Journal
FRONTIERS IN PHYSIOLOGY
Volume 11, Issue -, Pages -Publisher
FRONTIERS MEDIA SA
DOI: 10.3389/fphys.2020.00326
Keywords
mitochondria; H+ leak pathway; uncoupling; thermogenesis; brown fat; beige fat; patch-clamp technique; fatty acid
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Funding
- NIH/NIGMS [R01GM107710, R01GM118939]
- AHA Career Development Award [19CDA34630062]
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Mitochondria convert the chemical energy of metabolic substrates into adenosine triphosphate (ATP) and heat. Although ATP production has become a focal point of research in bioenergetics, mitochondrial thermogenesis is also crucial for energy metabolism. Mitochondria generate heat due to H+ leak across the inner mitochondrial membrane (IMM) which is mediated by mitochondrial uncoupling proteins. The mitochondrial H+ leak was first identified, and studied for many decades, using mitochondrial respiration technique. Unfortunately, this method measures H+ leak indirectly, and its precision is insufficient for the rigorous insight into the mitochondrial function at the molecular level. Direct patch-clamp recording of H+ leak would have a significantly higher amplitude and time resolution, but application of the patch-clamp technique to a small subcellular organelle such as mitochondria has been challenging. We developed a method that facilitates patch-clamp recording from the whole IMM, enabling the direct measurement of small H+ leak currents via uncoupling proteins and thus, providing a rigorous understanding of the molecular mechanisms involved. In this paper we cover the methodology of measuring the H+ leak in mitochondria of specialized thermogenic tissues brown and beige fat.
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