Respiration rates of ectothermic organisms are affected by environmental temperatures, and sustainable metabolism at high temperatures sometimes limits heat tolerance. Organisms are hypothesized to exhibit acclimatory metabolic compensation effects, decelerating their metabolic processes below Arrhenius expectations based on temperature alone. We tested the hypothesis that either heritable or plastic heat tolerance differences can be explained by metabolic compensation in the eurythermal freshwater zooplankton crustacean Daphnia magna. We measured respiration rates in a ramp-up experiment over a range of assay temperatures (5-37°C) in eight genotypes of D. magna representing a range of previously reported acute heat tolerances and, at a narrower range of temperatures (10-35°C), in D. magna with different acclimation history (either 10 or 25°C). We discovered no difference in temperature-specific respiration rates between heat-tolerant and heat-sensitive genotypes. In contrast, we observed acclimationspecific compensatory differences in respiration rates at both extremes of the temperature range studied. Notably, there was a deceleration of oxygen consumption at higher temperature in 25°Cacclimated D. magna relative to their 10°C-acclimated counterparts, observed in active animals, a pattern corroborated by similar changes in filtering rate and, partly, by changes in mitochondrial membrane potential. A recovery experiment indicated that the reduction of respiration was not caused by irreversible damage during exposure to a sublethal temperature. Response time necessary to acquire the respiratory adjustment to high temperature was lower than for low temperature, indicating that metabolic compensation at lower temperatures requires slower, possibly structural changes.

Introduction: Cold exposure is a challenging environmental stimulus for humans, particularly in vulnerable populations such as older adults. Decrements in cold tolerance observed in older adults may be mediated by an age-related reduction in the stress-induced cytoprotective mechanism of autophagy, which enhances cell survival by eliminating damaged cellular components. However, it is unknown how autophagy and accompanying cytoprotective pathways (i.e., heat shock proteins) respond to cold conditions in humans. Purpose: The purpose of this thesis was to evaluate the impact of aging on autophagic activity during cold exposure and assess strategies to reverse age-related autophagic dysfunction. Methods: We examined the influence of age on autophagy during acute cold exposure utilizing ex vivo, in vitro, and in vivo models in young (19-29 years) and older (54-76 years) adults. Autophagic activity in all investigations was assessed in peripheral blood mononuclear cells (i.e., immune cells) via Western blotting. Simulated hypothermic conditions (equivalent to 4-35°C core temperature) were evaluated using ex vivo whole-blood exposure. In vivo cold stress was achieved using cold-water immersions to elicit a physiologically relevant decrease in core temperature by 0.5 and 1.0°C. Techniques that potentially reversed autophagic impairments during cold exposure were assessed including, 1) an in vitro treatment of a known autophagic stimulator (rapamycin) in immune cells obtained from young and older males, and 2) an in vivo cold acclimation in young males with cold-water immersions (14°C for 60 min) repeated on 7 consecutive days. Results: Simulated hypothermia (4-35°C) induced autophagic dysfunction regardless of age or sex. Moderate cold stress (a 0.5°C reduction in core temperature) stimulated autophagy in young males. However, intense cold exposures (equivalent to ≥ 1.0°C decrease in core temperature) elicited signs of autophagic dysfunction and a shift towards apoptotic cell death. Additionally, older adults displayed evidence of autophagic dysfunction during each cold exposure, although age-related autophagic dysfunction was mitigated with acute rapamycin treatment. Further, cold acclimation robustly improved autophagic responses to cold exposure. Conclusion: Despite an observed age-related impairment in autophagic responses during cold exposure, this thesis provided the first evidence in humans that autophagic dysregulation during cold exposure can be reversed through the administration of autophagic stimulators and through cold acclimation.

No description available.

Žieminiai rapsai (Brassica napus L.) – vienas plačiausiai Lietuvoje auginamų augalų. Atsparumas šalčiui – vienas pagrindinių veiksnių, ribojančių žieminių rapsų auginimą vidutinio klimato regionuose. Žieminių rapsų atsparumo šalčiui tyrimai atlikti Lietuvos žemės ūkio universiteto Agrobiotechnologijos laboratorijoje. Tirtas žieminių rapsų veislių ‘Banjo’, ‘Elvis’, ‘Silvia’, ‘Valesca’ atsparumas šalčiui šaldant in vitro po atlydžio pakartotinai grūdintus ir negrūdintus ūglius. Visų tyrimo variantų ūgliai grūdinti 14 dienų 4 °C temperatūroje. Modeliuojant atlydžio sąlygas, grūdinti ūgliai auginti 18 °C temperatūroje 14 ir 21 dieną. Atsparumas šalčiui vertintas šaldant ūglius po 14 atlydžio dienų be pakartotinio grūdinimo ir pakartotinai grūdinant. Laboratoriniais tyrimais nustatyta, kad šaldant ūglius po 14 atlydžio dienų be pakartotinio grūdinimo žieminių rapsų atsparumas šalčiui ženkliai mažėjo. Pakartotinai grūdinant po atlydžio, šalčio nepažeistų ūglių kiekis iš esmės didėjo. Iš tirtų žieminių rapsų genotipų didžiausias nepažeistų ūglių procentas gautas šaldant veislių ‘Banjo’ ir ‘Valesca’ ūglius. Nustatytas teigiamas sacharozės poveikis šalčio nepažeistų ūglių kiekiui, tačiau tirtų žieminių rapsų atsparumą šalčiui labiau lėmė pakartotinis grūdinimas. / Winter rapeseed (Brassica napus L.) is one of the most important crops in Lithuania. Winter hardiness is a complex trait limiting cultivation of winter rapeseed to the regions of temperate climate. Cold resistance of four winter rapeseed cultivars in vitro and their ability to recover after de-acclimation and re-acclimation periods were investigated at the Laboratory of Agrobiotechnology of Lithuanian University of Agriculture. Shoots were acclimated at 4 °C for 14 days and transferred to freezing temperatures. For study influence of de-acclimation period on rapeseed cold resistance acclimated shoots were de-acclimated and subjected to freezing temperature with and without repeated acclimation. A significant decrease in frost resistance of shoots after 14 days de-acclimation period for all tested cultivars has been obtained. Repeated acclimation followed after de-acclimation period resulted increasing the shoot ability to cold re-acclimation. Cultivars ‘Banjo’ and ‘Valesca’ showed highest value of un-injured shoots under freezing treatment among tested winter rapeseed genotypes. Exogenous sucrose has positive effect on winter rapeseed cold resistance, but repeated acclimation is more important for shoot recovery after de-acclimation.

This thesis evaluated the role of Serine/Arginine-rich proteins, also known as SR proteins, in addition to LAMMER kinases in the cold acclimation response using Brassica napus and Arabidopsis thaliana. Transcription profile analyses of SR and LAMMER kinase genes in Brassica napus and BnCBF overexpressor lines showed that exposure to low temperatures led to increased transcript levels for nine SR genes and two kinases. BnCBF overexpression was found to exacerbate this response. This was associated with increases in SR protein abundance and phosphorylation status, suggesting that SR proteins play an essential role in cold acclimation. These findings prompted further studies to assess the role of SR proteins and kinases in the cold acclimation induced adjustment of photosynthesis, the acquisition of freezing tolerance and the transcriptional profile of CBF, SPS and COR genes, which play an important role in the transcriptional cascade allowing plants to undergo cold acclimation. Using Arabidopsis loss-of-function mutants of SR proteins and AME3 LAMMER kinase, it was shown that At-RSZ22 and At-SR45 are indispensable in the regulation of photosynthesis under non-acclimated and cold acclimation conditions. At-RSZ22a, At-SCL30 and At-RS41 were then proposed to play a crucial role in the cold acclimation induced adjustment of photosynthetic performance. Moreover, the deletion of At-AME3 kinase not only jeopardized the cold acclimation induced adjustment of photosynthetic performance, but also the acquisition of freezing tolerance. This was associated with attenuation of the transcription profile of key cold responsive genes and protein abundance of COR15 A/B and dehydrins. These findings prompted further physiological characterization of ame3 mutants, and the elucidation of Serine/Arginine-rich proteins capable of interacting with this LAMMER kinase of interest. Under cold stress and acclimation conditions, the deletion of At-AME3 LAMMER kinase impeded Photosystem I physiology and state-1 state-2 transitions. These findings were associated with decreases in Photosystem II and Photosystem I protein abundance. Yeast 2-hybrid assays showed that six SR proteins are capable of physically interacting with AME3. Taken together, the results of this study demonstrate that At-RSZ22 and At-SR45 are essential in the photosynthetic performance of Arabidopsis, that At-RSZ22a, At-SCL30 and At-RS41 play an essential role in the cold acclimation induced recovery of photosynthetic performance, and that At-AME3 plays an essential role in the cold acclimation response.

Wheat (Triticum aestivum L.) is occasionally exposed to low temperature during flowering and huge economic losses can occur especially in some key production countries such as Australia. Although it is generally predicted that there will be a rise in global temperature there are still predicted to be risks associated with low temperature for temperate crops. Post head emergence frost damage remains a major constraint to increasing wheat production. Five Iraqi varieties (Abu-Ghariab, Fatah, Sham6, IPA95 and IPA99) were screened for their frost hardiness (LT50). Abu-Ghariab was chosen for further investigations along with the European cv. Claire because the acclimated frost hardiness level in these two varieties (LT50 -8.07 and -8.01°C for Claire and Abu-Ghariab respectively) was found to be significantly lower than the other varieties in this study. Several techniques were employed including REC%, IR thermography and molecular analysis of cold acclimation in order to characterise the frost resistance of those two varieties. REC% revealed that both Claire and Abu-Ghariab could tolerate some freezing when the spikes were just visible (ZCK 51-60) with an acclimation shift of the LT50 of -1.6 and -2.11°C respectively but this was not apparent at later growth stages. Based on molecular analysis, cold acclimation was shown to be activated at ZCK 51-60 in both varieties. Cbf14 was expressed after 8 hours exposure to acclimatising temperatures (4°C) and then declined to a low, but still up-regulated level in both varieties and this led to expression of the COR15a protein. These molecular changes correlated with the frost tolerance recorded at ZCK 51-60. It was concluded that the possibility existed to up-regulate cold acclimation after spike emergence if there was enough environmental stimulus. Molybdenum (Mo) was demonstrated to work synergistically with low temperature in increasing the expression of Cbf14 and COR15a. The European wheat cv. Claire showed a higher capacity (-8.14°C) to be acclimated than Iraqi wheat (-7.40°C) under the effect of both Mo and acclimation temperatures. Mo alone increased the expression of Cbf14 in both varieties but did not increase the frost tolerance. Observations of ice nucleation using an infrared thermography (IR) revealed that supercooling is highly likely in spikes and some spikes avoided frost damage even when the temperature fell to -12°C. It was observed that the proportion of frozen spikes was 22.8% while the remainder supercooled. Spraying plants with distilled water was not effective in facilitating ice nucleation in wheat spikes. Observations also revealed that spikes that did freeze started freezing at temperatures of -4 to -5 °C close to temperature of the putative constitutive frost hardiness of un-acclimated wheat and it is suggested that this may reflect that many laboratory freezing experiments may not actually freeze until -5°C. The anthesis stage of wheat was found to be the most vulnerable stage and it needs to be given more attention in terms of research to up-regulate cold acclimation. Frost damage to wheat during flowering continues to be a serious problem in certain production areas and therefore continued effort in characterising and finding suitable solutions to this are imperative.

Rates and energy equivalents of consumption (C), egestion+ excretion (FU) , assimilation (A), respiration (R) and production (P) were measured in two groups of Sceloporus olivaceus: 1) a laboratory group acclimated at four seasonally encountered temperatures (15, 20, 25 and 30°C) ; and 2) four acclimatization groups collected at different seasons when ambient temperatures corresponded to 15, 20, 25 and 30°C.

ABSTRACT Phenotypic flexibility in basal metabolic rate (BMR) in response to short-term thermal acclimation was assessed in the Laughing Dove (Streptopelia senegalensis), a common resident bird species distributed throughout most of southern Africa. I hypothesised that S. senegalensis would display flexibility in BMR over short time scales and that this flexibility would be reversible. Additionally, I hypothesised BMR to be repeatable, and that changes in BMR would be correlated with changes in organ mass. I tested these hypotheses by measuring BMR in three groups of 10 birds before and after a short-term (21 day) thermal acclimation period to one of three air temperatures (10o, 22o & 35oC). After acclimation the three temperature groups were randomly divided and reverseacclimated for another 21 days to one of the two thermal environments not yet experienced. After this reverse-acclimation period BMR was measured again. The dry masses of the stomach, kidney, heart, intestines, liver and pectoral muscles of acclimated birds were used to determine possible mechanistic correlates of BMR adjustments. Additionally, by monitoring BMR every 4-6 days during cold (10oC) and heat (35oC) acclimation I was able to assess the temporal dynamics of adjustments in BMR in response to short-term thermal acclimation. BMR was both flexible and reversible in S. senegalensis as a consistent relationship between BMR and acclimation air temperature was observed after acclimation and reverse-acclimation. BMR increased with decreasing acclimation temperature. Furthermore, a significant proportion (25%) of the observed variation in BMR was repeatable in the 22oC group in spite of the change in BMR induced by thermal acclimation. The mechanistic correlate of BMR adjustment in S. senegalensis appears to be metabolic intensity and not organ size, as the only organ to show a significant increase in size was the intestine of the acclimated 10oC group, which was significantly heavier than the intestine of the 22oC group. BMR also decreases in response to the reduction of flight and/or exercise. Since this reduction was not accompanied by a correlated change in organ mass or body mass, the reduction in BMR as a response to captivity appears to be linked to metabolic intensity of the organs and skeletal muscles. In S. senegalensis adjustments in BMR occur during the first 30 days of captivity and thermal acclimation. The response in BMR to acclimation temperature is clearly evident as BMR of the heat-acclimated group was significantly lower than the coldacclimated group after 21 days. During the response period, which lasts approximately 30 days, BMR adjusts as a mechanism to offset the costs of thermoregulation and habituation to captivity while other metabolic parameters such as body mass, body temperature, and minimum wet thermal conductance adjust to captivity and the thermal environment. After 30 days BMR of the cold and heat-acclimated groups converge on 0.68W, indicating that once the associated metabolic parameters adjust and stabilize in response to the thermal environment, BMR continues to adjust to captivity.

Non-compensable cold exposure represents a potentially deadly threat to humans, as we lack highly specialized organs and mechanisms necessary to maintain our optimal core temperature of ~37°C. Repeated exposures to cold have been shown to induce protective physiological changes in cold responses through a process known as cold acclimatization (natural) or acclimation (in laboratory). The purpose of this thesis was to determine what physiological changes occur following an intense 7 day, 14°C cold water immersion acclimation protocol, during both non-compensable (Chapter 2) and compensable cold exposures (Chapter 3). This includes identifying changes in the contributions of the shivering (ST) and non-shivering (NST) thermogenic pathways to overall heat production. ST and NST changes were quantified via electromyography and indirect calorimetry, respectively. This 7 day cold water acclimation protocol resulted in a decrease in cooling rate, a significant increase in mean esophageal core temperature, a decrease in peak heart rate following immersion, and increased thermal comfort from day 1 to day 7 of the 1h 14°C cold water immersions. Further to these findings, changes in ST and NST were measured pre- and post-acclimation with a standardized compensable cold protocol using a liquid conditioned suit (LCS) which lowered Tskin to 26°C for 2.5h. The cold acclimation protocol resulted in a ~38% decrease in mean shivering over the 2.5h without any change in thermogenic rate from pre- to post-cold acclimation. In addition, no significant difference in fuel selection was observed. These results indicate that the short, intense cold acclimation protocol did result in a substantial change in the contribution of ST and NST to total heat production which could increase cold tolerance by reducing involuntary muscle contractions during ST.

Freshwater zooplankton crustacean Daphnia frequently face strong temperature fluctuations in its natural environment, which necessitates adaptive plastic responses. This study focuses on changes in lipid peroxidation and total oxidative capacity in Daphnia tissues in response to long-term and short-term temperature changes. Long-term acclimation to 28ºC helped Daphnia survive longer at lethally high temperatures. This difference, however, was not accompanied by changes in lipid peroxidation, indicating that it isn’t a good measure of damage or predictor of temperature tolerance. On the other hand, total oxidation capacity was lower 28ºC- than in 18ºC-acclimated Daphnia, suggesting that acclimation resulted in higher amounts of antioxidants in Daphnia tissues. Exposure to hypoxia, known to up-regulate antioxidant pathways in Daphnia, further elevated heat tolerance in 28ºC- acclimated individuals. Yet, manipulations of glutathione, an important antioxidant, while predictably affecting oxidative capacity, didn’t influence heat tolerance in Daphnia, suggesting that other antioxidants may play a significant role in it.