The effects of fasting and cold exposure on metabolic rate and mitochondrial proton leak in liver and skeletal muscle of an amphibian, the cane toad Bufo marinus

Abstract

SUMMARY Futile cycling of protons across the mitochondrial inner membrane contributes significantly to standard metabolic rate in a variety of ectothermic and endothermic animals, but adaptations of the mitochondrial bioenergetics to different environmental conditions have rarely been studied in ectotherms. Changes in ambient temperature and nutritional status have a great effect on the physiological demands of ectothermic amphibians and may require the adjustment of mitochondrial efficiency. In order to investigate the effect of temperature and nutritional status on the mitochondrial level,we exposed male cane toads to either 10°C or 30°C and fasted half of the animals in each group. Cold exposure resulted in a fourfold reduction of the resting metabolic rate whereas nutritional status had only minor effects. The mitochondrial adjustments to each condition were observed by comparing the proton leak kinetics of isolated liver and skeletal muscle mitochondria at 25°C. In response to cold exposure, liver mitochondria showed a decrease in proton conductance while skeletal muscle mitochondria were unchanged. Additional food deprivation had minor effects in skeletal muscle, but in liver we uncovered surprising differences in energy saving mechanisms between the acclimation temperatures: in warm-acclimated toads, fasting resulted in a decrease of the proton conductance whereas in cold-acclimated toads, the activity of the respiratory chain was reduced. To investigate the molecular mechanism underlying mitochondrial proton leakage, we determined the adenine-nucleotide transporter (ANT) content, which explained tissue-specific differences in the basal proton leak, but neither the ANT nor uncoupling protein (UCP) gene expression correlated with alterations of the proton leak in response to physiological stimuli.

The Long Life of Birds: The Rat-Pigeon Comparison Revisited

“…Blood was collected into heparinised vials following cardiac puncture and centrifuged to extract plasma, which was then stored in liquid nitrogen prior to later analyses. Separate aliquots of selected tissues were placed in buffer solutions for mitochondrial extraction (see [30]) prior to determining mitochondrial ROS production (see below).…”

“…Heart, pectoral muscle and liver mitochondria were isolated by differential centrifugation as described previously [30]. The protein concentration of mitochondrial suspensions was determined by the Biuret method using bovine BSA as a standard [31].…”

“…Mitochondrial oxygen consumption was determined as described previously [30]. Oxygen consumption was measured using a Clarke-type electrode (Rank Brothers Ltd, Cambridge, UK) maintained at 37°C for the rats and at 41°C for the pigeons and calibrated with air-saturated medium [120 mM KCl, 5 mM K 2 HPO 4 , 3 mM Hepes, 1 mM EGTA, 0.3% (w/v) defatted BSA, 7 µM rotenone (to inhibit complex I of the respiratory chain), adjusted to pH 7.2], which was assumed to contain 406/381 nmol oxygen/ml, respectively [32].…”

Brown adipose tissue: physiological function and evolutionary significance

“…The uncoupling protein 1 (UCP1) is critical for BAT-mediated NST. The origin of the UCP1 gene dates back to the roots of vertebrates more than 400 million years ago (MYA) but a thermogenic role is unlikely in ectothermic vertebrates (Jastroch et al 2005Trzcionka et al 2008). During mammalian evolution, the Monotremata separated from the mammalian lineage about 170 MYA in the late Jurassic period (Warren et al 2008).…”

Significant modulation of the hepatic proteome induced by exposure to low temperature inXenopus laevis

“…Cold exposure depresses their metabolic rates and has effects on mitochondrial bioenergetics (Boutilier et al, 1997; Trzcionka et al, 2008). The African clawed frog, Xenopus laevis (Daudin), has served as a crucial vertebrate model for biological research.…”