Signals and molecular mechanisms of temperature adaptation of mitochondrial functions in marine fish

Abstract

Temperature has a large impact on the velocity of biochemical and enzymatic processes and hence is a key factor defining the performance of ectothermic organisms. In marine fish, temperature acclimation is well known to induce adjustments of mitochondrial capacities and functions. This thesis sets out to elucidate the cellular signals and molecular mechanism involved in thermal adaptation of mitochondrial functions in marine fish. The basis for these mechanistic studies was laid by the establishment of a cellular system for the common eelpout Zoarces viviparus, a model organism to investigate thermal acclimation in fish. Primary culture developed for eelpout hepatocytes supported viability and physiological integrity of the cells for several days. Adjustments of mitochondrial functions occurred in response to varying ambient conditions, reaching steady state levels within the available time period. Primary culture of eelpout hepatocytes thus provided a suitable tool to investigate the mechanism involved in thermal adaptation.On this groundwork, the involvement of systemic signals in temperature-dependent mitochondrial adjustment was investigated by monitoring mRNA expression and capacities of the mitochondrial key enzymes citrate synthase (CS) and cytochrome c oxidase (COX). In vivo temperature acclimation of Z. viviparus to 4 and 11 degrees C resulted in an increase of CS activities in hepatocytes in the cold, while COX activities and the mRNA expression of the respective genes remained unaffected. In contrast, in vitro cold incubation of liver cells from warm acclimated animals left both mitochondrial enzymes unchanged and warm incubation of hepatocytes prepared from cold acclimated fish induced a simultaneous decrease of the activities of both enzymes and a decline of COX mRNA expression. The lack of cold acclimation in isolated liver cells and the differences between warm acclimation patterns in vivo vs. in vitro indicates the involvement of (a) systemic signal(s) in the induction or modulation of thermal adaptation. Therefore, the impact of several potential effectors on thermal adaptation of mitochondrial functions was studied. Adenosine may act as such a signal, since it is known to be a general indicator for bioenergetic disturbances. In line with a potential role for this metabolite in thermal adaptation, adenosine levels in serum and liver of Z. viviparus increased within 24 h of in vivo cold incubation and were still elevated after 3 days in liver. Adenosine treatment of isolated hepatocytes caused a reduction of COX activities, but induced an increase of COX mRNA expression. These effects were not receptor mediated, suggesting a diffusive entry and intracellular action of the metabolite. Adenosine may inhibit the translation of COX mRNA resulting in reduced COX activities, which in turn may cause a compensatory increase in COX mRNA levels. Thus it may act as a modulator in thermal adaptation by removing excess COX activities during warm acclimation or preventing its build-up during cold incubation. Temperature changes might also involve a stress response in ectothermic animals. Therefore the impact of epinephrine and cortisol on mitochondrial functions was elaborated. Cortisol treatment of isolated eelpout hepatocytes increased the mRNA expression of CS and of the nuclear encoded, but not of the mitochondrial encoded, COX subunit. Enzyme activities were not affected. This resembles the situation during the early phase of cold acclimation described for Z. viviparus and suggests an involvement of cortisol in the induction of cold acclimation. The response of isolated hepatocytes to epinephrine significantly depended on the season in which the experiment was performed. The catecholamine revealed almost no effects during summer incubations, but was found to increase activities of both enzymes during experiments performed in winter. Thus a potential role for epinephrine may be restricted to acclimatisation in winter.In conclusion, the findings of the present thesis confirm the requirement of systemic signals for the induction and modulation of thermal adaptation. They furthermore indicate a role for adenosine and the stress hormones epinephrine and cortisol in specific parts of this process.