Flow cytometric determination of genome size in European sunbleak Leucaspius delineatus (Heckel, 1843)

The aim of this study was to compare DNA content in hepatocyte and erythrocyte nuclei of the European sunbleak, Leucaspius delineatus, in relation to nuclear and cell size by means of flow cytometry and fluorescence microscopy. The DNA standards, chicken and rainbow trout erythrocytes, were prepared in parallel with both cell types, with initial separation of liver cells in pepsin solution followed by cell filtering. Standards and investigated cells were stained with a mixture of propidium iodide, citric acid, and Nonidet P40 in the presence of RNAse, and fluorescence of at least 50,000 nuclei was analyzed by flow cytometry. Average cell size was determined by flow cytometry, using fresh cell suspension in relation to latex beads of known diameter. The size of nuclei was examined on the basis of digital micrographs obtained by fluorescence microscopy after nuclei staining with DAPI. The sunbleak’s erythrocyte nuclei contain 2.25 ± 0.06 pg of DNA, whereas the hepatocyte nuclei contain 2.46 ± 0.06 pg of DNA. This difference in DNA content was determined spectroscopically using isolated DNA from the two cell types. The modal diameters of the erythrocytes and hepatocytes were estimated to be 5.1 ± 0.2 and 22.3 ± 5.0 μm, respectively, and the corresponding modal dimensions of their nuclei (measured as surface area) were 15.2 and 21.4 μm2, respectively. The nucleoplasmic index, as calculated from diameters estimated from surface area of nuclear profiles, was 2.51 for the erythrocytes compared with 0.08 for hepatocytes

Voluntary wheel running prevents the acidosis-induced decrease in skeletal muscle mitochondrial reactive oxygen species emission

Decreases in pH (acidosis) in vitro can alter skeletal muscle mitochondrial function [respiration and reactive oxygen species (ROS) emission]. However, because skeletal muscles readily adapt to exercise, the effects of acidosis may be different on sedentary vs. trained muscle. The aim of this work was to compare the effects of pH on skeletal muscle mitochondrial function between sedentary vs. exercise-trained male Sprague-Dawley rats ( n = 10 in each cohort). Rates of mitochondrial respiration and ROS emission were determined from the soleus muscle of both cohorts over a physiologic range of pH values (pH 6.2-7.1). Exercise-trained rats had 14% higher mean muscle buffering capacities; 46 and 40% greater enzyme activity of citrate synthase and lactate dehydrogenase, respectively; and greater activity of respiratory complexes I-IV. ADP-stimulated respiration with complex I and II substrates was ∼25% greater in exercise-trained rats but was unaffected by pH in either cohort. In both cohorts, lowering pH decreased respiration only in complex I- and complex II-supported nonphosphorylating (leak) state. However, as pH decreased, ROS emissions in complex I- and complex II-supported leak state decreased only in sedentary rats; in exercise-trained rats, ROS emissions in this state remained constant. We hypothesize that this effect may result from modulation at complex III, which declined 47% per unit pH in sedentary rats, in comparison to 23% in exercise-trained rats. Taken together, these data suggest that pH regulates mitochondrial respiratory complexes and that exercise training can decrease the effects of pH on skeletal muscle mitochondrial function.-Hedges, C. P., Bishop, D. J., Hickey, A. J. R. Voluntary wheel running prevents the acidosis-induced decrease in skeletal muscle mitochondrial reactive oxygen species emission

Insights on the impact of mitochondrial organisation on bioenergetics in high-resolution computational models of cardiac cell architecture

Recent electron microscopy data have revealed that cardiac mitochondria are not arranged in crystalline columns but are organised with several mitochondria aggregated into columns of varying sizes spanning the cell cross-section. This raises the question-how does the mitochondrial arrangement affect the metabolite distributions within cardiomyocytes and what is its impact on force dynamics? Here, we address this question by employing finite element modeling of cardiac bioenergetics on computational meshes derived from electron microscope images. Our results indicate that heterogeneous mitochondrial distributions can lead to significant spatial variation across the cell in concentrations of inorganic phosphate, creatine (Cr) and creatine phosphate (PCr). However, our model predicts that sufficient activity of the creatine kinase (CK) system, coupled with rapid diffusion of Cr and PCr, maintains near uniform ATP and ADP ratios across the cell cross sections. This homogenous distribution of ATP and ADP should also evenly distribute force production and twitch duration with contraction. These results suggest that the PCr shuttle and associated enzymatic reactions act to maintain uniform force dynamics in the cell despite the heterogeneous mitochondrial organization. However, our model also predicts that under hypoxia activity of mitochondrial CK enzymes and diffusion of high-energy phosphate compounds may be insufficient to sustain uniform ATP/ADP distribution and hence force generation

Changes in mitochondrial function and mitochondria associated protein expression in response to 2-weeks of high intensity interval training

Purpose: High-intensity short-duration interval training (HIT) stimulates functional and metabolic adaptation in skeletal muscle, but the influence of HIT on mitochondrial function remains poorly studied in humans. Mitochondrial metabolism as well as mitochondrial-associated protein expression were tested in untrained participants performing HIT over a 2-week period. Methods: Eight males performed a single-leg cycling protocol (12 × 1min intervals at 120% peak power output, 90 s recovery, 4 days/week). Muscle biopsies (vastus lateralis) were taken pre- and post-HIT. Mitochondrial respiration in permeabilized fibers, citrate synthase (CS) activity and protein expression of peroxisome proliferator-activated receptor gamma coactivator (PGC-1!) and respiratory complex components were measured. Results: HIT training improved peak power and time to fatigue. Increases in absolute oxidative phosphorylation (OXPHOS) capacities and CS activity were observed, but not in the ratio of CCO to the electron ransport system (CCO/ETS), the respiratory control ratios (RCR-1 and RCR-2) or mitochondrial-associated protein expression. Specific increases in OXPHOS flux were not apparent after normalization to CS, indicating that gross changes mainly resulted from increased mitochondrial mass. Conclusion: Over only 2 weeks HIT significantly increased mitochondrial function in skeletal muscle independentl

Spatiotemporal and demographic variation in the diet of New Zealand lesser short-tailed bats (Mystacina tuberculata)

Variation in the diet of generalist insectivores can be affected by site‐specific traits including weather, habitat, and season, as well as demographic traits such as reproductive status and age. We used molecular methods to compare diets of three distinct New Zealand populations of lesser short‐tailed bats, Mystacina tuberculata. Summer diets were compared between a southern cold‐temperate (Eglinton) and a northern population (Puroera). Winter diets were compared between Pureora and a subtropical offshore island population (Hauturu). This also permitted seasonal diet comparisons within the Pureora population. Lepidoptera and Diptera accounted for >80% of MOTUs identified from fecal matter at each site/season. The proportion of orders represented within prey and the Simpson diversity index, differed between sites and seasons within the Pureora population. For the Pureora population, the value of the Simpson diversity index was higher in summer than winter and was higher in Pureora compared to Eglinton. Summer Eglinton samples revealed that juvenile diets appeared to be more diverse than other demographic groups. Lactating females had the lowest dietary diversity during summer in Pureora. In Hauturu, we found a significant negative relationship between mean ambient temperature and prey richness. Our data suggest that M. tuberculata incorporate a narrower diversity of terrestrial insects than previously reported. This provides novel insights into foraging behavior and ecological interactions within different habitats. Our study is the first from the Southern Hemisphere to use molecular techniques to examine spatiotemporal variation in the diet of a generalist insectivore that inhabits a contiguous range with several habitat types and climates

Changes in mitochondrial function and mitochondria associated protein expression in response to 2-weeks of high intensity interval training

Purpose: High-intensity short-duration interval training (HIT) stimulates functional and metabolic adaptation in skeletal muscle, but the influence of HIT on mitochondrial function remains poorly studied in humans. Mitochondrial metabolism as well as mitochondrial-associated protein expression were tested in untrained participants performing HIT over a 2-week period. Methods: Eight males performed a single-leg cycling protocol (12 × 1min intervals at 120% peak power output, 90 s recovery, 4 days/week). Muscle biopsies (vastus lateralis) were taken pre- and post-HIT. Mitochondrial respiration in permeabilized fibers, citrate synthase (CS) activity and protein expression of peroxisome proliferator-activated receptor gamma coactivator (PGC-1!) and respiratory complex components were measured. Results: HIT training improved peak power and time to fatigue. Increases in absolute oxidative phosphorylation (OXPHOS) capacities and CS activity were observed, but not in the ratio of CCO to the electron ransport system (CCO/ETS), the respiratory control ratios (RCR-1 and RCR-2) or mitochondrial-associated protein expression. Specific increases in OXPHOS flux were not apparent after normalization to CS, indicating that gross changes mainly resulted from increased mitochondrial mass. Conclusion: Over only 2 weeks HIT significantly increased mitochondrial function in skeletal muscle independentl