Mitochondrial Localization and Function of Adenosine Receptors

G-protein coupled receptors (GPCRs) are typically expressed on the cell surface where they mediate extracellular signals from hormones, neurotransmitters and growth factors, among others. However, growing evidence support the intracellular localization of GPCRs, including mitochondria. In the present work, we assessed the presence and functionality of adenosine receptors in mitochondria by combining techniques such as western blotting, radioligand binding, electron microscopy, enzymatic activities determination, oxygen consumption measurement and 3D morphological analysis of mitochondrial networks. Our results demonstrate the mitochondrial localization of adenosine A₁ and A₂ receptors in pure mitochondria fractions isolated from mouse brain and liver, human brain, HeLa and SH-SY5Y cells. Adenylyl cyclase activity assays revealed that these receptors are functional in the mitochondria. Moreover, exposure of isolated mitochondria to selective A₁, A_2A and A_2B receptors agonists revealed these receptors as potential modulators of mitochondrial energy metabolism, since ATP production and coupling efficiency increased in the presence of BAY 60-6583 (A_2B agonist) whereas proton leak and acute response were higher with CGS 21680 (A_2A agonist). Also, proton leak, ATP production, acute response and acute respiration were increased in the presence of CPA (A₁ agonist). Interestingly, different mitochondrial morphological changes were detected in HeLa cells exposed to these receptors' agonists.