Future migratory behaviour predicted from premigratory levels of gill Na+/K+-ATPase activity in individual wild brown trout (Salmo trutta)
1 Institute of Biology, University of Southern Denmark, Campusvej 55, 5230
Odense M, Denmark
2 Danish Institute for Fisheries Research, Vejlsøvej 39, 8600
Silkeborg, Denmark
* Author for correspondence at present address: Department of Clinical Immunology, Sdr Boulevard 29, 5000 Odense C, Denmark (e-mail: christian.nielsen{at}ouh.fyns-amt.dk)
Accepted 10 November 2003
![]() |
Summary |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: brown trout, Salmo trutta, migration, prediction of migratory behaviour, gill, Na+/K+-ATPase
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The existing knowledge of smolt physiology and migration in several species
is primarily based on comparisons of migrants and residents at the population
level, with residents having lower average gill
Na+/K+-ATPase activity than migrants at the time of
migration (Hart et al., 1981;
Zaugg, 1981
;
Ewing et al., 1984
;
Rodgers et al., 1987
;
McCormick and Björnsson,
1994
; Ewing and Rodgers,
1998
).
The aims of the present study were (1) to investigate the potential of regression analysis for describing the relationship between premigratory gill Na+/K+-ATPase activity, measured at two dates during spring, and future migratory behaviour of individual wild brown trout and (2) to evaluate the ability of the regression equation obtained for trout caught in two tributaries to predict future migratory behaviour in an independent group of trout caught in a distinct location in the mainstream. This was done by repeatedly measuring gill Na+/K+-ATPase activity in both descending (i.e. caught in a trap) and non-descending (caught by upstream electro-fishing) individuals of passive integrated transponder (PIT)-tagged wild brown trout before the smolt run in a large Danish river system.
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Capture, tagging and sampling
Tributaries
Electro-fishing (1000 W pulsed DC-current) was conducted over a stretch of
3 km of Haldum Brook and River Granslev Aa on 29 February 2000 and 1 March
2000, respectively. The captured wild brown trout were size selected, and 200
fish between 11.3 cm and 18.4 cm were chosen in both of the tributaries. The
fish were anaesthetised in a 5 mg l1 methomidate (Marnil TM,
Wildlife labs, Inc., Fort Collins, USA) solution and the total length
(LT) recorded to the nearest millimetre. Each fish was
implanted peritoneally with a PIT tag. Just prior to the implantation of the
PIT tags, while the fish was under anaesthesia, a small gill biopsy (four or
five tips of gill filaments) was removed from the first gill arch of each fish
and frozen in SEI buffer (300 mmol l1 sucrose, 50 mmol
l1 imidazole, 20 mmol l1 Na-EDTA, pH 7.3)
using a non-lethal gill biopsy method
(McCormick, 1993). This method
is reported to have no effect on gill Na+/K+-ATPase
activity (Rodgers et al.,
1987
) or the subsequent growth and survival of the fish
(McCormick, 1993
). The tagging
and sampling procedure was conducted at the river banks of the tributaries,
and prior to release the fish were allowed to recover at the banks in tanks
supplied with freshwater from the rivers. On 11 April 2000 (Haldum Brook) and
12 April 2000 (River Granslev Aa), a second round of electro-fishing was
conducted over the same stretches of the tributaries as during the first
sampling. All caught wild trout were anaesthetised and examined for PIT tags.
A small gill biopsy was taken from the other first gill arch of each
PIT-tagged fish and the LT was measured. Fish were allowed
to recover as described above and were subsequently released.
Mainstream
Electro-fishing was conducted on 6 April 2000 over a stretch of 3 km of the
River Lille Aa approximately 2225 km upstream from the traps. Two
hundred wild trout between 11 cm and 20.5 cm were selected, implanted with PIT
tags, sampled and further treated in the same way as the trout from the
tributaries. Fish from the mainstream were only sampled once.
Migratory behaviour
Migratory behaviour was established when PIT-tagged individuals were either
caught in the traps (i.e. migrants) or by electro-fishing after the smolt run
had ceased (i.e. residents) in early June 2000.
Analysis
Na+/K+-ATPase activity was analysed in gill
homogenates at 27°C by the method of McCormick
(1993) using a microtitre
plate reader (Spectramax, Molecular Devices, Sunnyvale, CA, USA). Protein
content in the tissue homogenates was measured by the Lowry method
(Lowry et al., 1951
), modified
for the microtitre plate reader, i.e. reagent volumes being adjusted for
96-well plates. Experiments were carried out subject to a licence from the
Animal Experimentation Inspectorate, Danish Ministry of Justice.
Statistics
To ascertain if future seaward migration or freshwater residency in wild
brown trout can be related to a premigratory measurement, the relationship
between premigratory determinations of either gill
Na+/K+-ATPase activity or LT and the
future migratory behaviour (i.e. migration or residency) in individual trout
from the tributaries and from the mainstream was investigated. A logistic
regression model was used to investigate the relationship between a continuous
independent variable (i.e. premigratory gill
Na+/K+-ATPase activity or LT) and a
categorical dependent variable with only two possible outcomes (i.e. migration
or residency). Logistic regression analysis was performed on pooled data from
the two tributaries and subsequently on data from the mainstream.
The logistic regression procedure involves the iterative fitting of a
linear function of the independent variable (X), i.e. premigratory
gill Na+/K+-ATPase activity or LT,
to the natural logarithm of the odds ratio for the dependent variable, i.e.
probability (P) of migration divided by probability of residency
(1P), using the following equation:
![]() |
Linear regression analysis was used to evaluate the relationship between premigratory LT and gill Na+/K+-ATPase activity, and for migrants only between premigratory gill Na+/K+-ATPase activity and the number of days from premigratory sampling to migration. In all cases, a significance level of P=0.05 was used.
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
The logistic regression equation relating future migratory behaviour of trout from the tributaries to the change in gill Na+/K+-ATPase activity between the first and second sampling was highly significant (P<0.000001, r2=0.64, a=2.549, b=1.242, N=75=60 migrants + 15 residents). However, no improvement in the ability of the regression model to identify individuals as migrants or residents was observed when data from both sampling periods were combined, as compared with using only the second sampling: 95% of the 60 future migrants, 80% of the 15 future residents, and 92% of the total of 75 individuals were correctly identified.
In the mainstream, a highly significant logistic regression equation describing future migratory behaviour as a function of gill Na+/K+-ATPase activity was found (P<0.000001, r2=0.48, a=1.283, b=4.040, N=104=88 migrants + 16 residents; Fig. 2). The regression model correctly identified 94% of the 88 future migrants, i.e. individuals that actually migrated 1454 days (median 19 days) after sampling. Of the 16 future residents, i.e. individuals that remained resident 63 days after sampling, 69% were correctly identified. In summary, the regression equation correctly identified the migratory strategy in 90% of the total of 104 individual fish.
|
Is LT related to migratory behaviour or to gill Na+/K+-ATPase activity?
No relationship between future migratory strategy and
LT was found for either of the two samplings in the
tributaries or for the mainstream sampling (P>0.05). Furthermore,
no relationship between the April measurements in both tributaries and
mainstream of premigratory LT and gill
Na+/K+-ATPase activity was observed (linear regression,
P=0.26, N=178).
Is gill Na+/K+-ATPase activity related to the number of days from sampling to migration?
For migrants only from both tributaries and mainstream, the premigratory
gill Na+/K+-ATPase activity was not related to the
number of days from premigratory sampling in April to migration (linear
regression, P=0.31, N=147;
Fig. 3).
|
Can the regression model for trout from the tributaries predict future migratory behaviour of trout from the mainstream?
The ability of the highly significant logistic regression model obtained
from the tributary-collected trout sampled in mid-April to predict future
migratory behaviour in the independent group of trout caught in early April in
the mainstream (River Lille Aa) was explored. The regression curve for the
tributaries is included for comparison in
Fig. 2.
A threshold probability of migration was used to predict the behaviour of the mainstream individuals as either future migrants, comprising fish with probabilities of migration higher than or equal to the threshold, or future residents, consisting of fish with probabilities of migration lower than the threshold.
Predicting migration
The effect of changing the threshold probability of migration on the
ability of the logistic regression model to predict future migration is
illustrated in Fig. 4. Setting
the threshold probability at 0, i.e. predicting all individuals as future
migrants, resulted in a predicted number of migrants equal to 104. The 104
predicted migrants, of course, included the 88 actual migrants but also the 16
actual residents. Hence, 85% of the predictions, i.e. 88 out of 104, regarding
future migrants among the mainstream individuals were correct using this
threshold probability. As the threshold probability was increased, changes in
predictive ability were observed. The first improvement occurred at a
threshold probability of migration of 0.04 (corresponding to a threshold gill
Na+/K+-ATPase activity of 1.8 µmol ADP
mg1 protein h1), where the number of
predicted migrants decreased to 103, including all 88 actual migrants. As the
threshold was increased further, the number of falsely predicted migrants
continued to decrease. The number of actual migrants included in the predicted
migrants remained at 88 until a threshold probability of 0.06 (corresponding
to a threshold gill Na+/K+-ATPase activity of 2.1
µmol ADP mg1 protein h1), with 87% of
the predictions regarding future migrants being correct, and then subsequently
decreased. At a threshold probability of 0.70 (corresponding to a threshold
gill Na+/K+-ATPase activity of 4.4 µmol ADP
mg1 protein h1), the 75 predicted migrants
included only one individual that actually remained resident (with 99% of the
predictions regarding migrants being correct). At a threshold probability
higher than 0.99 (corresponding to a threshold gill
Na+/K+-ATPase activity of 6.9 µmol ADP
mg1 protein h1), all predicted migrants
actually migrated.
|
Predicting residency
Fig. 4 illustrates the
effect of changing the threshold probability of migration on the ability of
the logistic regression model to predict future residency in mainstream
individuals. Setting the threshold probability at 1, i.e. predicting all
individuals as future residents, resulted in a predicted number of residents
equal to 104. These 104 predicted residents included the 16 actual residents
but also the 88 actual migrants, with only 15% of the predictions regarding
residents being correct. Lowering the threshold probability of migration
results in a decrease in the number of falsely predicted future residents. At
a threshold probability of 0.70 (corresponding to a threshold gill
Na+/K+-ATPase activity of 4.4 µmol ADP
mg1 protein h1), all but one of the 16
actual residents were included in the number of predicted residents, along
with 14 actual migrants (with 52% of the predictions regarding residents being
correct). At a threshold probability of 0.20 (i.e. a threshold gill
Na+/K+-ATPase activity of 2.9 µmol ADP
mg1 protein h1), the 12 predicted
residents included two individuals that actually migrated (with 83% of the
predictions regarding residency being correct). Only at a threshold
probability of 0.06 (corresponding to a threshold gill
Na+/K+-ATPase activity of 2.1 µmol ADP
mg1 protein h1) were these two actual
migrants separated from the one remaining actual resident.
Overall predictive ability of the logistic regression model for migratory behaviour of trout from the tributaries
The maximum percentage of correct predictions of future migratory behaviour
in fish from the mainstream was observed at threshold probabilities between
approximately 0.15 and 0.45 (corresponding to threshold gill
Na+/K+-ATPase activities of 2.73.7 µmol ADP
mg1 protein h1), with an average of 91% of
all predictions being correct.
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The robustness of the logistic regression analysis approach in predicting future migration or residency was illustrated when the future migratory behaviour of the mainstream individuals was successfully predicted using the logistic regression equation from the tributaries. Depending on the threshold probability of migration chosen, up to 91% of the predictions of future migratory strategy (migration or residency) were correct.
The rationale for using the level of gill
Na+/K+-ATPase activity to predict the future migratory
behaviour of individual brown trout was primarily based on previous reports of
correlations between gill Na+/K+-ATPase activity and
migratory behaviour at the population level in several different salmonid
species [e.g. rainbow trout (Oncorhynchus mykiss),
Zaugg and Wagner, 1973;
chinook salmon (Oncorhynchus tshawytscha),
Buckman and Ewing, 1982
; coho
salmon (Oncorhynchus kisutch),
Rodgers et al., 1987
; Atlantic
salmon, Nielsen et al., 2001
;
brown trout, Aarestrup et al.,
2000
]. Furthermore, the ability of a smolt to acclimate to
seawater may be enhanced by premigratory physiological priming and by coupling
of migration with elevated gill Na+/K+-ATPase activity,
as the level of gill Na+/K+-ATPase activity is
positively coupled to SW-tolerance in salmonids (e.g.
Nielsen et al., 1999
).
The high degree of accuracy of the predictive model is interesting in
several respects. Firstly, the method used for determining gill
Na+/K+-ATPase activity is an in vitro assay
using homogenized solutions of a few gill filaments under standardized
conditions, representing the functional capacity under optimized in
vitro conditions and not necessarily the true in vivo activity.
Secondly, the premigratory enzyme measurement correctly predicts a future
behaviour occurring days or weeks later. Finally, the smolt transformation
consists of a series of more or less independent and concurring changes in
physiology, morphology, metabolism and behaviour, being individually
controlled and coordinated by several hormones (e.g.
Hoar, 1988). Migratory
behaviour could be dependent upon the synchronization of several of the
changes associated with smoltification and it is unlikely that an increase in
gill Na+/K+-ATPase activity is the physiological factor
initiating migration. A single premigratory measurement of gill
Na+/K+-ATPase activity did, however, prove to be a very
accurate determinant of future migratory behaviour of wild brown trout.
Migration may be seen as the climax of smolt development in the river and
it has been hypothesized that migration only occurs when the smolt has passed
a physiological threshold condition that assures responsiveness to external
release factors (Solomon,
1978; Aarestrup et al.,
2000
). The transition from nearly no migrants below gill
Na+/K+-ATPase activities of approximately 2.5 µmol
ADP mg1 protein h1 to almost all migrants
above approximately 4 µmol ADP mg1 protein
h1 (Figs 1B,
2) suggests a threshold value
for differentiation of wild brown trout into migrant and resident individuals
occurring within this range of enzyme activities.
A note of caution regarding the general applicability of predictive
logistic regression models obtained using the methods described in the present
study is appropriate. Whether a regression equation for fish from one river
system can be utilized to predict the behaviour of fish from a separate river
system within a given year remains to be established. It is, however,
questionable as the physiological state (e.g. level of gill
Na+/K+-ATPase activity) at which the fish become
responsive to triggering factors and the nature of these factors must be
adapted to local conditions. Fish from different river systems may therefore
respond to different proximate factors in order to reach sea during an optimal
period. Heggberget et al.
(1993) showed that smolts of
Atlantic salmon in three different river systems had developed a specific
proximate trigger system for migration adapted to local conditions, but
whether the physiological state of the smolts at the time of the initiation of
migration differed as well was not investigated. Smolts of five strains of
Atlantic salmon originating from different river systems have been shown to
initiate downstream migration differently when released into the same foreign
river (Nielsen et al., 2001
).
However, the level of gill Na+/K+-ATPase activity at the
time of initiation of migration was approximately the same in the different
strains, suggesting a comparable threshold level of gill
Na+/K+-ATPase activity in populations of salmonids of
the same species living in different river systems.
Similarly, the ability to predict behaviour in a given river system from a
regression model obtained in the same river in another year should be
ascertained. Photoperiod and, to a lesser degree, temperature regulate the
neuroendocrine alterations that result in the physiological changes during
smolting (Hoar, 1988). Water
temperature is believed to be the rate-determining factor of the physiological
development, and once a certain physiological smolt condition is reached
(Solomon, 1978
), water
temperature (Jonsson and Ruud-Hansen,
1985
) and discharge (Hansen and
Jonsson, 1985
) are factors known to have a rapid stimulatory
effect on the initiation of downstream migration in salmonids. The response to
these cues may have evolved by natural selection to increase the survival of
the smolts, because the timing of the environmental cues changes from year to
year (McCormick et al., 1998
;
Wedemeyer et al., 1980
). The
timing of smolt migration is reported to be controlled by a combination of
actual temperature and temperature increases in the water during spring
(Jonsson and Ruud-Hansen,
1985
). As the temperature profile in a particular river varies
from year to year, the time when the physiological development is reached is
likely to differ and the time of the main run of Atlantic salmon smolts is
reported to vary between 0 and 14 days between different years
(Jonsson and Ruud-Hansen,
1985
; Whalen et al.,
1999
).
Finally, the data obtained in the present study regarding the temporal applicability of the regression model should be considered. In the tributaries in late Feruaryearly March, i.e. two months before the main run, physiological differentiation in terms of gill Na+/K+-ATPase activity was not yet initiated and it was not possible to predict future migratory behaviour from gill Na+/K+-ATPase activity. In comparison, the premigratory measurement of gill Na+/K+-ATPase activity from mid-April, i.e. 23 weeks (median time before capture in trap) before the main run, very successfully predicted the future migratory behaviour of the individual fish from the tributaries. In Fig. 2, the displacement along the x-axis of the regression curve for the tributaries compared with the curve for the mainstream probably reflects that the mainstream sampling was performed 56 days before the mid-April sampling in the tributaries.
In conclusion, a non-lethal premigratory biochemical measurement of gill Na+/K+-ATPase activity was successful in selecting individual brown trout with a high probability of future migration. The present findings should encourage similar studies of the migratory behaviour in other species.
![]() |
Acknowledgments |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Aarestrup, K., Nielsen, C. and Madsen, S. S. (2000). Relationship between gill Na+,K+-ATPase activity and downstream displacement in domesticated and F1-offspring of wild anadromous brown trout (Salmo trutta) released into a Danish river. Can. J. Fish. Aquat. Sci. 57,2086 -2095.[CrossRef]
Björnsson, B. Th., Thorarensen, J., Hirano, T., Ogasawara, T. and Kristinsson, J. B. (1989). Photoperiod and temperature affect plasma growth hormone levels, growth condition factor and hypoosmoregulatory ability of juvenile Atlantic salmon (Salmo salar) during parrsmolt transformation. Aquaculture 82, 77-91.[CrossRef]
Buckman, M. and Ewing, R. D. (1982). Relationship between size and time of entry into the sea and gill (Na+,K+)-ATPase activity for juvenile spring chinook salmon. Trans. Am. Fish. Soc. 111,681 -687.
Ewing, R. D. and Rodgers, J. R. (1998). Changes in physiological indices of smolting during seaward migration of wild coho salmon, Oncorhynchus kisutch. Aquaculture 168, 69-83.[CrossRef]
Ewing, R. D., Evenson, M. D., Birks, E. K. and Hemmingsen, A. H. (1984). Indices of parrsmolt transformation in juvenile steelhead trout (Salmo gairdneri) undergoing volitional release at Cole Rivers Hatchery, Oregon. Aquaculture 40,209 -221.[CrossRef]
Hansen, L. P. and Jonsson, B. (1985). Downstream migration of hatchery reared smolts of Atlantic salmon (Salmo salar L.) in the River Imsa, Norway. Aquaculture 45,237 -248.[CrossRef]
Hart, C. E., Concannon, G., Fustish, C. A. and Ewing, R. D. (1981). Seaward migration and gill Na+,K+-ATPase activity of spring chinook salmon in an artificial stream. Trans. Am. Fish. Soc. 110, 44-50.
Heggberget, T. G., Johnsen, B. O., Hindar, K., Jonsson, B., Hansen, L. P., Hvidsten, N. A. and Jensen, A. J. (1993). Interactions between wild and cultured Atlantic salmon: a review of the Norwegian experience. Fish. Res. 18,123 -146.[CrossRef]
Hindar, K., Jonsson, B., Ryman, N. and Staahl, G. (1991). Genetic relationships among landlocked, resident, and anadromous brown trout, Salmo trutta L. Heredity 66,83 -91.
Hoar, W. S. (1988). The physiology of smolting salmonids. In Fish Physiology. Vol. XIB The Physiology of Developing Fish (ed. W. S. Hoar and D. J. Randall), pp.275 -343. London: Academic Press.
Jonsson, B. and Ruud-Hansen, J. (1985). Water temperature as the primary influence on timing of seaward migrations of Atlantic salmon Salmo salar smolt. Can. J. Fish. Aquat. Sci. 42,593 -595.
L'Abée-Lund, J. H., Jonsson, B., Jensen, A. J., Sættem, L. M., Heggberget, T. G., Johnsen, B. O. and Næsje, T. F. (1989). Latitudinal variation in life-history characteristics of sea-run migrant brown trout Salmo trutta. J. Anim. Ecol. 58,525 -542.
Lowry, O. H., Rosebrough, N. J., Farr, A. L. and Randall, R.
J. (1951). Protein measurement with the folin phenol reagent.
J. Biol. Chem. 193,265
-275.
Marshall, W. S. (1995). Transport processes in isolated teleost epithelia: opercular epithelium and urinary bladder. In Fish Physiology. Vol. 14 Cellular and Molecular Approaches to Fish Ionic Regulation (ed. C. M. Wood and T. J. Shuttleworth), pp.1 -23. New York: Academic Press.
McCormick, S. D. (1993). Methods for nonlethal gill biopsy and measurement of Na+,K+-ATPase activity. Can. J. Fish. Aquat. Sci. 50,656 -658.
McCormick, S. D. and Björnsson, B. Th. (1994). Physiological and hormonal differences among Atlantic salmon parr and smolts reared in the wild, and hatchery smolts. Aquaculture 121,235 -244.[CrossRef]
McCormick, S. D. and Saunders, R. L. (1987). Preparatory physiological adaptations for marine life of salmonids: osmoregulation, growth and metabolism. Am. Fish. Soc. Sym. 1,211 -229.
McCormick, S. D., Björnsson, B. Th., Sheridan, M., Eilertson, C., Carey, J. B. and O'dea, M. (1995). Increased daylength stimulates plasma growth hormone and gill Na+,K+-ATPase in Atlantic salmon (Salmo salar). J. Comp. Physiol. B 165,245 -254.
McCormick, S., Hansen, L. P., Quinn, T. P. and Saunders, R. L. (1998). Movement, migration and smolting of Atlantic salmon (Salmo salar). Can. J. Fish Aquat. Sci. 55,77 -92.[CrossRef]
Nielsen, C., Holdensgaard, G., Petersen, H. C., Björnsson, B. Th. and Madsen, S. S. (2001). Genetic differences in physiology, growth hormone levels and migratory behaviour of Atlantic salmon smolts. J. Fish Biol. 59, 28-44.[CrossRef]
Nielsen, C., Madsen, S. S. and Björnsson, B. Th. (1999). Changes in branchial and intestinal osmoregulatory mechanisms and growth hormone levels during smolting in hatchery-reared and wild brown trout. J. Fish Biol. 54,799 -818.[CrossRef]
Rodgers, J. D., Ewing, R. D. and Hall, J. D. (1987). Physiological changes during seaward migration of wild juvenile coho salmon (Oncorhynchus kisutch). Can. J. Fish. Aquat. Sci. 44,452 -457.
Solomon, D. J. (1978). Some observations on smolt migration in a chalkstream. J. Fish Biol. 12,571 -574.
Wedemeyer, G. A., Saunders, R. L. and Clarke, W. C. (1980). Environmental factors affecting smoltification and early marine survival of anadromous salmonids. Mar. Fish. Rev. 42,1 -14.
Whalen, K. G., Parrish, D. L. and McCormick, S. D. (1999). Migration timing of Atlantic salmon smolts relative to environmental and physiological factors. Trans. Am. Fish. Soc. 128,289 -301.
Zaugg, W. S. (1981). Advanced photoperiod and water temperature effects on gill Na+-K+-adenosine triphosphatase activity and migration of juvenile steelhead (Salmo gairdneri). Can. J. Fish. Aquat. Sci. 38,758 -764.
Zaugg, W. S. and Wagner, H. H. (1973). Gill ATPase activity related to parrsmolt transformation and migration in steelhead trout (Salmo gairdneri): influence of photoperiod and temperature. Comp. Biochem. Physiol. B 45,955 -965.[CrossRef][Medline]