Diving experience and the aerobic dive capacity of muskrats: does training produce a better diver?
Department of Zoology, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2
* Author for correspondence (e-mail: rmacarth{at}ms.umanitoba.ca)
Accepted 10 January 2003
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Summary |
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Key words: dive conditioning, body oxygen store, dive behaviour, myoglobin, haemoglobin, muscle buffering capacity, metabolism, aerobic dive limit, muskrat, Ondatra zibethicus
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Introduction |
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The possibility that increased encounters with diving hypoxia contribute to
boosting the oxygen storage capacity of divers presents an intriguing
hypothesis that is amenable to testing. Yet few experimental studies have
specifically addressed the benefits of prior dive experience to any aspect of
diving capacity. In a study of the tufted duck Aythya fuligula,
Stephenson et al. (1989)
compared several key respiratory variables of control birds with those of
dive-conditioned ducks that were required to swim 6.0 m underwater in order to
feed. Following a 6-month training period, total body oxygen stores were
similar in both groups, though the partitioning of these stores was altered
significantly. These authors reported a decline in the lung-air sac oxygen
reserve of dive-conditioned ducks that was offset by a commensurate gain in
their blood and muscle stores. In a parallel study of immature harbor seals
Phoca vitulina, Kodama et al.
(1977
) compared several blood
indices of seals raised in a dry enclosure (`non-divers') with those of
animals that were provided free access to a large seawater tank (`divers'). At
the end of the 10-month conditioning period, these authors reported
significant gains in the blood haematocrit (Hct) and haemoglobin (Hb) levels
of `divers', which they attributed to hypoxia-induced erythropoiesis
associated with intermittent diving.
The paucity of experimental studies linking underwater experience with
physiological diving capability is surprising, given the considerable
attention devoted to the potential benefits of altitude hypoxia in boosting
the body oxygen reserves of terrestrial mammals especially humans (see
Böning, 1997;
Rodríguez et al.,
2000
). Exercise physiologists have repeatedly demonstrated that
training at altitude increases Hct, Hb and serum erythropoietin levels
(Mairbäurl, 1994
;
Klausen et al., 1996
;
Böning, 1997
). There is
also evidence that altitude hypoxia in combination with exercise is more
potent in stimulating erythropoiesis than is hypoxia alone
(Mairbäurl, 1994
). In a
more recent study, Rodríguez et al.
(2000
) reported that brief,
intermittent episodes of hypobaric hypoxia presented over a 3-week period were
sufficient to stimulate erythropoiesis and boost red cell mass, Hct and Hb
levels of humans. The effects of hypoxia conditioning on muscle oxygen
reserves are equivocal, though Terrados et al.
(1990
) observed increased Mb
levels in human leg muscle following hypobaric hypoxic training.
Thus, despite the considerable research effort focused on the physiological
benefits of hypoxia training in terrestrial mammals, little is currently known
about the ability of natural divers to modulate body oxygen reserves or other
indices of dive capacity in response to chronic changes in diving activity. An
excellent model for investigating this problem is provided by the semi-aquatic
muskrat Ondatra zibethicus Link 1795. Earlier research of muskrats
inhabiting a northern prairie marsh
(MacArthur, 1990;
MacArthur et al., 2001
)
established that the total body oxygen reserves of this rodent increase
2942% between summer and winter. In both studies, significant gains
were observed in the mass-specific blood volume, Hct, Hb and skeletal muscle
Mb of winter-caught animals. The oxygen-binding affinity of muskrat blood is
likewise elevated in winter compared to summer, and this increase appears to
be linked to a reduction in red cell 2,3-diphosphoglycerate (2,3-DPG) content
(MacArthur, 1984a
).
Winter-acclimatized muskrats also appear to be superior divers, exhibiting
greater cumulative and average dive times and longer dive:pause ratios than
adults tested in summer (MacArthur et al.,
2001
).
The underlying basis for these physiological and behavioural changes is
currently unknown, though one possibility relates to the increased reliance on
diving by muskrats after marshes freeze over, when most foraging and home
range movements dictate the necessity for underwater travel
(MacArthur, 1992). It is
conceivable that the winter gains in body oxygen reserves, diving ability, and
perhaps even whole blood oxygen affinity, develop in response to the increased
dependence on diving during the ice-bound season.
The primary goal of this study was to test the hypothesis that an increased dependence on diving leads to phenotypic upregulation in the body oxygen stores, aerobic dive limit (ADL) and diving ability of muskrats. We also considered the possibility that dive conditioning could result in greater reliance on anaerobic pathways in the primary swimming muscles of this species. To address these questions, we compared several key variables in acclimated muskrats trained to swim an underwater course to a feeding station, with those of animals precluded from diving but required to travel identical distances in water in order to feed. The specific variables examined included lung, blood and muscle oxygen stores, resting and diving metabolic rates, ADL, muscle buffering capacity, red cell 2,3-DPG content, and selected behavioural indices of dive performance.
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Materials and methods |
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Training tanks and testing protocol
Acclimated muskrats were randomly assigned to one of two identically
constructed fibreglass-lined, plywood tanks (internal dimensions: 217
cmx126 cmx60 cm) installed in a controlled-environment room set at
14±1°C with a 12 h:12 h L:D photoperiod. Each tank was partitioned
into eight interconnected swimming lanes by removable dividers, forming a 16
m-long maze. Detachable hardware cloth screens mounted on top of the dividers
prevented subject animals from escaping.
A wire cage (53 cmx53 cmx48 cm) containing two nest boxes (see above) was positioned above the maze, in one corner of the tank. An opening in the cage floor provided access, via a ramp, to the swimming maze. At the opposite end of the maze, a ramp provided access to a dry feeding station, consisting of a 53 cmx53 cmx48 cm cage mounted above the tank. Food consisted of ad libitum rations of rodent chow supplemented daily by fresh apples and carrots. In each training tank, animals could reach the feeding station only by traversing the 16 m maze separating the two cages.
The `control' tank was filled to a depth of 1012 cm with 14°C water, leaving an 1820 cm air space between the water surface and the tank cover. This shallow depth precluded diving but required muskrats to swim at the surface while negotiating the maze. In the experimental (`diving') tank, the maze was completely submerged (water depth=32 cm) and muskrats could move between their nest box and feeding station only by diving. During the first 34 days following release of muskrats into the diving tank, the water level was reduced to provide a 23-cm air space beneath the tank cover. This was necessary to allow animals to become familiar with the maze before requiring them to dive, thereby minimizing stress and the risk of accidental drowning in the early stages of training. When we were confident that muskrats could negotiate the maze, the water level was raised to eliminate the breathing space beneath the screen, thereby limiting all movement in the tank to underwater swimming. Based on 32 timed dives recorded from five trained muskrats, the time required for animals to dive from nest box to feeding station was 43.8±2.4 s (mean ± 1 S.E.M.; range 26.076.1 s).
Over the 34 day period immediately before introducing subjects into
their assigned tanks, a series of baseline (pre-training) measurements were
recorded from each animal. These included determinations of blood Hb and Hct
levels, resting rate of oxygen consumption
(O2) in air, as
well as voluntary dive times and
O2 recorded from
each animal during a 15 min diving trial (see below). Following baseline
measurements, each muskrat was released into its assigned holding tank where
it remained for a period of 911 weeks. In all tests, a pair of muskrats
was held in each tank, thus every dive-training session was accompanied by a
control run on an identical number of `surface-swimming' muskrats. In total,
12 animals (7 males, 5 females) were tested in the control tank, and 12
animals (5 males, 7 females) in the diving tank. The initial (pre-training)
body mass was similar in both groups (`surface swimmers' 813.0±38.5 g;
`divers' 787.6±37.7 g). Of the seven subadults included in the study
(438623 g), three were assigned to the control tank and four to the
diving tank. Within each test group, we observed no differences between adults
and subadults, hence data for both age classes were pooled.
Food rations were replenished daily and each tank was drained and cleaned at 2 day intervals. During the final 34 days of each training session, muskrats were removed from each tank for brief periods, in order to gather behavioural and metabolic data comparable to those collected prior to training. At the conclusion of testing, animals were killed and blood and tissue samples harvested for determination of relevant biochemical and respiratory measures (see below).
Metabolic and behavioural recordings in air and water
Pre- and post-training measurements of resting
O2 were obtained
from fasted muskrats at thermoneutrality (15±0.5°C) using positive
pressure, open-circuit respirometry
(MacArthur and Campbell,
1994
). Short-term (15 min) diving trials were conducted in a
fibreglass-lined, plywood tank (183 cmx175 cmx72 cm) housed in a
controlled-environment room. The tank was filled to a depth of 6870 cm
with warm (2930°C) water to minimize thermal stress for subjects. A
wire screen cover secured to a frame 3 cm below water level prevented diving
muskrats from surfacing at any point in the tank except in a 20.5 liter
respirometry chamber (MacArthur and
Krause, 1989
). Animals could swim or float at the surface in the
chamber but were prevented from leaving the water. At both the pre-training
and post-training stage, each animal was tested twice (on separate days). In
each case, the first trial provided a training session to familiarize the
animal with the tank and only data for the second trial were used in analyses.
The behavioural data gathered included frequency and duration of all
exploratory dives, as well as cumulative dive time for the 15 min trial.
Diving
O2 was
estimated following the procedure of MacArthur and Krause
(1989
).
Body oxygen stores
To obtain pre-training measures of blood Hb and Hct levels, blood samples
were drawn by cardiac puncture from animals lightly anaesthetized with an
inhalant anaesthetic (Halothane, MTC Pharmaceuticals). Following training and
completion of all metabolic and behavioural testing, each animal was
anaesthetized with an intramuscular injection of ketamine hydrochloride
(Rogar/STB Inc.) given in combination with xylazine (Haver-Lockhart) and
atropine sulphate. The left jugular vein was exposed, cannulated, and a blood
sample drawn for Hb, Hct and red cell 2,3-DPG determinations (MacArthur,
1984a;
1990
). Mean corpuscular Hb
concentration (MCHC) was calculated from Hb and Hct measurements
(Schalm et al., 1975
). Blood
volume was calculated from Hct and the plasma dilution of Evans Blue dye
(Swan and Nelson, 1971
;
El-Sayed et al., 1995
).
Subsequently, the muskrat was euthanized and muscle samples and the intact
lungs were removed from the carcass immediately. The ventricles and biceps
brachii, biceps femoris and gastrocnemius muscles were dissected free from fat
and connective tissue and immediately frozen at 70°C. Muscle Mb
concentrations were subsequently measured according to the technique described
by Reynafarje (1963
). Lung
volume, corrected to standard temperature and pressure (STPD), was determined
gravimetrically (MacArthur,
1990
).
Muscle oxygen stores were estimated from the mean Mb concentration of the
three skeletal muscles sampled, assuming skeletal muscle constitutes 44% of
the ingesta-free body mass of these rodents
(MacArthur et al., 2001). The
potential lung and blood oxygen stores of each individual were calculated in
accordance with MacArthur
(1990
) and followed
conventional protocol (Kooyman,
1989
).
Muscle buffering capacity and glycogen content
Intracellular buffering capacities of forelimb (biceps brachii) and hind
limb (biceps femoris) muscles were determined as described by Castellini and
Somero (1981). A 0.5 g sample
of frozen muscle was homogenized in 0.15 mol l1 NaCl and
titrated with 0.2 mol l1 NaOH using a Corning model 360i pH
meter equipped with an ISFET electrode (Fisher Scientific Canada, Whitby, ON,
Canada). Buffering capacity, measured in slykes, is defined as the µmoles
of base required to titrate the pH of 1 g wet mass of muscle by 1 pH unit,
over the pH range 6 to 7 (Van Slyke,
1922
). The glycogen content of these muscles was determined using
a spectrophotometric procedure based on a calibration curve derived for
glucose standards (Kemp and Kits Van
Heijningen, 1954
). Absorbances of standards and unknowns were read
at 520 nm using a Spectronic 601 Spectrophotometer (Rochester, NY, USA).
Treatment of data
Mean values for `surface swimmers' and `divers' were compared using one- or
two-tailed independent-samples t-tests
(Zar, 1984). Comparisons of
pre- and post-training values derived for the same individuals were made using
paired t-tests. For analyses of muscle Mb, glycogen and buffering
capacity, two-way analysis of variance (ANOVA) was used to evaluate sampling
site and treatment effects. Post hoc testing for differences in Mb
content between muscle sampling sites was made using Tukey's HSD test
(Zar, 1984
). All statistical
procedures were performed using SPSS software (version 9, 1999), following
KolmogorovSmirnov or ShapiroWilk tests for normality and
Levene's test for homogeneity of variance (SPSS). In the few cases when data
did not meet normality or homogeneity of variance assumptions, the data were
either transformed prior to statistical testing or the non-parametric
MannWhitney U-test was performed. Values are presented as
means ±1 S.E.M.
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Results |
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We considered the possibility that pre-training Hb and Hct levels might differ for muskrats captured in different months (JulyDecember), even though all animals were acclimated to the laboratory holding facilities for 46 weeks prior to testing. Results of a one-way ANOVA revealed that there was no relationship between month of capture and the initial (pre-training) blood Hb concentration of acclimated muskrats (F3,20=1.923, P=0.158). However, the pre-training Hct levels of acclimated animals did vary with time of capture (F3,20=5.516, P=0.006), averaging 45.60±1.14% for July (N=6), 41.88±1.21% for August (N=4), 47.28±1.03% for October (N=10) and 49.65±1.01% for December (N=4) captures.
Training effects on metabolic rate and diving behaviour
Following the 9- to 11-week training session, mean mass-specific resting
O2 values were
18.1% lower in `surface swimmers' (t=4.07, d.f.=11, P=0.002)
and 9.4% lower in `divers' (t=1.49, d.f.=11, P=0.164),
compared to their respective pre-training values
(Table 1). Mean diving
O2 values of
`surface swimmers' followed a similar trend
(Table 1), and were 22.1% lower
following training (t=2.67, d.f.=11, P=0.022). In contrast,
diving
O2 of
`divers' was not significantly affected by training (t=0.70, d.f.=11,
P=0.498). It is noteworthy that body mass was similar for `surface
swimmers' and `divers' prior to training, but animals in both groups
experienced significant (P<0.05) mass gains during the training
session. Mean mass gained by `divers' was 16.9±3.41%, compared to
19.5±4.73% for `surface swimmers'. However, these mass changes could
not account for the observed training effects on resting and diving
O2, since
conversion of
O2
to mass-independent units (ml O2 g0.67
h1) (Campbell and
MacArthur, 1998
) did not alter the observed metabolic trends.
Post-training metabolic rates tended to be highest for the dive-conditioned
animals, though only the increase observed in resting
O2 in air was
significant (Table 1).
We found little evidence that our training regime altered the diving behaviour of muskrats in either test group. Post-training measures of cumulative dive time, total number of dives, and overall mean dive duration did not differ from the respective pre-training values in either test group (Table 1) (P>0.05). However, the two indices of maximal dive time tended to be lower following training, especially in `divers' (Table 1). In this test group, the mean of the five longest and the single longest exploratory dive were reduced by 19.9% (t=2.179, d.f.=11, P=0.052) and 39.8% (t=1.951, d.f.=11, P=0.077), respectively, after training. We detected no differences in the dive behaviour of `surface swimmers' and `divers' following training (Table 1).
Training effects on blood, lung and muscle respiratory variables
The requirement for underwater swimming imposed on `divers' clearly
elevated the Hct and blood oxygen capacity of muskrats assigned to this test
group (Fig. 1). Compared to
initial (pre-training) values, blood Hct and Hb concentration of `divers' were
increased 10.8% (t=4.684, d.f.=11, P=0.0005) and 6.4%
(t=2.483, d.f.=11, P=0.015), respectively, after training.
This compares with differences of only 0.10.5% between pre- and
post-training measures of Hct and Hb levels recorded from `surface swimmers'
(P>0.05) (Fig. 1).
Final (post-training) Hct, Hb concentration, blood oxygen capacity and MCHC
were significantly higher in `divers' than in `surface swimmers'
(Fig. 1,
Table 2). The mean blood oxygen
store calculated for `divers' (22.9 ml O2 kg1)
was nearly 26% higher than that (18.2 ml O2 kg1)
derived for `surface swimmers' (t=1.990, d.f.=22, P=0.030)
(Fig. 2).
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The dive-training regime imposed on muskrats in this study had no apparent effect on lung volume, whole blood and plasma volumes, erythrocyte 2,3-DPG concentration (Table 2), nor on the glycogen level or buffering capacity of skeletal muscles (Table 3). Ventricular and skeletal muscle Mb levels were also similar in `surface swimmers' and `divers' following training (Tables 2 and 3). Consequently, the final lung and muscle oxygen stores calculated for each group were virtually identical (Fig. 2). However, owing to the significant gain in blood oxygen reserves, the mean total body oxygen store of `divers' (37.8 ml O2 STPD kg1) was 13.5% higher (t=1.874, d.f.=22, P=0.037) than for `surface swimmers' (33.3 ml O2 STPD kg1; Fig. 2).
|
Site-specific variability in muscle buffering capacity and glycogen
and Mb content
Although there was a tendency for glycogen level and buffering capacity to
be higher in the hind limb swimming muscles (biceps femoris) than in the
non-propulsive muscles of the forelimbs (b. brachii), these trends were not
statistically significant (P>0.05). Results of a two-way ANOVA
performed on the Mb data revealed a significant muscle site effect
(F1,88=30.12, P<0.0001), but no evidence of a
test group effect (F1,88=0.387, P=0.536) or a
muscle site x test group interaction (F3,88=0.194,
P=0.900). Though Mb levels of the hind limb muscles, especially
gastrocnemius, tended to slightly exceed those of the forelimbs
(Table 3), post hoc
comparisons (Tukey's HSD test) indicated no differences (P>0.05)
in mean Mb content amongst any of the skeletal muscles sampled. By comparison,
Mb content of the ventricles was consistently lower
(P=0.0060.0001) than levels recorded from any of the limb
muscles, accounting for the muscle site effect observed with ANOVA.
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Discussion |
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The most striking observation was the significant gain in the blood oxygen storage capacity of dive-conditioned muskrats. Not only did Hct, Hb and blood oxygen capacity of `divers' increase over the 9- to 11-week training period, but the final (post-training) values of this test group exceeded those of `surface swimmers' by 712% (Fig. 1; Table 2). We attribute the gains in Hb and Hct by dive-conditioned muskrats to intermittent hypoxia associated with underwater swimming through the 16 m maze separating nest box from feeding station in the dive tank. This conclusion is strengthened by the absence of any change whatsoever in the blood parameters of control animals (`surface swimmers') held concurrently in an identical tank furnished only with shallow water (Fig. 1). Though we cannot preclude the possibility that these differences may also reflect a stronger exercise response by the dive-trained group, it was our impression that the effort required to negotiate the maze was similar, whether muskrats were diving or swimming at the surface.
The results of this study are consistent with the comparison by Kodama et
al. (1977) of `diving'
(N=4) and `non-diving'(N=3) juvenile harbor seals reared in
captivity. Over a 10 month period, these authors reported modest gains in
several blood parameters, including Hb and Hct levels, in `divers' that had
continuous access to an outdoor seawater tank. To our knowledge, the only
other study that specifically addressed the benefits of dive conditioning in a
natural diver was by Stephenson et al.
(1989
) on tufted ducks. These
authors reported no significant gains in the Hb content, blood oxygen capacity
or red blood cell count of dive-conditioned ducks following a 6 month training
period when birds were required to swim a distance of 6 m underwater in order
to feed. However, their dive-conditioned ducks exhibited a 34% increase in
plasma volume, leading these researchers to conclude that this hypervolemia
was probably accompanied by dive-induced erythropoeisis, thereby avoiding the
haemodilution response commonly observed in training.
Our results harmonize with the extensive literature in exercise physiology
documenting elevated serum erythropoietin (EPO), Hb and Hct levels of mammals,
especially humans, following training under hypobaric hypoxic conditions (see
Böning, 1997;
Rodríguez et al.,
2000
). Several of these studies have established that intermittent
hypoxia, including that simulating sleep apnoea in humans, is sufficient to
stimulate EPO synthesis and/or elevate Hb and Hct levels
(Nattie and Doble, 1984
;
Knaupp et al., 1992
;
Rodríguez et al.,
2000
). Though EPO was not measured in this study, we assume that
the recurrent apnoeic episodes associated with underwater swimming provided a
sufficient hypoxic signal to stimulate renal EPO synthesis in dive-conditioned
muskrats. It is well-established that this glycoprotein stimulates
erythropoiesis by enhancing mitotic frequency and promoting iron uptake and Hb
synthesis by erythroid-committed cells in bone marrow
(Jelkmann, 1986
;
Kranz, 1991
). Consequently,
the observed gains in Hct, Hb and blood oxygen capacity of `divers' probably
reflect elevated EPO production by the kidneys of these animals.
The degree of hypoxia experienced by muskrats in this study is unknown, as
is the level required to promote EPO synthesis. In an earlier study of
controlled dives by restrained muskrats, MacArthur
(1986) reported that the
arterial partial pressure of oxygen (PaO2) fell
to 3.06 kPa after 8090 s of submergence. Though this is nearly twice
the mean duration of voluntary dives by muskrats in the maze, the earlier
study involved non-exercising rather than swimming muskrats, and the resultant
end-dive PaO2 was well below the
PaO2 (7.327.98 kPa) required to induce
polycythemia in the laboratory rat (Nattie
and Doble, 1984
).
To our knowledge, there is no evidence to suggest that EPO might also
account for the modest gain in MCHC of dive-trained animals
(Table 2). Human studies, for
example, have reported no change (Klausen
et al., 1991) or even a drop
(Richalet et al., 1994
) in
MCHC with hypoxia-induced increases in blood EPO level. Though plasma
osmolality was not recorded in this study, it is possible that changes in this
variable may have contributed to the observed differences in MCHC between
`surface swimmers' and `divers'. Irrespective of the proximal cause, an
elevated MCHC should theoretically enhance blood oxygen transport capacity
while limiting the attendant rise in blood viscosity
(Rodríguez et al.,
1999
). In this context, it is noteworthy that the MCHC of recently
captured, winter-acclimatized muskrats (38.5%) is nearly 10% higher than for
animals captured in summer (35.1%)
(MacArthur, 1984a
).
Despite the pre-eminent role often ascribed to muscle Mb in the maturation
of diving proficiency in marine birds and mammals (see
Ponganis et al., 1999;
Noren et al., 2001
), we found
no evidence suggesting this variable is responsive to dive conditioning in
muskrats (Tables 2 and
3). The Mb content of
ventricles and fore- and hindlimb skeletal muscles were virtually identical in
`divers' and `surface swimmers' (Table
3). Though Mb content tended to be higher in the hindlimb swimming
muscles than in the forelimb muscles, a trend previously noted in
field-acclimatized muskrats (MacArthur,
1990
; MacArthur et al.,
2001
), the differences in this study were not statistically
significant (P>0.05) (Table
3). The mean skeletal muscle Mb levels of `divers' and `surface
swimmers' following training (12.312.4 mg g1 wet
tissue; Table 2) were similar
to the level (12.1 mg g1 wet tissue) reported for recently
captured, summer-acclimatized adult muskrats
(MacArthur et al., 2001
).
However, these post-training values were 1011% lower than the mean
muscle Mb level (13.8 mg1 g wet tissue) of recently
captured, winter-acclimatized animals
(MacArthur et al., 2001
).
In their investigation of tufted ducks, Stephenson et al.
(1989) reported no change in
myocardial Mb, but significant gains in the Mb content of the pectoralis and
locomotor limb muscles of dive-conditioned birds. Though researchers have
speculated that the intermittent hypoxia accompanying early dive experience
may contribute to the upregulation of muscle Mb in young avian and mammalian
divers (Ponganis et al., 1999
;
Noren et al., 2001
),
experimental support for this hypothesis remains limited
(Stephenson et al., 1989
;
Terrado et al., 1990). In a previous investigation of developmental changes in
body oxygen stores of wild-caught muskrats
(MacArthur et al., 2001
), we
reported that skeletal muscle Mb content is strongly age- and mass-dependent
in animals ranging from 254 to 600 g (Mb=27.7xMass1.63,
r2=0.82). The present study suggests that as muskrats grow
and gain experience swimming underwater, dive-induced hypoxia probably has
minimal effect on the acquisition of adult levels of Mb.
Not surprisingly, lung oxygen stores were similar for `divers' and `surface
swimmers'. Our finding that red cell 2,3-DPG concentration also remained
unchanged (Table 2) suggests
there may be little, if any, modulation in the oxygen binding properties of Hb
by this organophosphate in response to dive conditioning. In an earlier study,
MacArthur (1984a) reported
that the affinity of muskrat blood for oxygen was greatest in winter and
suggested that at least one contributing factor was the reduction in red cell
2,3-DPG concentration observed during this season. Whatever the proximal cause
for the enhanced binding affinity of muskrat blood for oxygen in winter, this
shift should, in theory, enable diving animals to extract a greater fraction
of available oxygen from their lung reserves during under-ice excursions
(Snyder, 1983
;
MacArthur, 1984a
).
Metabolic and behavioural responses to training
An unexpected finding in this study was the tendency for resting and diving
O2 of control
animals to decline significantly over the course of the training period.
Metabolic rates of `divers' appeared to be more stable over time, though
resting
O2 of
this test group exhibited a slight, albeit non-significant decline (9.4%)
during the training session (Table
1). We considered the possibility that these trends might reflect
the tendency for animals in both groups to gain appreciable body mass during
training sessions (19.5% in `surface swimmers'; 16.9% in `divers'). However,
mass change alone cannot account for the observed depression in resting
metabolic rate, since conversion of
O2 to
mass-independent units (Campbell and
MacArthur, 1998
) did not eliminate this effect. It is relevant to
note that extensive subcutaneous and intra-abdominal fat depots were evident
in most subject animals at the time of necropsy. We suspect that the observed
gains in body mass were due mainly to fat accretion by muskrats maintained on
a high plane of nutrition in captivity and that the increase in body lipid
content may be implicated in the apparent decline in resting
O2
(Hayward, 1965
). Unfortunately,
body composition analyses were not performed on any of the subjects, hence we
cannot assess the extent to which variation in
O2 between and
within test groups may have arisen from differences in body lipid or protein
content. For example, the finding that mean post-training diving
O2 of `divers'
(2.22 ml g1 h1) was 14.4% higher than for
`surface swimmers' (1.94 ml g1 h1) could
indicate a gain in relative muscle mass by the dive-trained muskrats.
While the difference in diving
O2 between the
two test groups was not statistically significant
(Table 1), it nevertheless
offset the 1314% gain in total body oxygen stores by dive-conditioned
muskrats. Consequently, the calculated ADL
(MacArthur et al., 2001
) of
this group (61.3 s) was indistinguishable from that of control animals (61.8
s), a finding that may at least partially explain the observed similarities in
diving behaviour of animals in the two test groups
(Table 1).
Though we examined only two indices of anaerobic potential in skeletal muscle (glycogen content and buffering capacity), there was no evidence that either variable was upregulated in response to dive conditioning. The mean time required by muskrats to negotiate the underwater maze was 43.8 s, which is well within their calculated ADL, and it is doubtful that these animals routinely resorted to anaerobic metabolism during underwater swimming in the dive tank.
Concluding remarks
On balance, the results of this study demonstrate that the blood oxygen
stores of muskrats can be improved with an enforced diving regime over a
relatively short time course (911 weeks). Total body oxygen stores of
dive-conditioned animals were boosted 1314% beyond those of animals
that were prevented from diving (Fig.
2). However, equally striking was our failure to detect any
evidence whatsoever that the accessible oxygen reserves in lungs and muscles
can be manipulated by dive training. In contrast, Stephenson et al.
(1989) reported significant
gains in both blood and muscle oxygen stores of dive-conditioned ducks, but at
the expense of reduced oxygen availability in their respiratory system
following training. On the basis of this study, we cannot conclude that an
increase in diving activity is the sole, or even the primary, factor
responsible for the marked elevation in body oxygen reserves of muskrats in
winter (MacArthur, 1990
;
MacArthur et al., 2001
). These
earlier investigations established that total body oxygen stores of
field-acclimatized muskrats increased 2942% between summer and winter,
with most of the increase accounted for by gains in blood oxygen capacity.
Several factors may have contributed to our failure to observe changes of
similar magnitude in the laboratory. One is the possibility that 46
weeks was an insufficient period to fully acclimate field-caught muskrats
prior to training. This argument is supported by the observation that
pre-training Hct levels tended to be lowest for acclimated muskrats caught in
JulyAugust (41.945.6%, N=10) and highest for animals
captured in OctoberDecember (47.349.6%, N=14), a
similar trend to that reported for field-acclimatized animals
(MacArthur et al., 2001). On
the other hand, we found no evidence that month of capture significantly
affected the extent to which Hct or Hb levels were elevated following dive
conditioning. We similarly observed no relationship between time of capture
and the post-training Hct, Hb or muscle Mb levels of muskrats assigned to
either test group.
The absence of a stronger training response could also reflect the
artificial nature of the dive-conditioning protocol that required acclimated
muskrats to negotiate an underwater maze separating nest box from feeding
station. This limitation notwithstanding, the average dive time of muskrats
travelling between these sites in the dive tank (4344 s) was consistent
with average under-ice transit times calculated for free-living muskrats
swimming between resting and feeding shelters in winter (2042 s;
MacArthur, 1992). However, we
also recognize that other factors may be implicated in boosting body oxygen
reserves of winter-acclimatized muskrats in nature, including exposure to low
ambient temperatures (Deb and Hart,
1956
; Morrison et al.,
1966
) and the hypoxic-hypercapnic atmospheres of the winter
shelters frequented by these rodents
(MacArthur, 1984b
). Clearly,
further research is needed to assess the relative contributions of each of
these variables to the seasonal adjustments in body oxygen reserves and diving
proficiency of muskrats inhabiting northern marshes.
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