Oxidative activity of poikilotherm mitochondria as a function of temperature

In Newell & Northcroft (1967) evidence was presented which showed that the metabolism of certain intertidal invertebrates at rest does not vary greatly with temperature change. The following work was undertaken to determine whether temperature-independent metabolism could be demonstrated in suspensions of mitochondria extracted from poikilotherm tissue and whether it could be inferred that there was any difference in the ability of poikilotherms from different habitats to adjust to short-term temperature change.
It was found that the rate of reaction/temperature curve of mitochondria isolated from the skate Raia had a flattened region below 10C but above this the rate of oxidation of succinate and pyruvate increased regularly with temperature. The curve for intertidal forms had a gradual slope extending as high as 20C in the mussel Mytilus edulis while in the terrestrial snail Helix aspersa the gradual slope reached 27.5C after which the rate of oxidation of substrate increased sharply before declining towards higher temperatures. Finally, the curve for mitochondria isolated from the desert locust Schistocerca gregaria showed a gradual slope up to 35C after which a sharp increase in the rate of oxidation of substrate occurred. Thus the extent of the gradual slope corresponds with the normal environmental temperature range to which the tissues of the poikilotherms are subjected and the general form of the R-T curve is such that rapid fluctuations within the normal environmental temperature range have relatively little effect on the rate of mitochondrial activity.     

Longevity of animals under reactive oxygen species stress and disease susceptibility due to global warming

The world is projected to experience an approximate doubling of atmospheric CO_2 concentration in the next decades. Rise in atmospheric CO_2 level as one of the most important reasons is expected to contribute to raise the mean global temperature 1.4 ℃-5.8 ℃ by that time. A survey from 128 countries speculates that global warming is primarily due to increase in atmospheric CO_2 level that is produced mainly by anthropogenic activities. Exposure of animals to high environmental temperatures is mostly accompanied by unwanted acceleration of certain biochemical pathways in their cells. One of such examples is augmentation in generation of reactive oxygen species(ROS) and subsequent increase in oxidation of lipids, proteins and nucleic acids by ROS. Increase in oxidation of biomolecules leads to a state called as oxidative stress(OS). Finally, the increase in OS condition induces abnormality in physiology of animals under elevated temperature. Exposure of animals to rise in habitat temperature is found to boost the metabolism of animals and a very strong and positive correlation exists between metabolism and levels of ROS and OS. Continuous induction of OS is negatively correlated with survivability and longevity and positively correlated with ageing in animals. Thus, it can be predicted that continuous exposure of animals to acute or gradual rise in habitat temperature due to global warming may induce OS, reduced survivability and longevity in animals in general and poikilotherms in particular. A positive correlation between metabolism and temperature in general and altered O_2 consumption at elevated temperature in particular could also increase the risk of experiencing OS in homeotherms. Effects of global warming on longevity of animals through increased risk of protein misfolding and disease susceptibility due to OS as the cause or effects or both also cannot be ignored. Therefore, understanding the physiological impacts of global warming in relation to longevity of animals will become very crucial challenge to biologists of the present millennium.     

Respiration of mitochondria isolated from tulip bulbs stored at 5 and 17°C

For the production of good quality flowers, tulip ( Tulipus gesneriana L.) bulbs need a period of low temperature. In cultivar Apeldoom a treatment of 12 weeks at 5 C can be used. In the bulb scales the respiratory metabolism has to adjust to this low temperature. Mitochondria isolated from the bulb scales are able to use succinate. NADH and pyruvate as respiratory substrates. Respiratory characteristics of these mitochondria changed after transfer of the bulbs to 5°C and the adaptation was complete within 2 weeks. Both state 3 respiration and respiratory control increased. Alternative pathway capacity was constitutively present at both 5°C and 17°C: it was low and substrate dependent at both temperatures. During the following weeks of the treatment no significant changes took place. However, when bulbs were transferred to 17°C after storage at 5°C. various responses could be demonstrated. In bulbs only cooled for a short period no readjustment to this higher temperature occurred. In bulbs stored for longer periods the change depended on the duration of the 5°C treatment. The nature of the re-adaptation is discussed     

A re-interpretation of the effect of temperature on the metabolism of certain marine invertebrates

Measurements by nieans of an oxygen electrode of the rate of oxygen consumption of a wide variety of common intertidal invertebrates show that at least two rates of uptake can be distinguished in the intact animal. First, there is a rapid rate of oxygen uptake corresponding with activity and second there is a slower rate which corresponds with periods of quiescence and which alternates with the fast rate. Intermediate rates correspond with intermediate rates of overt activity. The maximal and minimal rates of oxygen uptake are affected differently by temperature change. The fast rate corresponding to activity ("active metabolism") increases with temperature in approximate agreement with Arrhenius's law. The slow rate corresponding to quiescence ("maintenance metabolism") does not vary with temperature over much of the range 7 to 22.5°C. It is concluded that, contrary to common belief, the intertidal invertebrates studied have a "basal metabolic rate" with a Q₁₀ of approximately one over much of the normal environmental temperature range and in this respect are well suited to life in a habitat where rapid fluctuations in temperature occur.     

Changes in parameters of the plasmalemma ATPase during cold acclimation of apple (Mains domestica) tree bark tissues

A plasmalemma fraction was isolated from homogenized apple tree (Mains domestica Borkh 'Golden Delicious') hark tissues using aqueous phase partitioning and ultra-centrifugation. Results of marker enzyme assays indicated that a membrane preparation highly enriched in plasma membranes was obtained. ATPase activity in this preparation possessed a high specificity for ATP as substrate, was inhibited by vanadate, diethylstilbcsterol and dicyclohexylcarbocHimide, and was insensitive to inhibitors of mitochondria! and tonoplasl ATPases. Specific activity of the plasma-lemma ATPase increased during cold acclimation prior to the attainment of vegetative maturity. Kinetic parameters (Km, Vln) determined from assays performed at different temperatures (10, 30°C) indicated a differential effect of cold acclimation on enzyme activity. Vm increased during cold acclimation, whereas Km increased when determined at 30°C but declined at 10°C. Acclimation treatments during April and May resulted in alteration of ATPase kinetics in the absence of any increase in bark frost hardiness. Changes in ATPase kinetics may be related more to enhanced low temperature metabolism than to frost hardiness per se.     

Tissue-specific depression of mitochondrial proton leak and substrate oxidation in hibernating arctic ground squirrels

A significant proportion of standard metabolic rate is devoted to driving mitochondrial proton leak, and this futile cycle may be a site of metabolic control during hibernation. To determine if the proton leak pathway is decreased during metabolic depression related to hibernation, mitochondria were isolated from liver and skeletal muscle of nonhibernating (active) and hibernating arctic ground squirrels (Spermophilus parryii). At an assay temperature of 37 degrees C, state 3 and state 4 respiration rates and state 4 membrane potential were significantly depressed in liver mitochondria isolated from hibernators. In contrast, state 3 and state 4 respiration rates and membrane potentials were unchanged during hibernation in skeletal muscle mitochondria. The decrease in oxygen consumption of liver mitochondria was achieved by reduced activity of the set of reactions generating the proton gradient but not by a lowered proton permeability. These results suggest that mitochondrial proton conductance is unchanged during hibernation and that the reduced metabolism in hibernators is a partial consequence of tissue-specific depression of substrate oxidation.     

Mitochondrial metabolic suppression and reactive oxygen species production in liver and skeletal muscle of hibernating thirteen-lined ground squirrels

During hibernation, animals cycle between periods of torpor, during which body temperature (T(b)) and metabolic rate (MR) are suppressed for days, and interbout euthermia (IBE), during which T(b) and MR return to resting levels for several hours. In this study, we measured respiration rates, membrane potentials, and reactive oxygen species (ROS) production of liver and skeletal muscle mitochondria isolated from ground squirrels (Ictidomys tridecemlineatus) during torpor and IBE to determine how mitochondrial metabolism is suppressed during torpor and how this suppression affects oxidative stress. In liver and skeletal muscle, state 3 respiration measured at 37°C with succinate was 70% and 30% lower, respectively, during torpor. In liver, this suppression was achieved largely via inhibition of substrate oxidation, likely at succinate dehydrogenase. In both tissues, respiration by torpid mitochondria further declined up to 88% when mitochondria were cooled to 10°C, close to torpid T(b). In liver, this passive thermal effect on respiration rate reflected reduced activity of all components of oxidative phosphorylation (substrate oxidation, phosphorylation, and proton leak). With glutamate + malate and succinate, mitochondrial free radical leak (FRL; proportion of electrons leading to ROS production) was higher in torpor than IBE, but only in liver. With succinate, higher FRL likely resulted from increased reduction state of complex III during torpor. With glutamate + malate, higher FRL resulted from active suppression of complex I ROS production during IBE, which may limit ROS production during arousal. In both tissues, ROS production and FRL declined with temperature, suggesting ROS production is also reduced during torpor by passive thermal effects.     

The influence of some ecological factors on the metabolism—temperature curve of the larvae of Limnephilus rhombicus (Trichoptera, Limnephilidae)

SUMMARY. The oxygen uptake of Limnephilus rhombicus , a typical inhabitant of lentic waters, was measured in three different experimental situations: in still water with a plastic mesh as a substrate, in stirred water with a plastic mesh and in still water without any substrate. Water flow caused a small increase in oxygen uptake at all temperatures but principally above 15°C. The presence or absence of the plastic substrate had little influence on oxygen uptake; L. rhombicus can be active over the whole range of temperatures studied but its activity is always small. Its metabolism—temperature curve stands out in having a plateau between 5 and 10°C, with a steeper part between 10 and 20°C, the slope of which varies according to the conditions. Maximum oxygen uptake at high temperatures was relatively low (1235 ± 153 μl O₂g dry wt-1h-1 at 25°C).
The results are compared with those obtained previously for the larvae of Microptema testacea which inhabit flowing zones in brooks. L. rhombicus larvae appear both as cold-water animals and as animals from highly variable biotopes. They are in fact very ubiquitous and occupy all lentic biotopes from cold and mountainous districts to temperate lowlands.     

Regulation of nonphosphorylating electron transport pathways in soybean cotyledon mitochondria and its implications for fat metabolism

The respiration of mitochondria isolated from germinating soybean cotyledons was strongly resistant to antimycin and KCN. This oxygen uptake was not related to lipoxygenase which was not detectable in purified mitochondria. The antimycin-resistant rate of O₂ uptake was greatest with succinate as substrate and least with exogenous NADH. Succinate was the only single substrate whose oxidation was inhibited by salicyl hydroxamic acid alone, indicating engagement of the alternative oxidase. Concurrent oxidation of two or three substrates led to greater involvement of the alternative oxidase. Despite substantial rotenone-resistant O₂ uptake with NAD-linked substrates, respiratory control was observed in the presence of antimycin, indicating restriction of electron flow through complex I. Addition of succinate to mitochondria oxidizing NAD-linked substrates in state four stimulated O₂ uptake substantially, largely by engaging the alternative oxidase. We suggest that these properties of soybean cotyledon mitochondria would enable succinate received from the glyoxysome during lipid metabolism to be rapidly oxidized, even under a high cytosolic energy charge.     

SEASONALITY AND THERMAL ACCLIMATION OF REACTIVE OXYGEN METABOLISM IN FUCUS VESICULOSUS (PHAEOPHYCEAE)

The intertidal brown macroalga Fucus vesiculosus L. acclimates its defense against reactive oxygen in response to both (1) growth at different temperatures in laboratory culture and (2) seasonal changes in environmental conditions. Fucus vesiculosus was grown in seawater at 0° C, 20° C, and at 0° C with a 3-h daily emersion at −10° C. Algae grown at low temperature, both with and without freezing, produced less reactive oxygen after severe freezing stress than those grown at 20° C. These differences were correlated with growth temperature-induced changes in activities of superoxide dismutase (SOD), glutathione reductase, and ascorbate peroxidase. The contents of tocopherols increased with increased cultivation temperature, whereas the activity of catalase and the content of glutathione and ascorbate did not change. Growth at 0° C increased the resistance of photosynthesis to freezing and reduced photoinhibition in high light at 5° C; the latter effect was further increased in algae subject to daily freezing. These data suggest that elevated activity of reactive oxygen scavenging enzymes, especially SOD, increases the resistance to photoinhibition, at least at low temperature, as well as being important for freezing tolerance. Seasonal changes in reactive oxygen metabolism showed a similar pattern to those elicited by temperature in laboratory culture. Summer samples had lower activities of most reactive oxygen scavenging enzymes than algae collected in autumn and winter when water temperatures were lower. In contrast to the laboratory experiments, ascorbate content did change and was lower during the winter than summer, whereas the content of glutathione was not influenced by season. Overall, the data not only indicate that temperature plays an important role in the regulation of stress tolerance and reactive oxygen metabolism but also suggest that other factors are also involved.     

MORPHOLOGY OF ISOLATED RABBIT HEART MUSCLE MITOCHONDRIA AND THE OXIDATION OF EXTRAMITOCHONDRIAL REDUCED DIPHOSPHOPYRIDINE NUCLEOTIDE

The morphology of rabbit heart muscle mitochondria isolated in several media has been compared by electron microscopy. The internal structure of isolated mitochondria differs from that of in situ mitochondria, with the type and degree of alteration depending on the isolation medium. Examination of the isolated mitochondria after incubation revealed that additional morphological changes occurred during incubation, but these changes were less pronounced when the incubation was conducted in a complete medium containing substrate. The isolated mitochondria have been shown to be capable of catalyzing a slow aerobic oxidation of extramitochondrial reduced diphosphopyridine nucleotide. The rate of DPNH oxidation observed is sufficient to account for the ability of the mitochondria to oxidize lactate in the presence of catalytic amounts of DPNH. The suspensions used were essentially free of mitochondrial fragments, which are known to oxidize DPNH. Possible relationships of these findings to metabolism in situ are discussed. The results indicate the desirability of correlating biochemical activities with the morphology of isolated mitochondria.     

Top-down control analysis of the effect of temperature on ectotherm oxidative phosphorylation

Top-down control and elasticity analysis was conducted on mitochondria isolated from the midgut of the tobacco hornworm (Manduca sexta) to assess how temperature affects oxidative phosphorylation in a eurythermic ectotherm. Oxygen consumption and protonmotive force (measured as membrane potential in the presence of nigericin) were monitored at 15, 25, and 35 degrees C. State 4 respiration displayed a Q(10) of 2.4-2.7 when measured over two temperature ranges (15-25 degrees C and 25-35 degrees C). In state 3, the Q(10)s for respiration were 2.0 and 1.7 for the lower and higher temperature ranges, respectively. The kinetic responses (oxygen consumption) of the substrate oxidation system, proton leak, and phosphorylation system increased as temperature rose, although the proton leak and substrate oxidation system showed the greatest thermal sensitivity. Whereas there were temperature-induced changes in the activities of the oxidative phosphorylation subsystems, there was no change in the state 4 membrane potential and little change in the state 3 membrane potential. Top-down control analysis revealed that control over respiration did not change with temperature. In state 4, control of respiration was shared nearly equally by the proton leak and the substrate oxidation system, whereas in state 3 the substrate oxidation system exerted over 90% of the control over respiration. The proton leak and phosphorylation system account for <10% of the temperature-induced change in the state 3 respiration rate. Therefore, when the temperature is changed, the state 3 respiration rate is altered primarily because of temperature's effect on the substrate oxidation system.     

Isolation and characterisation of a novel extremely thermophilic, anaerobic, chemo-organotrophic eubacterium

Abstract A new, extremely thermophilic, anaerobic, chemo-organotrophic bacterium was isolated from intertidal habitats where seepage of geothermally heated water occurs. The antibiotic sensitivity pattern and the presence of muramic acid strongly suggest an eubacterial nature of the novel isolate. Growth was measured between pH 4.8–8.2 (optimal pH 7.0) and at temperatures up to 90°C with a doubling time of 50 min at optimal temperatures of 80–85°C. This is the highest optimal growth temperature for an eubacterium described so far.
The Gram-negative, non-motile, non-sporulating, short rod to coccal shaped cells were enclosed in a sphere-like cell envelope protruding from either end. A wide range of carbohydrates, including xylose, glucose, fructose, maltose, starch, carboxymethylcellulose, and amylopectin were used in an obligately fermentative metabolism.
Morphological, physiological and molecular properties (mol% G + C = 46) are distinct from other known extremely thermophilic eubacterial genera.     

Effects of hypothermia on canine kidney mitochondria

The effect of hypothermia on the function of isolated dog kidney cortex mitochondria was determined with an FAD- and NAD⁺-linked substrate. In dog kidney mitochondria, temperatures of 10 °C or less suppress ADP stimulation of respiration but have little or no effect upon uncoupler, Ca²⁺ or valinomycin-K⁺ stimulation of respiration. This suggests that the adenine nucleotide translocase which catalyses the transport of ADP into the mitochondria limits the rate of respiration and generation of ATP at 10 °C in kidneys undergoing preservation. The coupling of oxidation to phosphylation, as determined by measuring the amount of ATP formed at low temperatures, indicates, however, that mitochondria are fully coupled at both 10 and 5 °C. The respiratory control index at 15 °C is greater (with pyruvate plus malate) than at 30 or 10 °C and suggests that 15 °C may be the optimum perfusion temperature for maintaining adenine nucleotide levels in the perfused kidney.     

Effects of temperature on the rate of development of eggs of a planktonic water mite, Piona exigua

SUMMARY. 1. The eggs of a planktonic, predatory water mile Piona exigua Viets were reared in the laboratory at constant temperatures ranging from 10°C to 280C. Egg development rates increased with increase in temperature.
2. Five models describing the effect of temperature on egg development times were fitted to the data.
3. The variances of the egg hatching times at each temperature differed widely, but these variances were not made more equal by In-transformation.
4. A simple power curve, fitted to the raw data, accounted for ≥.97% of the variation in the data. Two further models, both of which included a third parameter, provided a better fit at the temperature extremes.
5. Close examination of the patterns of variance at difference temperatures may provide information relating to the pattern of emergence of aquatic poikilotherms.
6. At temperatures below 230C. eggs of Piona exigua developed more rapidly than eggs of Piona constricta , from Heney Lake. Quebec (Riessen, 1982)     

Impact of temperature on growth and metabolic efficiency of Penicillium roqueforti– correlations between produced heat, ergosterol content and biomass

The influence of temperature on the growth of the mould Penicillium roqueforti growing on malt extract agar was studied by correlating the produced heat (measured by isothermal calorimetry), ergosterol content (quantified by GC-MS/MS) and biomass of the mould at 10, 15, 20, 25 and 30°C. The results were analysed with a simple metabolic model from which the metabolic efficiency was calculated. The results show that the impact of temperature on growth rate and metabolic efficiency are different: although the mould fungus had the highest growth rate (in terms of thermal power, which was continuously measured) at 25°C, the substrate carbon conversion efficiency (biomass production divided by substrate consumption, both counted as moles carbon) was the highest at 20°C. The temperature of the most rapid growth did not therefore equal the temperature of the most efficient growth.     

Short-term effects of triiodothyronine on the bowfin, Amia calva (Holostei), and the lake char, Salvelinus namaycush (Teleostei).

To assess the role of triiodothyronine (T3) in mediating short-term changes in metabolism, such as those occurring in circadian patterns, we examined the effects of intraperitoneal injection of T3 on the oxidation of substrates by isolated mitochondria from liver of the bowfin, Amia calva, and red muscle and liver of the lake char, Salvelinus namaycush. Selected enzymes were measured in red muscle and liver of the lake char. Three hours after intraperitoneal injection of T3, oxidation of some substrates by mitochondria isolated from the liver of the bowfin was reduced. Similar treatment had no effect on substrate oxidation in liver mitochondria isolated from lake char. Oxidation of substrates by lake char red muscle mitochondria was stimulated by T3 injection. Citrate synthase levels were increased in red muscle suggesting that changes in enzyme activity may be in part responsible for the short-term mitochondrial responses to T3 injection.     

The relationship between respiration rate and peroxidase activities in maize root mitochondria

In the present study we provide the evidence of different respiration rates and peroxidase activities in maize (Zea mays L.) mitochondria isolated from germinated seeds and roots of 2-week-old seedlings. The negative relationships between mitochondrial respiration rate measured with NADH as substrate and activities of peroxidases that oxidized NADH in both oxidative and peroxidative cycles were found. The possible role of peroxidase in the regulation of reactive oxygen species metabolism in expense of NADH oxidation was hypothesized.     

Temperature reaction norms of Daphnia magna: the effect of food concentration

1. The effects of temperature and food concentration on the fitness of Daphnia magna were tested in a 4×4 factorial flow-through design. Food ranged from 0.1 to 1.0 mg C L⁻¹ and temperature ranged from 15 to 30 °C. 2. The juvenile growth rate ( g _j) was used to construct reaction norms for temperature at varying food concentrations. Two clones isolated from the same pond at different seasons did not differ with respect to their temperature responses. Reaction norms had the shape of an optimum curve with highest values around 20 °C. There was a significant temperature–food interaction as the temperature response was most pronounced when the food was not limiting. 3. Differences in fitness were a consequence of different responses of physiological parameters to food and temperature. Age and size at first reproduction, as well as egg numbers, decreased with increasing temperature and decreasing food concentration. 4. As the temperature effect was strongest at the highest food concentrations, it can be concluded that environmental warming may affect D. magna more through a temperature rise earlier in spring rather than in summer.     

Thermal plasticity of skeletal muscle mitochondrial activity and whole animal respiration in a common intertidal triplefin fish,Forsterygion lapillum (Family: Tripterygiidae)

Oxygen demand generally increases in ectotherms as temperature rises in order to sustain oxidative phosphorylation by mitochondria. The thermal plasticity of ectotherm metabolism, such as that of fishes, dictates a species survival and is of importance to understand within an era of warming climates. Within this study the whole animal O₂ consumption rate of a common New Zealand intertidal triplefin fish, Forsterygion lapillum, was investigated at different acclimation temperatures (15, 18, 21, 24 or 25 °C) as a commonly used indicator of metabolic performance. In addition, the mitochondria within permeabilised skeletal muscle fibres of fish acclimated to a moderate temperature (18 °C Cool acclimation group—CA) and a warm temperature (24 °C. Warm acclimation group—WA) were also tested at 18, 24 and 25 °C in different states of coupling and with different substrates. These two levels of analysis were carried out to test whether any peak in whole animal metabolism reflected the respiratory performance of mitochondria from skeletal muscle representing the bulk of metabolic tissue. While standard metabolic rate (SMR- an indicator of total maintenance metabolism) and maximal metabolic rate ( \(\dot{M}\) O_{2 max}) both generally increased with temperature, aerobic metabolic scope (AMS) was maximal at 24 °C, giving the impression that whole animal (metabolic) performance was optimised at a surprisingly high temperature. Mitochondrial oxygen flux also increased with increasing assay temperature but WA fish showed a lowered response to temperature in high flux states, such as those of oxidative phosphorylation and in chemically uncoupled states of respiration. The thermal stability of mitochondria from WA fish was also noticeably greater than CA fish at 25 °C. However, the predicted contribution of respirational flux to ATP synthesis remained the same in both groups and WA fish showed higher anaerobic activity as a result of high muscle lactate loads in both rested and exhausted states. CA fish had a comparably lower level of resting lactate and took 30 % longer to fatigue than WA fish. Despite some apparent acclimation capacity of skeletal muscle mitochondria, the ATP synthesis capacity of this species is constrained at high temperatures, and that a greater fraction of metabolism in skeletal muscle appears to be supported anaerobically at higher temperatures. The AMS peak at 24 °C does not therefore represent utilisation efficiency of oxygen but, rather, the temperature where scope for oxygen flow is greatest.