Combined effects of cadmium, temperature and hypoxia-reoxygenation on mitochondrial function in rainbow trout (Oncorhynchus mykiss)

•Ternary interactions of Cd, temperature and H-R alter their individual and binary effects on mitochondrial bioenergetics.•Oxidative stress explains many effects of Cd, H-R and temperature on mitochondria.•Cd accumulation does not explain increased sensitivity of mitochondria to multiple stressors.•Cd induces hormetic responses during H-R stress.•Cd at low dose blunts conversion of complex I active (A)- to deactive (D)-form after H-R.

Although aquatic organisms face multiple environmental stressors that may interact to alter adverse outcomes, our knowledge of stressor-stressor interaction on cellular function is limited. We investigated the combined effects of cadmium (Cd), hypoxia-reoxygenation (H-R) and temperature on mitochondrial function. Liver mitochondria from juvenile rainbow trout were exposed to Cd (0-20 μM) and H-R (0 and 5 min) at 5, 13 and 25 °C followed by measurements of mitochondrial Cd load, volume, complex І active (A) ↔ deactive (D) transition, membrane potential, ROS release and ultrastructural changes. At high temperature Cd exacerbated H-R-imposed reduction of maximal complex I (CI) respiration whereas at low temperature 5 and 10 μM stimulated maximal CI respiration post H-R. The basal respiration showed a biphasic response at high temperatures with low Cd concentrations reducing the stimulatory effect of H-R and high concentrations enhancing this effect. At low temperature Cd monotonically enhanced H-R-induced stimulation of basal respiration. Cd and H-R reduced both the P/O ratio and the RCR at all 3 temperatures. Temperature rise alone increased mitochondrial Cd load and toxicity, but combined H-R and temperature exposure reduced mitochondrial Cd load but surprisingly exacerbated the mitochondrial dysfunction. Mitochondrial dysfunction induced by H-R was associated with swelling of the organelle and blocking of conversion of CІ D to A form. However, low amounts of Cd protected against H-R induced swelling and prevented the inhibition of H-R-induced CI D to A transition. Both H-R and Cd dissipated mitochondrial membrane potential Δψm and damaged mitochondrial structure. We observed increased reactive oxygen species (H2O2) release that together with the protection afforded by EGTA, vitamin E and N-acetylcysteine against the Δψm dissipation suggested direct involvement of Cd and oxidative stress. Overall, our findings indicate that mitochondrial sensitivity to Cd toxicity was enhanced by the effects of H-R and temperature, and changes in mitochondrial Cd load did not always explain this effect.

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