Monitoring mitochondrial calcium and metabolism in the beating MCU-KO heart
Optical methods for measuring intracellular ions including Ca2+ revolutionized our understanding of signal transduction. However, these methods are not extensively applied to intact organs due to issues including inner filter effects, motion, and available probes. Mitochondrial Ca2+ is postulated to...
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Veröffentlicht in: | Cell reports (Cambridge) 2021-10, Vol.37 (3), p.109846-109846, Article 109846 |
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Sprache: | eng |
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Zusammenfassung: | Optical methods for measuring intracellular ions including Ca2+ revolutionized our understanding of signal transduction. However, these methods are not extensively applied to intact organs due to issues including inner filter effects, motion, and available probes. Mitochondrial Ca2+ is postulated to regulate cell energetics and death pathways that are best studied in an intact organ. Here, we develop a method to optically measure mitochondrial Ca2+ and demonstrate its validity for mitochondrial Ca2+ and metabolism using hearts from wild-type mice and mice with germline knockout of the mitochondria calcium uniporter (MCU-KO). We previously reported that germline MCU-KO hearts do not show an impaired response to adrenergic stimulation. We find that these MCU-KO hearts do not take up Ca2+, consistent with no alternative Ca2+ uptake mechanisms in the absence of MCU. This approach can address the role of mitochondrial Ca2+ to the myriad of functions attributed to alterations in mitochondrial Ca2+.
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•Rhod-2 loading in mouse hearts allows measurement of intraventricular mitochondrial Ca2+•Absorption spectra alternated with fluorescence are used to correct inner filter effects•Optical spectroscopy is used to monitor changes in mitochondrial Ca2+ and metabolism•Mitochondrial Ca2+ uptake is blocked in hearts with MCU ablation or inhibition
Kosmach et al. develop an optical spectroscopy-based method using Ca2+ indicator dye Rhod-2-AM to monitor mitochondrial Ca2+ in the ex vivo perfused mouse heart. Using this method with hearts under beta-adrenergic stimulation, we find that there are no alternative Ca2+ uptake pathways in germline MCU-KO hearts. |
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ISSN: | 2211-1247 2211-1247 |
DOI: | 10.1016/j.celrep.2021.109846 |