). For each sample, four successive volumes (1 µl)
of a concentrated extract aliquot were spotted on a Chromarod with a 2 µl
microsyringe. After drying in a dessicator, the rod was read directly by the
Iatroscan without any development. A calibration curve based on a mixture of
pure standards (Sigma-Aldrich, St Louis, MO, USA) was used to calculate the
total lipid concentration of the lipid extracts
(Parrish and Ackman,
1985).
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FAME preparation, purification and measurement
FAME from the separated lipid fractions were obtained after a
base-catalysed transesterification with sodium methylate (0.5 mol
l-1, 1 h, 80°C) (Christie,
1984).
FAME were purified by HPLC, using a preparative 100 mmx10 mm i.d. stainless steel column filled with an SPE Si-NH2 phase (IST). The polarity gradient was obtained by increasing the proportions of chloroform and methanol in heptane. As above, the column effluent was by-passed so that 90% of the purified FAME was recovered for gas chromatography, while the detector measured 10%.
Purified FAME were then analysed by gas chromatography (Packard 439) on a BPX-70 highly polar capillary column 30 mx0.25 mm i.d.x0.25 µm film (SGE, Courtaboeuf, France), with hydrogen as carrier gas at 1.1 ml min-1. The temperature gradient ran from 90 to 210°C at a rate of 1°C min-1. Identification of FAME was completed by comparison with pure individual standards (Sigma-Aldrich), standard mixtures (Supelco, Bellemonte, PA, USA) and hydrogenated samples.
Statistical analysis
Differences amongst the dietary groups for any given variable were assessed
by one-way analysis of variance (ANOVA) with Bonferroni post-hoc
tests to identify where significant differences lay. In those cases where the
groups did not exhibit homogeneity of variance, a Kruskal-Wallis
non-parametric ANOVA and Mann-Whitney post-hoc tests were used. Links
between heart polar lipids and physiological traits were explored by Principal
Component Analysis (PCA) followed by Pearson correlation tests, using
StatBox6® software. Only FA that represented at least 1% of tissue FA in
at least one lipid fraction (polar or neutral) were used for the PCA and
Pearson correlation tests. A probability of less than 5% (P<0.05)
was taken as the fiducial level for statistical significance.
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Exercise performance, cardiac performance and respiratory metabolism
Seabass fed the CO and PO diets achieved a mean (± s.e.m.)
Ucrit of 3.2±0.04 and 3.24±0.05 BL
s-1, respectively, which were not significantly different
(Fig. 1). In contrast, fish fed
FO only achieved a Ucrit of 2.93±0.12 BL
s-1, which was significantly lower than the other two groups
(Fig. 1).
There was no significant difference in
routine between diets
(Fig. 2), nor in routine
VSH and fH
(Table 2). There were, however,
differences in the cardiac response to exercise. As shown in
Fig. 2,
max was significantly
higher in CO and PO fish than in those from the FO group. As a consequence,
the increase in during exercise was
also significantly higher for CO and PO fish
(Fig. 2). In all fishes, the
increased during exercise was
primarily a consequence of increased fH, with a smaller
contribution from increases in VSH
(Table 2), but there were no
differences in the maxima for these latter two variables amongst the three
diets (Table 2).
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IMR was not significantly different between diets
(Fig. 3) but there were
significant differences in respiratory metabolism during exercise. Exercise
elicited an exponential increase in
in all
fish (data not shown) until a plateau was reached just prior to fatigue
(Ucrit). As shown in
Fig. 3, AMR was significantly
higher in the CO and PO groups relative to the FO group. As a direct
consequence, net aerobic scope was also significantly higher in the CO and PO
groups, relative to the FO group (Fig.
3).
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Tissue FA composition
As anticipated, there was a clear influence of dietary FA supply upon the
FA composition of neutral and polar lipids in the heart
(Table 3) and muscle
(Table 4). In polar lipids of
the heart (Table 3), the PO
fish had higher levels of stearic acid than found in the other two groups.
Both the CO and PO fish had higher levels of OA and LA in their polar lipids
than the FO fish did (Table 3).
On the other hand, the FO fish had higher levels of n-3HUFA such as EPA and
its elongation product 22:5n-3. DHA comprised about 28% of the heart polar
lipids in all groups, with no differences between them. The groups did not
differ in their overall levels of SFA, MUFA and PUFA but the FO group had a
much higher ratio of n-3 to n-6 PUFA in its polar lipids by comparison with
the other two groups. In the neutral lipids, stearic acid, OA, LA and n-3HUFA
exhibited the same overall pattern of distribution as seen for the polar
lipids (Table 3). However, the
dominant FA in the neutral lipids of all groups was not DHA but, rather, OA.
Furthermore, for their neutral lipids, the PO group had a higher total content
of SFA than the other two groups, and it also had a higher total content of
MUFA than the FO group (heart neutral MUFA levels varied greatly in the CO
fishes, Table 3). Conversely,
the PO group had a significantly lower level of total PUFA than the CO and FO
groups. Finally, the FO group, as expected, had a much higher ratio of total
n-3 to n-6 PUFA than in the other groups, although the differences were less
pronounced than in the polar lipids (Table
3).
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As can be seen in Table 4, skeletal white muscle also exhibited this general pattern of diversity in FA distribution between the three dietary groups. Indeed, the differences observed in the white muscle were, if anything, more pronounced than in the ventricular muscle (Table 3 cf. Table 4). This demonstrates that the diets had generated systemic differences in tissue FA profile amongst the three groups.
Principal component analysis
The PCA revealed some clear associations between the measured physiological
traits and levels of particular FA in the polar and neutral lipids of the
heart (Fig. 4). The analysis
also separated the groups along the primary axis, with the CO and PO fish
localised close together and apart from the FO fish
(Fig. 4). In particular, on
axis 1, fishes from the CO and PO groups with elevated levels of OA, LA and
total MUFA in their lipids had high Ucrit,
max, scope to increase
during exercise, AMR, and AS
(Fig. 4). On the other hand,
fishes from the FO group with high levels of n-3PUFA and a high ratio of n-3
to n-6 PUFA had low values for these traits of exercise and cardiac
performance but high values for growth and condition factor
(Fig. 4).
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These associations between FA and physiological traits were borne out by
the Pearson correlations derived from the PCA.
Table 5 shows the correlations
for polar lipids. Specific growth rate showed a weak negative correlation with
LA levels and a positive correlation with n-3 to n-6 PUFA ratio. Immobile
metabolic rate showed a strong positive correlation with stearic acid and
total SFA in polar lipids, and a negative correlation with DHA and total PUFA,
this latter probably because total PUFA were dominated by DHA
(Table 5). The performance
traits (Ucrit,
max, scope to increase
during exercise, AMR and AS) were all
correlated positively with LA and OA in polar lipids, but negatively with EPA,
22:5n-3 and ratio of n-3 to n-6 PUFA (Table
5). Table 6 shows
the correlations for neutral lipids. Once again, growth was correlated
negatively with stearic acid and total SFA levels in neutral lipids, while
both growth and condition factor were related positively with EPA levels.
Routine VSH was correlated positively with AA, DHA and
total PUFA in neutral lipids (Table
6). Performance traits were related positively to LA levels in
neutral lipids (but not neutral OA levels, which did not emerge from the PCA),
and related negatively to EPA levels and ratio of n-3 to n-6 PUFA
(Table 6).
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;
McKenzie et al., 1995;
McKenzie et al., 1997;
McKenzie et al., 1998;
McKenzie et al., 2000;
Agnisola et al., 1996;
Wagner et al., 2004). As
expected, the best physiological performance was observed in the animals fed
the CO and PO diets, rich in OA and LA. The PCA revealed a number of
associations between the percentage content of specific FAs in the polar
and/or neutral pools of ventricular muscle and particular traits of growth,
metabolism and performance.
The significantly higher Ucrit measured in the CO and
PO groups, as compared to the FO group, was linked to the high levels of OA
and LA in their tissues. These results are consistent with a previous study on
Atlantic salmon (McKenzie et al.,
1998), which found that incremental substitutions of fish oil by
canola oil in the dietary lipids led to incremental increases in swimming
performance, with a direct positive relation between tissue OA and LA (derived
from the dietary canola oil) and Ucrit. The current data
extend these observations to demonstrate that improved exercise performance
was linked to improved maximum cardiac performance, higher active metabolic
rates and greater aerobic scope in fish with tissues rich in OA and LA.
It is possible that the greater aerobic scope and higher swimming
performance of the CO and PO fish was a consequence of the improved cardiac
performance. It has been argued that aerobic myocardial performance may be a
primary factor limiting AMR and Ucrit in active teleosts
(Farrell, 2002;
Claireaux et al., 2005;
Clark et al., 2005). A recent
study has demonstrated that intrinsic individual diversities in
max are directly linked to
parallel diversities in AMR, AS and Ucrit in rainbow trout
(Claireaux et al., 2005). This
linkage has yet to be demonstrated for seabass, but there is some evidence for
a role of cardiac performance in defining aerobic scope and
Ucrit. In many active teleosts, including rainbow trout
and seabass, both and
plateau as fish approach Ucrit in swim tests
(Kiceniuk and Jones, 1977;
Kolok and Farrell, 1994;
Thoraresen et al., 1996; Gallaugher et
al., 2001; Chatelier et al.,
2005) and this has been taken as circumstantial evidence that it
is limitations to cardiac work that are constraining aerobic scope
(Farrell, 2002). In the
seabass, the simultaneous plateau of both
and
during
exercise corresponds with the initiation of an intermittent `burst and coast'
swimming pattern (Chatelier et al.,
2005) that indicates recruitment of anaerobic white muscle
(Day and Butler, 1996) and
which precedes fatigue. Visual observation of the ventral aortic probe trace
at this time of the experiment also revealed significant cardiac arrhythmias,
further circumstantial evidence that impaired cardiac performance was linked
to the onset of fatigue. This arrhythmia always occurred at higher swimming
speeds in the CO and PO fish, when compared to the FO animals.
At present, it is only possible to speculate about the mechanism by which
the tissue OA and LA might exert their effects upon cardiorespiratory
performance. They may be related to the fact that aerobic metabolism, and
aerobic work, is fuelled primarily by ß-oxidation of neutral FA in fish
(Hochachka and Somero, 1984;
Richards et al., 2002). There
is in vitro evidence to suggest that OA and LA are preferred over
other FA, especially HUFA, as substrates for ß-oxidation
(Sidell and Driedzic, 1985;
Henderson and Sargent, 1985;
Egginton, 1996). It is
conceivable, therefore, that higher levels of these preferred substrates in
the tissues might allow the animals to achieve higher rates of aerobic work
(McKenzie, 2001). It was
unexpected, therefore, that only levels of neutral LA in tissue lipids emerged
as being related to high performance in the PCA, and that levels of neutral OA
and total neutral MUFA showed no association. This might argue against such a
`substrate' mechanism.
The linkage between levels of OA and LA in polar lipids and increased
cardiac (and exercise) performance may also be a result of membrane processes.
Studies on isolated mammalian cardiomyocytes revealed that extracellular
application of EPA and other PUFA produced a prompt and reversible
concentration-dependent inhibition of the L-type calcium current, thereby
limiting calcium entry into the cells
(Xiao et al., 1997;
Leaf et al., 1999). It was
proposed that such effects should protect against calcium overload and
arrhythmia (Xiao et al., 1997;
Leaf et al., 1999). Any such
beneficial effect of EPA on cardiac arrhythmia was not revealed in our study.
There is, however, preliminary evidence to indicate that OA has an inhibitory
effect on L-type calcium channels and could, therefore, protect hearts against
arrhythmia at high workloads (A. Chatelier, N. Imbert, J. L. Zambonino
Infante, D. J. McKenzie and P. Bois, manuscript submitted for publication).
Such an effect might have contributed to the improved cardiac performance in
the CO and PO fish.
The relatively poorer exercise performance of the seabass with tissues rich
in EPA and with a high n-3 to n-6 PUFA ratio is also consistent with previous
results (McKenzie et al.,
1998) on Atlantic salmon. These results are, however, in direct
contrast to those reported by Wagner et al., who found improved exercise
performance in salmon fed a diet rich in n-3HUFA
(Wagner et al., 2004). The
explanation for these opposing results presumably lies in the enormous
complexity of factors within such diet studies. In particular, the oils used
in dietary studies all provide a complex mixture of FA. Many of these FA have
specific biological roles that interact with each other
(Sargent et al., 1999), such
that each study is effectively unique unless great care is taken to match the
ingredients. There may also be a minimum threshold of action for some FA. In
the current study, the relatively poor cardiorespiratory performance of fish
with tissues rich in EPA should not be taken as an indicator of overall
`reduced fitness', since tissue levels of this FA were correlated positively
with high fish growth rates. It is interesting that the PCA revealed that
these effects were correlated with EPA (and its elongation product 22:5 n-3)
rather than DHA, the other essential n-3HUFA provided in the FO diet.
The high IMR observed in fish with high tissue total SFA, revealed in the
PCA and Pearson correlations, are consistent with previous studies
(McKenzie et al., 1997;
McKenzie et al., 2000) where
sturgeon and eels fed SFA had significantly higher rates of metabolism than
those fed n-3HUFA. The results of the PCA indicate that this effect of SFA on
IMR in seabass may have been a result of a membrane-related mechanism, as the
associations were only observed for polar lipids. It is also interesting that
it was high tissue levels of DHA in polar lipids that were correlated with low
IMR in the PCA, with no other HUFA implicated.
Once again, it is only possible to speculate about the mechanism(s) for the
effects on metabolic rate of SFA versus DHA
(McKenzie, 2001). It has been
suggested that effects of n-3HUFA versus SFA on metabolic rate are
due to membrane-related processes, and particularly to differences in energy
consumption by membrane-bound ATPases, consequent to changes in membrane FA
composition and physico-chemical properties
(Hulbert and Else, 1999;
Hulbert et al., 2005). These
authors, however, report that n-3HUFA raise metabolic rate, while SFA lower
it, in a number of terrestrial endotherms (birds and mammals)
(Hulbert and Else, 1999;
Hulbert et al., 2005). This is
exactly opposite to the effects on metabolic rate reported here and previously
(McKenzie et al., 1997;
McKenzie et al., 2000). On the
other hand, hearts isolated from rats fed with n-3HUFA (from fish oil) had
significantly lower rates of myocardial oxygen consumption than hearts from
rats fed SFA (from coconut oil) (Pepe and
MacLennan, 2002), which was attributed to differences in
mitochondrial membrane composition that influenced proton leak
(Pepe and MacLennan, 2002).
The basis for such contrasting results is not clear and, while there can be no
doubt that dietary and tissue FA exert profound effects upon the metabolic and
cardiorespiratory physiology of vertebrates, further work is needed to explore
the mechanisms for these effects and thereby, hopefully, to explain
contrasting results.
The positive correlation between routine VSH and tissue
total PUFA levels, and in particular HUFA such as AA and DHA, cannot be
explained at present. Agnisola et al. found, when working spontaneously in
vitro, that hearts isolated from sturgeon fed a diet rich in n-3HUFA had
a greater routine VSH, than hearts isolated from sturgeon
fed a diet rich in SFA (Agnisola et al.,
1996).
In the present study at 20°C, seabass exhibited a much greater AMR,
aerobic scope, max and
Ucrit than the seabass at 15°C studied by Chatelier et
al. (Chatelier et al., 2005).
This demonstrates that temperature exerts a profound effect upon the
cardiorespiratory performance of seabass. In both the present study and that
of Chatelier et al. (Chatelier et al.,
2005), the increase in
during exercise was attributable to an increase in fH
rather than VSH. Rapid changes in water salinity had no
effect on cardiac or swimming performance
(Chatelier et al., 2005). This
indicates that diet may be at least as important an environmental variable as
salinity for defining the performance of seabass in their natural environment.
Migrations of seabass may expose the animals to entirely different FA
availabilities, particularly between marine foodwebs that are rich in n-3HUFA
and estuarine foodwebs where FA such as LA and OA may be more common
(Galois et al., 1996).
Exploring the associations between tissue FA composition and traits of growth,
performance and respiratory metabolism in wild seabass is, therefore, an
interesting avenue for future research.
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