Is the Mitochondrial Membrane Potential (∆Ψ) Correctly Assessed? Intracellular and Intramitochondrial Modifications of the ∆Ψ Probe, Rhodamine 123

• Published in: International journal of molecular sciences Vol. 23; no. 1; p. 482
• Main Authors: Zorova, Ljubava D, Demchenko, Evgeniya A, Korshunova, Galina A, Tashlitsky, Vadim N, Zorov, Savva D, Andrianova, Nadezda V, Popkov, Vasily A, Babenko, Valentina A, Pevzner, Irina B, Silachev, Denis N, Plotnikov, Egor Y, Zorov, Dmitry B
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 01.01.2022; MDPI
• Subjects: Amiodarone; Animals; Astrocytes - metabolism; ATP synthase; Bioenergetics; Brain tumors; Cancer therapies; Cell Extracts; Cell Line, Tumor; Cytochrome P450; Cytochromes P450; Electric components; energetics; esterase; Flow cytometry; Fluorescence; Fluorescent indicators; Glioma - metabolism; Glioma cells; Homeostasis; Hydrogen ions; Intracellular; Mass spectrometry; Mass spectroscopy; Membrane potential; Membrane Potential, Mitochondrial; Membranes; Mitochondria; Mitochondria - metabolism; Molecular Probes - metabolism; Proteins; Rats; Rhodamine; Rhodamine 123 - metabolism; Time Factors; Tumor cells; Tumors; Xenobiotics; fluorescence; mitochondria; membrane potential; cancer; esterase; cytochrome P450; tumor cells; energetics
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms23010482

The mitochondrial membrane potential (∆Ψ) is the driving force providing the electrical component of the total transmembrane potential of hydrogen ions generated by proton pumps, which is utilized by the ATP synthase. The role of ∆Ψ is not limited to its role in bioenergetics since it takes part in other important intracellular processes, which leads to the mandatory requirement of the homeostasis of ∆Ψ. Conventionally, ∆Ψ in living cells is estimated by the fluorescence of probes such as rhodamine 123, tetramethylrodamine, etc. However, when assessing the fluorescence, the possibility of the intracellular/intramitochondrial modification of the rhodamine molecule is not taken into account. Such changes were revealed in this work, in which a comparison of normal (astrocytic) and tumor (glioma) cells was conducted. Fluorescent microscopy, flow cytometry, and mass spectrometry revealed significant modifications of rhodamine molecules developing over time, which were prevented by amiodarone apparently due to blocking the release of xenobiotics from the cell and their transformation with the participation of cytochrome P450. Obviously, an important role in these processes is played by the increased retention of rhodamines in tumor cells. Our data require careful evaluation of mitochondrial ∆Ψ potential based on the assessment of the fluorescence of the mitochondrial probe.