Effects of increases in temperature and genipin on mitochondrial function in primary fibroblast cells of dogs of different sizes and ages.

In homeotherms, heat loss and the need for heat production are increased in small animals compared with large ones, and mitochondria are the primary organelles responsible for heat production in animals. Because of this, in animals of different body sizes, mitochondria may differ in their sensitivity to temperature and ability to produce heat via ATP uncoupling. Here, we used primary fibroblast cells isolated from large breed puppies and senior dogs, and small breed puppies and senior dogs to determine whether mitochondria across body sizes and ages in dogs function similarly, or whether smaller breeds tend to be more uncoupled and temperature sensitive due to their whole-animal phenotype of low body temperature and high mass specific metabolic rate. We compared metabolic rates in cells exposed to 37 and 40 °C to determine thermal sensitivities. We found that basal respiration and proton leak increased across all size and age classes in the 40 °C group, while coupling efficiency and spare respiratory capacity decreased, indicating that higher temperatures uncouple mitochondria in dogs, but body size does not affect temperature sensitivity. Additionally, to elucidate mechanistic differences in uncoupling, we treated cells with the UCP2 inhibitor genipin and found that genipin-treated cells increased proton leak in small senior dogs, but reduced proton leak in large puppies. The different responses to UCP2 inhibition correlate with metabolomic differences previously found in dogs of different size and age classes, and our data suggest that the role of UCP2 is dependent on the metabolic phenotype of a cell.