Polar Biol (2006) 30:39–43
DOI 10.1007/s00300-006-0157-y
ORIGINAL PAPER
Inducible heat tolerance in Antarctic notothenioid fishes
Jason E. Podrabsky Æ George N. Somero
Received: 27 February 2006 / Revised: 30 April 2006 / Accepted: 8 May 2006 / Published online: 22 June 2006
Springer-Verlag 2006
Abstract Significant increases in heat tolerance (time
of survival at 14C) were observed for some, but not
all, species of notothenioid fishes collected from
McMurdo Sound, Antarctica (7751¢S) following
acclimation to 4C. The increase in thermal tolerance
was rapid in Trematomus bernacchii, developing within
1–2 days of acclimation to 4C. Long-term (6–8 weeks)
acclimation to 4C led to greater heat tolerance in
Trematomus pennellii than in T. bernacchii. Unlike its
demersal congeners, the cryopelagic notothenioid Pagothenia borchgrevinki did not increase heat tolerance
during warm acclimation. A deep-living zoarcid fish,
Lycodichthys dearborni, also failed to increase heat
tolerance, but survived significantly (> threefold)
longer at 14C than the notothenioids.
Introduction
Ectothermic animals endemic to Antarctic waters are
markedly stenothermal; many invertebrates and fishes
acclimatized to typical ambient water temperatures
(–1.8 to 2C) have upper incipient lethal temperatures
near or below 5–6C (Somero and DeVries 1967;
Clarke and Johnston 1996; Peck 2002, 2005; Peck et al.
J. E. Podrabsky (&)
Department of Biology, Portland State University,
Portland, OR 97207, USA
e-mail: [email protected]
G. N. Somero
Hopkins Marine Station, Stanford University,
Pacific Grove, CA 93950, USA
2004). Failure of important physiological activities such
as locomotion and burrowing may occur at even lower
temperatures (Peck et al. 2004). Little is known, however, about the capacities of Antarctic ectotherms to
acclimate to warmer temperatures, a question that is
significant in the context of increasing seawater temperatures due to climate change. In some areas of the
Southern ocean bordering Antarctica, sea surface
temperatures already have risen by approximately 1C
since the 1950s (Meredith and King 2005). Here, we
report the effects of short- (1–4 day) and long-term
(6–8 weeks) acclimation to 4C on thermal tolerance
(survival at 14C) of selected notothenioid and zoarcid
fishes from McMurdo Sound, Antarctica (7751¢S,
16638¢E). Previous work showed that specimens of
notothenioids acclimated to –1.86C had upper lethal
temperatures near 5–6C (Somero and DeVries 1967).
We wished to determine whether notothenioids and
zoarcids could alter their sensitivities to elevated
temperatures during warm acclimation and, if so, how
rapidly this capacity was gained.
Materials and methods
Specimens of four species of the Family Nototheniidae
(Suborder Notothenioidei), Trematomus bernacchii,
Trematomus pennellii, Trematomus hansoni and Pagothenia borchgrevinki, and the zoarcid Lycodichthys
dearborni were caught in McMurdo Sound by hookand-line fishing or in baited wire traps during the
months of December 2005 and January 2006. Specimens were returned to the aquarium facility in the
Crary Science and Engineering Center at McMurdo
Station and held in flow-through seawater aquaria at
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40
Results
Fish placed into 14C water typically showed rapid
bursts of swimming immediately after transfer. Subsequently, fish generally became quiescent until
shortly before death, when another, often brief bout
of swimming occurred. Death was commonly accompanied by loss of equilibrium (fishes lying on their
backs or sides), flaring of the opercula, and rigidity of
the trunk muscle. No effect of body mass on survival
time was found for any species (ANCOVA
p = 0.378).
Specimens of notothenioids acclimated (or field
acclimatized) to –1.8C survived at 14C for 10–
20 min, in agreement with earlier measurements
made on some of these species (Somero and DeVries
1967; Fig. 1a). Field-acclimatized and –1.8C-acclimated specimens of T. bernacchii did not differ in
survival time at 14C, suggesting that no effects on
123
Log10 (min to death)
a 5
4
3
2
1
0
4
5
6
7
8 9 10 11 12 13 14 15 16
Temperature,°C
b 200
Minutes to death at 14°C
ambient temperature (–1.8C) until used for experimentation. Fish were fed on chopped fish muscle every
few days during the acclimation period.
The protocol used to measure thermal tolerance
followed that of Somero and DeVries (1967). Fish were
transferred acutely from the –1.8C holding tank to a
30 l clear plastic aquarium containing aerated seawater
at 14 ± 0.1C. The choice of 14C as the thermal stress
temperature is based on the finding that –1.8C-acclimated notothenioids survive for approximately 10 min
at this temperature (Fig. 1a). Thus, 14C exposure allowed rapid screening of many specimens.
Acclimation at 4C involved two protocols: direct
transfer into 4C water or a slow increase in water
temperature from –1.8 to 4C over 24 h. Long-term
acclimation involved holding specimens of T. bernacchii and T. pennellii for 6–8 weeks following direct
transfer to 4C. Shorter term acclimation was done
with T. bernacchii, P. borchgrevinki, and L. dearborni
to examine the kinetics of the acclimation response
(change in survival time at 14C) over the first few days
of 4C acclimation. Due to limitations in the number of
available animals we were only able to follow the
T. bernacchii exposed to a slow temperature increase
for 3 days compared to 4 days for the fish directly
transferred to 4C.
Analysis of variance (ANOVA) and analysis of
covariance (ANCOVA) were used to examine the effects of body mass, species, and acclimation history of
the specimens. Specific comparisons between groups
were made using t tests or the Student–Newman–Keuls
(SNK) test where appropriate.
Polar Biol (2006) 30:39–43
180
160
140
120
100
80
60
40
20
0
E
-1.8°C
4.0°C
Field
D
A
B
L.d.
P.b.
B
T.h.
a
C
B,a
T.b.
Species
T.p.
Fig. 1 Thermal tolerance of Antarctic notothenioid and zoarcid
fishes following long-term acclimation to –1.8 or 4C. a Time to
mortality (min) of four notothenioid species acclimated to –1.8C
[L. dearborni (open square, L.d., n = 7), P. borchgrevinki (open
triangle, P.b., n = 4), T. hansoni (inverted triangle, T.h., n = 3),
T. bernacchii (open circle, T.b., n = 8), T. pennellii (open
diamond, T.p., n = 5)]. Filled symbols are data from Somero
and DeVries (1967); open symbols are data from the present
study. b Time to mortality (min) of notothenioid and zoarcid fish
following long-term (6–8 weeks) acclimation to 4 or –1.8C. Bars
represent means ± SEM. Bars labeled with the same letters are
not significantly different. Field-acclimatized T. bernacchii
(n = 5) did not differ in tolerance from fish laboratory-acclimated (n = 8) at –1.8C (lower case letters, t test p = 0.134). For
–1.8C-acclimated specimens, T. pennellii survived significantly
longer at 14C than other notothenioids, and L. dearborni
survived significantly longer than all notothenioids (upper case
letters, ANOVA, SNK p < 0.021). Long-term acclimation to 4C
led to significant increases (ANOVA p = 0.00004) in tolerance of
14C in T. bernacchii (n = 6) and T. pennellii (n = 5); the latter
species was significantly more heat tolerant than T. bernacchii
following 4C acclimation
thermal tolerance resulted from several weeks of
laboratory holding at –1.8C (Fig. 1b). T. pennellii
was significantly more tolerant of 14C than the other
notothenioid species. All four notothenioids were
less tolerant of 14C than the zoarcid species
Polar Biol (2006) 30:39–43
41
Minutes to death at 14°C
50
40
*
30
*
20
*
*
*
*
10
0
0
1
2
3
4
Days at 4°C
Fig. 2 Time course of acquisition of resistance to high temperature in T. bernacchii. Acute transfer of T. bernacchii to 4C
water (open circles) and increasing water temperature from –1.8
to 4C over 24 h (filled circles; 1 day point = 24 h at 4C) led to
similar kinetics of warm acclimation (ANOVA p = 0.06). Both
acclimation regimes induced a significant increase in thermal
tolerance within 24 h (ANOVA p = 0.008). Asterisk denotes
tolerances to 14C that are significantly greater than the
tolerance at t = 0 (ANOVA, SNK p < 0.015). Symbols are
means ± SEM. Time zero points are data presented in Fig. 1. For
all other data points n = 3
80
Minutes to death at 14°C
L. dearborni (Fig. 1b). Long-term acclimation of
T. bernacchii and T. pennellii to 4C led to significant
increases in heat tolerance, seven- and sixfold,
respectively (Fig. 1b). The heat tolerance of
4C-acclimated T. pennellii was significantly greater
than that of T. bernacchii.
The rate of acquisition of heat tolerance by T. bernacchii was rapid: within a day, fish subjected to either
acute transfer to 4C or a slow increase to this temperature over 24 h showed significant increases in
survival times at 14C (Fig. 2). The time-dependence
of acquisition of heat tolerance by T. bernacchii may
reflect a biphasic process. For the acutely transferred
specimens, the rapid increase in tolerance over the first
2 days of acclimation was followed by a decline in
tolerance by day 4. Presumably, a second phase of
warm acclimation was responsible for the higher level
of heat tolerance observed in fish acclimated to 4C for
6–8 weeks (Fig. 1b). Neither P. borchgrevinki nor
L. dearborni (Fig. 3) exhibited a significant increase in
survival time at 14C during short-term acclimation at
4C. However, linear regression analysis indicates a
significant positive slope in the data for P. borchgrevinki, which may indicate a slow acclimation of thermal
tolerance is possible over a longer time period than
examined in our study.
70
60
50
40
30
20
10
0
0
1
2
3
4
5
Days at 4°C
Fig. 3 Thermal tolerance of the zoarcid L. dearborni (open
circles) and the notothenioid P. borchgrevinki (filled circles),
during acclimation to 4C (acute transfer). No significant
increase in tolerance of 14C occurred during the 3–4 day
acclimation period for either species (ANOVA p > 0.30).
However, a significant slope was identified for the P. borchgrevinki data (p = 0.015), but not the L. dearborni data (p = 0.709).
Symbols are means ± SEM. Time zero points are data presented
in Fig. 1. For all other data points n = 3–5
Discussion
The teleost suborder Notothenioidei primarily comprises species endemic to Antarctic waters (Gon and
Heemstra 1990; Eastman 1993). Notothenioids have
radiated to fill many niches that opened during the
extreme cooling of the high latitude Southern Ocean,
which began approximately 50 million years ago and
was greatly accelerated when deep-water flow began in
the Drake Passage approximately 25 million years ago
(Eastman 1993). Approximately 10–15 million years
ago, a substantial sea ice cover began appearing near
the Antarctic continent, and water temperatures began
falling to their current low levels near the freezing
point of seawater, –1.86C. The Antarctic notothenioids thus have evolved in cold, thermally stable waters
for millions of years, and many aspects of their physiology and biochemistry indicate a high level of cold
adaptation (Clarke and Johnston 1996; Chen et al.
1997; Fields and Somero 1998; Kawall et al. 2002).
Evolution in thermally stable waters also has led to a
pronounced stenothermy in these fishes, as indicated
by their upper incipient lethal temperature of 5–6C in
the case of specimens acclimated or acclimatized to
normal ambient temperatures.
The present study suggests, however, that at least
some notothenioids may be capable of significantly
increasing their heat tolerance. For example, in
T. pennellii acclimation to 4C increased the time of
tolerance of 14C from 25 min to nearly 150 min
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42
(Fig. 1b). In T. bernacchii, tolerance time rose from
near 10 min to almost 68 min. These times of survival
to acute lethal heat shock do not, of course, reveal the
maximal temperatures at which indefinite survival can
occur. Further studies of long-term acclimation to
increasing temperatures may reveal this ultimate limit.
Future work may also reveal which physiological processes acclimate to temperature. No change in tissue
oxygen consumption during acclimation of T. bernacchii to 4C was observed (Somero et al. 1968), but
Seebacher et al. (2005) reported alterations in locomotory capacity and enzymatic activities in P. borchgrevinki following acclimation to 4C.
The discoveries that T. pennellii is more heat
tolerant than the other notothenioids studied and that
inter-specific variation exists in the ability to acquire
increased heat tolerance during acclimation to 4C
were unanticipated in view of the common thermal
exposure the species are likely to encounter in their
habitats and the evolutionary histories of these coldadapted species (Eastman 1993). In McMurdo Sound,
T. bernacchii, T. pennellii and P. borchgrevinki normally live at temperatures close to the freezing point of
seawater throughout their depths of occurrence. Annual variation in seawater temperature in McMurdo
Sound is no greater than 0.5C (Hunt et al. 2003). All
three species are circum-Antarctic in distribution (Gon
and Heemstra 1990; Eastman 1993), and all likely
encounter similar temperatures throughout their
biogeographic ranges. The physiological and genetic
differences among these species that determine their
different thermal tolerances and degrees of acclimatory plasticity merit investigation.
The zoarcid fish L. dearborni has only been collected in the Ross Sea, at depths of 550–588 m
(Anderson 1990; Gon and Heemstra 1990). Little is
known about the thermal physiology of zoarcids, a
primarily deep-sea family. A study by Mark et al.
(2002) indicated that another Antarctic zoarcid,
Pachycara brachycephalum, tolerated temperatures up
to 12C, but optimal physiological function occurred
only below about 6C. Thus, while Antarctic zoarcids
appear to be less stenothermal than notothenioids,
they too tolerate only a relatively narrow range of
temperatures compared to temperate eurythermal
fishes.
The kinetics of the change in heat tolerance during
warm-acclimation of T. bernacchii may be biphasic.
While this statement is presently based on limited data,
we feel that the implications of these data warrant
discussion. A rise in tolerance of 14C occurred rapidly
and the kinetics of acclimation following acute- and
slow ramp-up exposure to 4C were not statistically
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Polar Biol (2006) 30:39–43
different. The apparent reduction in tolerance of 14C
between days 2 and 4 of acclimation in the acute
transfer experiment and the subsequent acquisition of
a higher level of tolerance later in the 6–8 week period
of 4C acclimation may reflect the two-stage stress
response described recently by Kültz (2005). The initial
effects of stress lead to a ‘‘cellular stress response
(CSR)’’, which develops rapidly (within hours to days)
and leads to repair of cellular damage, especially to
membranes and proteins. Thereby, the CSR facilitates
the short-term survival of the cell following stress.
Then, a ‘‘cellular homeostatic response (CHR)’’ is
triggered, which involves a host of other molecular
level changes that differ from those associated with the
CSR and effect a restoration of a cellular homeostasis,
e.g., in ion balance, redox balance, and cell division.
On-going studies of temperature-induced changes in
gene expression in Antarctic notothenioids, using
cDNA microarrays, may reveal how the molecular
underpinnings of thermal tolerance change over poststress recovery in these species (B. Buckley and G. N.
Somero, in preparation). Notably, in T. bernacchii the
CSR does not involve the production of increased
amounts of heat-shock proteins (Hofmann et al. 2000)
or messenger RNA (Place and Hofmann 2004;
B. Buckley and G. N. Somero, in preparation).
Although rapid induction of heat-shock proteins is
commonly thought to be a key event in the CSR (see
Kültz 2005), T. bernacchii and at least some Antarctic
notothenioids (Place and Hofmann 2004) lack this
capacity. Thus, the induced thermal tolerance found in
T. bernacchii and other Antarctic notothenioids may
occur through different mechanisms than those
common to other species.
Acknowledgement We gratefully acknowledge the assistance
of Dr. G. Hofmann and colleagues in collection and maintenance
of specimens (through support of NSF grant OPP03-01927). We
also express our gratitude to the staff of the Crary Science and
Engineering Center for their valuable assistance. This work was
supported by NSF grant OPP05-04072 to Dr. Donal Manahan.
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