Oxygen limitation of thermal tolerance in cod, Gadus morhua L., studied by magnetic resonance imaging and on-line venous oxygen monitoring

The hypothesis of an oxygen-limited thermal tolerance due to restrictions in cardiovascular performance at extreme temperatures was tested in Atlantic cod, Gadus morhua (North Sea). Heart rate, changes in arterial and venous blood flow, and venous oxygen tensions were determined during an acute temperature change to define pejus ("getting worse") temperatures that border the thermal optimum range. An exponential increase in heart rate occurred between 2 and 16 degrees C (Q(10) = 2.38 +/- 0.35). Thermal sensitivity was reduced beyond 16 degrees C when cardiac arrhythmia became visible. Flow-weighted magnetic resonance imaging (MRI) measurements of temperature-dependent blood flow revealed no exponential but a hyperbolic increase of blood flow with a moderate linear increase at temperatures >4 degrees C. Therefore, temperature-dependent heart rate increments are not mirrored by similar increments in blood flow. Venous Po(2) (Pv(O(2))), which reflects the quality of oxygen supply to the heart of cod (no coronary circulation present), followed an inverse U-shaped curve with highest Pv(O(2)) levels at 5.0 +/- 0.2 degrees C. Thermal limitation of circulatory performance in cod set in below 2 degrees C and beyond 7 degrees C, respectively, characterized by decreased Pv(O(2)). Further warming led to a sharp drop in Pv(O(2)) beyond 16.1 +/- 1.2 degrees C in accordance with the onset of cardiac arrhythmia and, likely, the critical temperature. In conclusion, progressive cooling or warming brings cod from a temperature range of optimum cardiac performance into a pejus range, when aerobic scope falls before critical temperatures are reached. These patterns might cause a shift in the geographical distribution of cod with global warming.     

Climate variations and the physiological basis of temperature dependent biogeography: systemic to molecular hierarchy of thermal tolerance in animals

The physiological mechanisms limiting and adjusting cold and heat tolerance have regained interest in the light of global warming and associated shifts in the geographical distribution of ectothermic animals. Recent comparative studies, largely carried out on marine ectotherms, indicate that the processes and limits of thermal tolerance are linked with the adjustment of aerobic scope and capacity of the whole animal as a crucial step in thermal adaptation on top of parallel adjustments at the molecular or membrane level. In accordance with Shelford's law of tolerance decreasing whole animal aerobic scope characterises the onset of thermal limitation at low and high pejus thresholds (pejus=getting worse). The drop in aerobic scope of an animal indicated by falling oxygen levels in the body fluids and or the progressively limited capacity of circulatory and ventilatory mechanisms. At high temperatures, excessive oxygen demand causes insufficient oxygen levels in the body fluids, whereas at low temperatures the aerobic capacity of mitochondria may become limiting for ventilation and circulation. Further cooling or warming beyond these limits leads to low or high critical threshold temperatures (T(c)) where aerobic scope disappears and transition to an anaerobic mode of mitochondrial metabolism and progressive insufficiency of cellular energy levels occurs. The adjustments of mitochondrial densities and their functional properties appear as a critical process in defining and shifting thermal tolerance windows. The finding of an oxygen limited thermal tolerance owing to loss of aerobic scope is in line with Taylor's and Weibel's concept of symmorphosis, which implies that excess capacity of any component of the oxygen delivery system is avoided. The present study suggests that the capacity of oxygen delivery is set to a level just sufficient to meet maximum oxygen demand between the average highs and lows of environmental temperatures. At more extreme temperatures only time limited passive survival is supported by anaerobic metabolism or the protection of molecular functions by heat shock proteins and antioxidative defence. As a corollary, the first line of thermal sensitivity is due to capacity limitations at a high level of organisational complexity, i.e. the integrated function of the oxygen delivery system, before individual, molecular or membrane functions become disturbed. These interpretations are in line with the more general consideration that, as a result of the high level of complexity of metazoan organisms compared with simple eukaryotes and then prokaryotes, thermal tolerance is reduced in metazoans. A similar sequence of sensitivities prevails within the metazoan organism, with the highest sensitivity at the organismic level and wider tolerance windows at lower levels of complexity. However, the situation is different in that loss in aerobic scope and progressive hypoxia at the organismic level define the onset of thermal limitation which then transfers to lower hierarchical levels and causes cellular and molecular disturbances. Oxygen limitation contributes to oxidative stress and, finally, denaturation or malfunction of molecular repair, e.g. during suspension of protein synthesis. The sequence of thermal tolerance limits turns into a hierarchy, ranging from systemic to cellular to molecular levels.     

Cold tolerance and the regulation of cardiac performance and hemolymph distribution in Maja squinado (Crustacea: decapoda)

Elevated Mg(2+) levels in the hemolymph ([Mg(2+)](HL)) of brachyuran crabs have recently been demonstrated to limit cold tolerance by reducing motor and circulatory activity. Therefore, the limiting function of elevated [Mg(2+)](HL) on circulatory performance and arterial hemolymph flow was investigated by the pulsed-Doppler technique in the spider crab Maja squinado during progressive cooling from 12 degrees to 0 degrees C. [Mg(2+)](HL) were reduced from control levels of 39.9 mmol L(-1) to levels of 6.1 mmol L(-1) by incubation in magnesium reduced seawater. At 12 degrees C cardiac output was 13.9+/-2.4 mL kg(-1) min(-1) and stroke volume 0.2+/-0.04 mL kg(-1) min(-1) in control animals. In [Mg(2+)](HL)-reduced animals cardiac output increased to 43.6+/-5.0 mL kg(-1) min(-1) and stroke volume rose to 0.6+/-0.1 mL kg(-1) min(-1). Temperature reduction in control animals revealed a break point at 8 degrees C linked to a major redirection of hemolymph flow from lateral to sternal and hepatic arteries. Cardiac output and heart rate dropped sharply during cooling until transiently constant values were reached. Further heart rate reduction occurred below 4.5 degrees C. Such a plateau was not detected in [Mg(2+)](HL)-reduced animals where the break point decreased to 6 degrees C, also indicated by a sharp drop in heart rate and cardiac output and the redirection of hemolymph flow. It is concluded that progressive cooling brings the animals from a temperature range of optimum cardiac performance into a deleterious range when aerobic scope for activity falls before critical temperatures are reached. Reduction of [Mg(2+)](HL) shifts this transition to lower temperatures. These findings support a limiting role for [Mg(2+)](HL) in thermal tolerance.     

Oxygen-limited thermal tolerance in Antarctic fish investigated by MRI and (31)P-MRS

The hypothesis of an oxygen-limited thermal tolerance was tested in the Antarctic teleost Pachycara brachycephalum. With the use of flow-through respirometry, in vivo (31)P-NMR spectroscopy, and MRI, we studied energy metabolism, intracellular pH (pH(i)), blood flow, and oxygenation between 0 and 13 degrees C under normoxia (PO(2): 20.3 to 21.3 kPa) and hyperoxia (PO(2): 45 kPa). Hyperoxia reduced the metabolic increment and the rise in arterial blood flow observed under normoxia. The normoxic increase of blood flow leveled off beyond 7 degrees C, indicating a cardiovascular capacity limitation. Ventilatory effort displayed an exponential rise in both groups. In the liver, blood oxygenation increased, whereas in white muscle it remained unaltered (normoxia) or declined (hyperoxia). In both groups, the slope of pH(i) changes followed the alpha-stat pattern below 6 degrees C, whereas it decreased above. In conclusion, aerobic scope declines around 6 degrees C under normoxia, marking the pejus temperature. By reducing circulatory costs, hyperoxia improves aerobic scope but is unable to shift the breakpoint in pH regulation or lethal limits. Hyperoxia appears beneficial at sublethal temperatures, but no longer beyond when cellular or molecular functions become disturbed.     

Thermal physiology of the common eelpout (Zoarces viviparus)

We investigated the temperature dependence of some physiological parameters of common eelpout (Zoarces viviparus) from different locations (North Sea, Baltic Sea and Norwegian Sea) on acclimation temperature (3 degrees C and 12 degrees C) and acute temperature variation. The lethal limit of 12 degrees C-acclimated eelpout was determined as the critical thermal maximum [loss of equilibrium (LE) and onset of muscular spasms (OS)] and it was found to be 26.6 degrees C for LE and 28.8 degrees C for OS for all populations. However, these parameters do not have any relevant ecological interpretation. We therefore investigated the effect of gradually increased water temperature on standard metabolic rate (measured as resting oxygen consumption Mo2) and critical oxygen concentration ([O2]c) of eelpouts. Acclimation to low temperature (3 degrees C) resulted in partial compensation of Mo2, paralleled by a decrease of activation energy for Mo2 (from 82 kJ mol(-1) at 12 degrees C to about 50 kJ mol(-1) at 3 degrees C) in North Sea and Baltic Sea eelpouts. At the same time, Norwegian eelpout showed no acclimation of oxygen demand to warm temperature (12 degrees C) at all. The scope for eelpout aerobic metabolism shrank considerably with increased acclimation temperature, as [O2]c approached water oxygen concentrations. At 22.5+/-1 degrees C the [O2]c reached air saturation, which is equivalent to the upper critical temperature (TcII) and at this temperature the aerobic scope for the metabolism completely disappeared. In line with previous insight, the comparative analysis of the temperature dependence of Mo2 of Z. viviparus from different populations suggests that a pejus (sub-critical) temperature for this species is about 13-15 degrees C. In conclusion, the capacity to adjust aerobic metabolism relates to thermal tolerance and the bio-geographical distribution of the species. Global warming would thus be likely to cause a shift in the distribution of this species to the North.     

A role for oxygen delivery and extracellular magnesium in limiting cold tolerance of the sub-antarctic stone crab Paralomis granulosa?

A low capacity for regulation of extracellular Mg(2+) has been proposed to exclude reptant marine decapod crustaceans from temperatures below 0°C and thus to exclude them from the high Antarctic. To test this hypothesis and to elaborate the underlying mechanisms in the most cold-tolerant reptant decapod family of the sub-Antarctic, the Lithodidae, thermal tolerance was determined in the crab Paralomis granulosa (Decapoda, Anomura, Lithodidae) using an acute stepwise temperature protocol (-1°, 1°, 4°, 7°, 10°, and 13°C). Arterial and venous oxygen partial pressures (Po(2)) in hemolymph, heartbeat and ventilation beat frequencies, and hemolymph cation composition were measured at rest and after a forced activity (righting) trial. Scopes for heartbeat and ventilation beat frequencies and intermittent heartbeat and scaphognathite beat rates at rest were evaluated. Hemolymph [Mg(2+)] was experimentally reduced from 30 mmol L(-1) to a level naturally observed in Antarctic caridean shrimps (12 mmol L(-1)) to investigate whether the animals remain more active and tolerant to cold (-1°, 1°, and 4°C). In natural seawater, righting speed was significantly slower at -1° and 13°C, compared with acclimation temperature (4°C). Arterial and venous hemolymph Po(2) increased in response to cooling even though heartbeat and ventilation beat frequencies as well as scopes decreased. At rest, ionic composition of the hemolymph was not affected by temperature. Activity induced a significant increase in hemolymph [K(+)] at -1° and 1°C. Reduction of hemolymph [Mg(2+)] did not result in an increase in activity, an increase in heartbeat and ventilation beat frequencies, or a shift in thermal tolerance to lower temperatures. In conclusion, oxygen delivery in this cold-water crustacean was not acutely limiting cold tolerance, and animals may have been constrained more by their functional capacity and motility. In contrast to earlier findings in temperate and subpolar brachyuran crabs, these constraints remained insensitive to changing Mg(2+) levels.     

Thermal acclimation is not necessary to maintain a wide thermal breadth of aerobic scope in the common killifish (Fundulus heteroclitus)

Loss of aerobic scope at high and low temperatures is a physiological mechanism proposed to limit the thermal performance and tolerance of organisms, a theory known as oxygen- and capacity-limited thermal tolerance (OCLTT). Eurythermal organisms maintain aerobic scope over wide ranges of temperatures, but it is unknown whether acclimation is necessary to maintain this breadth. The objective of this study was to examine changes in aerobic scope in Fundulus heteroclitus, a eurythermal fish, after acclimation and acute exposure to temperatures from 5° to 33°C. The range of temperatures over which aerobic scope was nonzero was similar in acclimated and acutely exposed fish, suggesting that acclimation has modest effects on the thermal breadth of aerobic scope. However, in acclimated fish, there was a clear optimum temperature range for aerobic scope between 25° and 30°C, whereas aerobic scope was relatively constant across the entire temperature range with acute temperature exposure. Therefore, the primary effect of acclimation was to increase aerobic scope between 25° and 30°C, which paradoxically resulted in a narrower temperature range of optimal performance in acclimated fish compared to acutely exposed fish. There was only weak evidence for correlations between the thermal optimum of aerobic scope and the thermal optimum of measures of performance (specific growth rate and gonadosomatic index), and indicators of anaerobic metabolism (lactate accumulation and lactate dehydrogenase activity) only increased at high temperatures. Together these data fit many, but not all, of the predictions made by OCLTT.     

Cadmium-dependent oxygen limitation affects temperature tolerance in eastern oysters (Crassostrea virginica Gmelin)

Marine ectotherms, including oysters are exposed to variable environmental conditions in coastal shallow waters and estuaries. In the light of global climate change, additional stressors like pollution might pose higher risk to populations. On the basis of the concept of oxygen- and capacity-limited thermal tolerance in aquatic ectotherms (40), we show that a persistent pollutant, cadmium, can have detrimental effects on oysters (Crassostrea virginica). During acute warming from 20 to 28 degrees C (4 degrees C/48 h) standard metabolic rate (SMR) rose in control and cadmium-exposed (50 microg Cd2+/l) animals, with a consistently higher SMR in Cd-exposed oysters. Additionally, Cd-exposed oysters showed a stronger temperature-dependent decrease in hemolymph oxygen partial pressures. This observation indicates that the effect of temperature on aerobic metabolism was exacerbated due to the additional Cd stress. The oxygen delivery systems could not provide enough oxygen to cover Cd-induced elevated metabolic demands at high temperatures. Interestingly, cardiac performance (measured as the heart rate and hemolymph supply to tissues) rose to a similar extent in control and Cd-exposed oysters with warming indicating that cardiac output was unable to compensate for elevated energy demand in Cd-exposed oysters. Together with the literature data on metal-induced reduction of ventilatory capacity, these findings suggest that synergistic effects of elevated temperatures and cadmium exposure led to oxygen limitation by impaired performance in oxygen supply through ventilation and circulation. Overall, cadmium exposure resulted in progressive hypoxemia in oysters at high temperatures, suggesting that the thermal tolerance window is narrowed in marine ectotherms inhabiting polluted areas compared with pristine environments.     

The acute temperature tolerance of green sturgeon (Acipenser medirostris) and the effect of environmental salinity

We investigated the effect of environmental salinity on the upper thermal tolerance of green sturgeon (Acipenser medirostris), a threatened species whose natural habitat is vulnerable to temperature and salinity variation as a result of global climate change. Freshwater (FW)-reared sturgeon were gradually acclimated to salinities representing FW, estuary water (EST), or San Francisco Bay water (BAY) at 18 degrees C, and their critical thermal maximum (CTMax) was measured by increasing temperature 0.3 degrees C/min until branchial ventilation ceased. CTMax was 34.2+/-0.09 degrees C in EST-acclimated fish, with FW- and BAY-acclimated fish CTMax at 33.7+/-0.08 and 33.7+/-0.1 degrees C, respectively. Despite the higher CTMax in EST-acclimated fish, FW-acclimated sturgeon ventilation rate reached a peak that was 2 degrees C higher than EST- and BAY-acclimated groups and had a greater range of temperatures within which they exhibited normal ventilatory function as assessed by Q(10) calculation. The osmoregulatory consequences of exposure to near-lethal temperatures were assessed by measuring plasma osmolality and hematocrit, as well as white muscle, brain, and heart tissue water contents. Hematocrit was increased following CTMax exposure, most likely owing to the elevated metabolic demands of temperature increase, and plasma osmolality was significantly increased in EST- and BAY-acclimated fish, which was likely the result of a greater osmotic gradient across the gill as metabolism increased. To our knowledge, this represents the first evidence for an effect of salinity on the upper thermal tolerance of sturgeon, as well as the first investigation of the osmoregulatory consequences of exposure to near-lethal temperatures. J. Exp. Zool. 309A:477-483, 2008. (c) 2008 Wiley-Liss, Inc.     

Heat shock protein induction in the freshwater prawn Macrobrachium malcolmsonii: acclimation-influenced variations in the induction temperatures for Hsp70

The intracellular build-up of thermally damaged proteins following exposure to heat stress results in the synthesis of heat shock proteins (Hsps). In the present study, the upper thermal tolerance and expression of heat shock protein 70 (Hsp70) were examined in juveniles of the freshwater prawn Macrobrachium malcolmsonii that had been acclimated at two different temperatures, i.e. 20 degrees C (group A) and 30 degrees C (group B), in the laboratory for 30 days. Upper thermal tolerance was determined by a standard method. For heat-shock experiments, prawns in groups A and B were exposed to various elevated temperatures for 3 h each, followed by 1 h recovery at the acclimation temperature. Endogenous levels of Hsp70 were determined in the gill, heart, hepatopancreas and skeletal muscle tissues by Western blotting analysis of one dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The critical thermal maximum (CT max) for prawns in groups A and B was 37.7+/-0.27 degrees C and 41.41+/-0.16 degrees C, respectively. In general, Western blotting analysis for Hsp70 revealed one band at the 70 kDa region, containing both constitutive (Hsc70) and inducible (Hsp70) isoforms, in the gill and heart tissues; these were not detected in the hepatopancreas and skeletal muscle tissues. The onset temperature for Hsp70 induction in both gill and heart tissues was 30 degrees C for prawns in group A and 34 degrees C for those in group B. The optimum induction temperatures (at which Hsp70 induction was maximum) were found to be 34 degrees C and 32 degrees C, respectively, in the gill and heart tissues of group A prawns, and 38 degrees C and 36 degrees C, respectively, for group B prawns. These results suggest that the temperature at which acclimation occurs influences both upper thermal tolerance and Hsp70 induction in M. malcolmsonii.     

Temperature-dependent oxygen extraction from the ventilatory current and the costs of ventilation in the cephalopod Sepia officinalis

Earlier work found cuttlefish (Sepia officinalis) ventilatory muscle tissue to progressively switch to an anaerobic mode of energy production at critical temperatures (T _c) of 7.0 and 26.8°C. These findings suggested that oxygen availability limits thermal tolerance. The present study was designed to elucidate whether it is the ventilatory apparatus that sets critical temperature thresholds during acute thermal stress. Routine metabolic rate (rmr) rose exponentially between 11 and 23°C, while below (8°C) and above (26°C) this temperature range, rmr was significantly depressed. Ventilation frequency (f _V) and mean mantle cavity pressure (MMP) followed an exponential relationship within the entire investigated temperature range (8–26°C). Oxygen extraction from the ventilatory current (EO₂) decreased in a sigmoidal fashion with temperature, falling from > 90% at 8°C to 32% at 26°C. Consequently, ventilatory minute volume (MV_V) increased by a factor of 20 from 7 to 150% body weight min⁻¹ in the same temperature interval. Increases in MMP and MV_V resulted in ventilatory muscle power output (P _out) increasing by a factor of > 80 from 0.03 to 2.4 mW kg⁻¹ animal. Nonetheless, costs for ventilatory mechanics remain below 1.5% rmr in the natural thermal window of the population (English Channel, 9–17°C), owing to very low MMPs of < 0.05 kPa driving the ventilatory stream, and may maximally rise to 8.6% rmr at 26°C. Model calculations suggest that the ventilatory system can maintain high arterial PO₂ values of > 14 kPa over the entire temperature interval. We therefore conclude that the cuttlefish ventilation system is probably not limiting oxygen transfer during acute thermal stress. Depression of rmr, well before critical temperatures are being reached, is likely caused by circulatory capacity limitations and not by fatigue of ventilatory muscle fibres.     

Effect of temperature on ethanol tolerance of a thermophilic anaerobic ethanol producer Thermoanaerobacter A10: modeling and simulation

The low ethanol tolerance of thermophilic anaerobic bacteria (<2%, v/v) is a major obstacle for their industrial exploitation for ethanol production. The ethanol tolerance of the thermophilic anaerobic ethanol-producing strain Thermoanaerobacter A10 was studied during batch tests of xylose fermentation at a temperature range of 50-70 degrees C with exogenously added ethanol up to approximately 6.4% (v/v). At the optimum growth temperature of 70 degrees C, the strain was able to tolerate 4.7% (v/v) ethanol, and growth was completely inhibited at 5.6% (v/v). A higher ethanol tolerance was found at lower temperatures. At 60 degrees C, the strain was able to tolerate at least 5.1% (v/v) ethanol. A generalized form of Monod kinetic equation proposed by Levenspiel was used to describe the ethanol (product) inhibition. The model predicted quite well the experimental data for the temperature interval 50-70 degrees C, and the maximum specific growth rate and the toxic power (n), which describes the order of ethanol inhibition at each temperature, were estimated. The toxic power (n) was 1.33 at 70 degrees C, and corresponding critical inhibitory product concentration (P(crit)) above which no microbial growth occurs was determined to be 5.4% (v/v). An analysis of toxic power (n) and P(crit) showed that the optimum temperature for combined microbial growth and ethanol tolerance was 60 degrees C. At this temperature, the toxic power (n), and P(crit) were 0.50, and 6.5% (v/v) ethanol, respectively. From a practical point of view, the model may be applied to compare the ethanol inhibition (ethanol tolerance) on microbial growth of different thermophilic anaerobic bacterial strains.     

Metabolic demand,oxygen supply,and critical temperatures in the Antarctic bivalve Laternula elliptica

Oxygen consumption (Mo(2)), heartbeat rate and form, and circulating hemolymph oxygen content were measured in relation to temperature in the large Antarctic infaunal bivalve Laternula elliptica. After elevations in temperature from 0 degrees to 3 degrees, 6 degrees, and then 9 degrees C, Mo(2) and heartbeat rate rose to new levels, whereas maximum circulating hemolymph oxygen content fell. At 0 degrees C, Mo(2) was 19.6 micromol O(2) h(-1) for a standard animal of 2-g tissue ash-free dry mass, which equates to a 8.95-g tissue dry-mass or 58.4-g tissue wet-mass animal. Elevation of metabolism following temperature change had acute Q(10) values between 4.1 and 5, whereas acclimated figures declined from 3.4 (between 0 degrees and 3 degrees C) to 2.2 (3 degrees -6 degrees C) and 1.9 (6 degrees -9 degrees C). Heartbeat rate showed no acclimation following temperature elevations, with Q(10) values of 3.9, 3.2, and 4.3, respectively. Circulating hemolymph oxygen content declined from 0 degrees to 3 degrees and 6 degrees C but stayed at a constant Po(2) (73-78 mmHg) and constant proportion ( approximately 50%) of the oxygen content of the ambient water. At 9 degrees C, Mo(2) and heartbeat rate both peaked at values 3.3 times those measured at 0 degrees C, which may indicate aerobic scope in this species. After these peaks, both measures declined rapidly over the ensuing 5 d to the lowest measured in the study, and the bivalves began to die. Hemolymph oxygen content fell dramatically at 9 degrees C to values between 2% and 12% of ambient water O(2) content and had a maximum Po(2) of around 20 mmHg. These data indicate an experimental upper lethal temperature of 9 degrees C and a critical temperature, where a long-term switch to anaerobic metabolism probably occurs, of around 6 degrees C for L. elliptica. Concurrent measures of mitochondrial function in the same species had indicated strong thermal sensitivity in proton leakage costs, and our data support the hypothesis that as temperature rises, mitochondrial maintenance costs rapidly outstrip oxygen supply mechanisms in cold stenothermal marine species.     

Metabolic and cardiorespiratory responses of summer flounder Paralichthys dentatus to hypoxia at two temperatures

To quantify the tolerance of summer flounder Paralichthys dentatus to episodic hypoxia, resting metabolic rate, oxygen extraction, gill ventilation and heart rate were measured during acute progressive hypoxia at the fish's acclimation temperature (22° C) and after an acute temperature increase (to 30° C). Mean ±s.e. critical oxygen levels (i.e. the oxygen levels below which fish could not maintain aerobic metabolism) increased significantly from 27 ± 2% saturation (2·0 ± 0·1 mg O(2) l(-1) ) at 22° C to 39 ± 2% saturation (2·4 ± 0·1 mg O(2) l(-1) ) at 30° C. Gill ventilation and oxygen extraction changed immediately with the onset of hypoxia at both temperatures. The fractional increase in gill ventilation (from normoxia to the lowest oxygen level tested) was much larger at 22° C (6·4-fold) than at 30° C (2·7-fold). In contrast, the fractional decrease in oxygen extraction (from normoxia to the lowest oxygen levels tested) was similar at 22° C (1·7-fold) and 30° C (1·5-fold), and clearly smaller than the fractional changes in gill ventilation. In contrast to the almost immediate effects of hypoxia on respiration, bradycardia was not observed until 20 and 30% oxygen saturation at 22 and 30° C, respectively. Bradycardia was, therefore, not observed until below critical oxygen levels. The critical oxygen levels at both temperatures were near or immediately below the accepted 2·3 mg O(2) l(-1) hypoxia threshold for survival, but the increase in the critical oxygen level at 30° C suggests a lower tolerance to hypoxia after an acute increase in temperature.     

Effects of temperature acclimation on cardiorespiratory performance of the Antarctic notothenioid Trematomus bernacchii

Notothenioid fishes of the Southern Ocean have evolved under cold and stable temperatures for millions of years. Due to rising temperatures in the Southern Ocean, investigating thermal limits and the capacities for inducing a temperature acclimation response in notothenioids has become of increasing interest. Here, we investigated effects of temperature acclimation on cardiorespiratory responses and cardiac and skeletal muscle energy metabolism in a benthic Antarctic notothenioid, Trematomus bernacchii. We acclimated specimens to ?1, 2 and 4.5 °C for 14 days and quantified heart rates and ventilation rates during an acute increase in temperature. Ventilation rates showed an effect of acclimation both at initial steady-state acclimation conditions and during an acute temperature increase, suggesting a partial thermal compensatory response. However, acclimation did not affect heart rates at steady-state acclimation conditions and the temperatures at which onset of cardiac arrhythmia occurred, suggesting lack of inducible thermal tolerance in cardiac performance. Citrate synthase (CS), lactate dehydrogenase (LDH) and 3-hydroxyacyl dehydrogenase activities in skeletal muscle tissues suggested acclimation-induced shifts in metabolic fuel preferences, and a marked increase in LDH activity with acclimation to 4.5 °C showed an increase in anaerobic metabolism. In heart tissue, CS and LDH activities decreased with acclimation to 4.5 °C, suggesting reduced cardiac ATP production. Overall, the data suggest a partial acclimatory response to temperature by T. bernacchii and support the hypothesis that reduced cardiac acclimatory capacity may play a role in limiting the thermal plasticity of T. bernacchii.     

Physiological ecology of aquatic overwintering in ranid frogs

In cold-temperate climates, overwintering aquatic ranid frogs must survive prolonged periods of low temperature, often accompanied by low levels of dissolved oxygen. They must do so with the energy stores acquired prior to the onset of winter. Overwintering mortality is a significant factor in their life history, occasionally reaching 100% due to freezing and/or anoxia. Many species of northern ranid frogs overwinter in the tadpole stage, which increases survival during hypoxic episodes relative to adults, as well as allowing for larger sizes at metamorphosis. At temperatures below 5 degrees C, submerged ranid frogs are capable of acquiring adequate oxygen via cutaneous gas exchange over a wide range of ambient oxygen partial pressures (PO(2)), and possess numerous physiological and behavioural mechanisms that allow them to maintain normal rates of oxygen uptake across the skin at a relatively low PO(2). At levels of oxygen near and below the critical PO(2) that allows for aerobic metabolism, frogs must adopt biochemical mechanisms that act to minimise oxygen utilisation and assist in maintaining an aerobic state to survive overwintering. These mechanisms include alterations in mitochondrial metabolism and affinity, changes in membrane permeability, alterations in water balance, and reduction in cellular electrochemical gradients, all of which lead to an overall reduction in whole-animal metabolism. Winter energetic requirements are fueled by the energy stores in liver, muscle, and fat depots, which are likely to be sufficient when the water is cold and well oxygenated. However, under hypoxic conditions fat stores cannot be utilised efficiently and glycogen stores are used up rapidly due to recruitment of anaerobiosis. Since ranid frogs have minimal tolerance to anoxia, it is untenable to suggest that they spend a significant portion of the winter buried in anoxic mud, but instead utilise a suite of behavioural and physiological mechanisms geared to optimal survival in cold, hypoxic conditions.     

Production of reactive oxygen species by isolated mitochondria of the Antarctic bivalve Laternula elliptica (King and Broderip) under heat stress

Formation of reactive oxygen species (ROS) in mitochondrial isolates from gill tissues of the Antarctic polar bivalve Laternula elliptica was measured fluorimetrically under in vitro conditions. When compared to the rates measured at habitat temperature (1 degrees C), significantly elevated ROS formation was found under temperature stress of 7 degrees C and higher. ROS formation correlated significantly with oxygen consumption in individual mitochondrial preparations over the entire range of experimental temperatures (1-12 degrees C). ROS generation per mg of mitochondrial protein was significantly higher in state 3 at maximal respiration and coupling to energy conservation, than in state 4+, where ATPase-activity is inhibited by oligomycin and only proton leakage is driving the residual oxygen consumption. The percent conversion of oxygen to the membrane permeant hydrogen peroxide amounted to 3.7% (state 3) and 6.5% (state 4+) at habitat temperature (1 degrees C), and to 7% (state 3) and 7.6% (state 4+) under experimental warming to 7 degrees C. This is high compared to 1-3% oxygen to ROS conversion in mammalian mitochondrial isolates and speaks for a comparatively low control of toxic oxygen formation in mitochondria of the polar bivalve. However, low metabolic rates at cold Antarctic temperatures keep absolute rates of mitochondrial ROS production low and control oxidative stress at habitat temperatures. Mitochondrial coupling started to fall beyond 3 degrees C, closely to pejus temperature (4 degrees C) of the bivalve. Accordingly, the proportion of state 4 respiration increased from below 30% at 1 degrees C to over 50% of total oxygen consumption at 7 degrees C, entailing reduced ADP/O ratios under experimental warming. Progressive mitochondrial uncoupling and formation of hazardous ROS contribute to bias mitochondrial functioning under temperature stress in vitro. Deduced from a pejus temperature, heat stress commences already at 5 degrees C, and is linked to progressive loss of phosphorylation efficiency, increased mitochondrial oxygen demand and elevated oxidative stress above pejus temperatures.     

Divergence in aerobic scope and thermal tolerance is related to local thermal regime in two populations of introduced Nile perch (Lates niloticus)

We tested whether thermal tolerance and aerobic performance differed between two populations of Nile perch (Lates niloticus) originating from the same source population six decades after their introduction into two lakes in the Lake Victoria basin in East Africa. We used short-term acclimation of juvenile fish to a range of temperatures from ambient to +6°C, and performed critical thermal maximum (CT_max) and respirometry tests to measure upper thermal tolerance, resting and maximum metabolic rates, and aerobic scope (AS). Across acclimation temperatures, Nile perch from the cooler lake (Lake Nabugabo, Uganda) tended to have lower thermal tolerance (i.e., CT_max) and lower aerobic performance (i.e., AS) than Nile perch from the warmer waters of Lake Victoria (Bugonga region, Uganda). Effects of temperature acclimation were more pronounced in the Lake Victoria population, with the Lake Nabugabo fish showing less thermal plasticity in most metabolic traits. Our results suggest phenotypic divergence in thermal tolerance between these two introduced populations in a direction consistent with an adaptive response to local thermal regimes.     

Critical thermal limits, temperature tolerance and water balance of a sub-Antarctic kelp fly, Paractora dreuxi (Diptera: Helcomyzidae)

Paractora dreuxi displays distinct ontogenetic differences in thermal tolerance and water balance. Larvae are moderately freeze tolerant. Mean larval onset of chill coma was -5.1 degrees C, and onset of heat stupor was 35.5 degrees C. Larval supercooling point (SCP) was -3.3 degrees C with 100% recovery, although mortality was high below -4 degrees C. Starvation caused SCP depression in the larvae. Adults were significantly less tolerant, with critical thermal limits of -2.7 and 30.2 degrees C, no survival below the SCP (-9.6 degrees C), and no change in SCP with starvation. Moderate freeze tolerance in the larvae supports the contention that this strategy is common in insects from southern, oceanic islands. Fly larvae survived desiccation in dry air for 30 h, and are thus less desiccation tolerant than most other sub-Antarctic insect larvae. Water loss rates of the adults were significantly lower than those of the larvae. Lipid metabolism did not contribute significantly to water replacement in larvae, which replaced lost body water by drinking fresh water, but not sea water. Kelp fly larvae had excellent haemolymph osmoregulatory abilities. Current climate change has led to increased temperatures and decreased rainfall on Marion Island. These changes are likely to have significant effects on P. dreuxi, and pronounced physiological regulation in larvae suggests that they will be most susceptible to such change.     

The effects of acclimation on thermal tolerance, desiccation resistance and metabolic rate in Chirodica chalcoptera (Coleoptera: Chrysomelidae)

Despite much focus on species responses to environmental variation through space and time, many higher taxa and geographic areas remain poorly studied. We report the effects of temperature acclimation on thermal tolerance, desiccation rate and metabolic rate for adult Chirodica chalcoptera (Coleoptera: Chrysomelidae) collected from Protea nerifolia inflorescences in the Fynbos Biome in South Africa. After 7 days of acclimation at 12, 19 and 25 degrees C, critical thermal maxima (mean+/-s.e.: 41.8+/-0.2 degrees C in field-fresh beetles) showed less response (<1 degrees C change) to temperature acclimation than did the onset of the critical thermal minima (0.1+/-0.2, 1.0+/-0.2 and 2.3+/-0.2 degrees C, respectively). Freezing was lethal in C. chalcoptera (field-fresh SCP -14.6 degrees C) and these beetles also showed pre-freeze mortality. Survival of 2 h at -10.1 degrees C increased from 20% to 76% after a 2 h pre-exposure to -2 degrees C, indicating rapid cold hardening. Metabolic rate, measured at 25 degrees C and adjusted by ANCOVA for mass variation, did not differ between males and females (2.772+/-0.471 and 2.517+/-0.560 ml CO2 h(-1), respectively), but was higher in 25 degrees C-acclimated beetles relative to the field-fresh and 12 degrees C-acclimated beetles. Body water content and desiccation rate did not differ between males and females and did not respond significantly to acclimation. We place these data in the context of measured inflorescence and ambient temperatures, and predict that climate change for the region could have effects on this species, in turn possibly affecting local ecosystem functioning.