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Published 16 August 2004. doi:10.1083/jcb.200403102
The Rockefeller University Press, 0021-9525 $8.00
JCB, Volume 166, Number 4, 527-536
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Article

 In vivo monitoring of Ca2+ uptake into mitochondria of mouse skeletal muscle during contraction

Rüdiger Rudolf1, Marco Mongillo1,2, Paulo J. Magalhães1, and Tullio Pozzan1,2

1 Department of Biomedical Sciences, Consiglio Nazionale Delle Ricerche (CNR) Institute of Neurosciences, University of Padua, I-35121 Padua, Italy
2 Venetian Institute of Molecular Medicine (VIMM), I-35129 Padua, Italy

Address correspondence to Tullio Pozzan, Dept. of Biomedical Sciences, University of Padua, Viale G. Colombo 3, I-35121 Padua, Italy. Tel.: 39-049-827-6070. Fax: 39-049-827-6049. email: tullio.pozzan{at}unipd.it

Although the importance of mitochondria in patho-physiology has become increasingly evident, it remains unclear whether these organelles play a role in Ca2+ handling by skeletal muscle. This undefined situation is mainly due to technical limitations in measuring Ca2+ transients reliably during the contraction–relaxation cycle. Using two-photon microscopy and genetically expressed "cameleon" Ca2+ sensors, we developed a robust system that enables the measurement of both cytoplasmic and mitochondrial Ca2+ transients in vivo. We show here for the first time that, in vivo and under highly physiological conditions, mitochondria in mammalian skeletal muscle take up Ca2+ during contraction induced by motor nerve stimulation and rapidly release it during relaxation. The mitochondrial Ca2+ increase is delayed by a few milliseconds compared with the cytosolic Ca2+ rise and occurs both during a single twitch and upon tetanic contraction.

Key Words: calcium; cameleon; in vivo; Na+/Ca2+ exchange; two-photon microscopy

Abbreviations used in this paper: 2mtYC2, YC with mitochondrial targeting signal; [Ca2+], concentration of Ca2+; [Ca2+]c, cytoplasmic concentration of Ca2+; [Ca2+]m, mitochondrial Ca2+; FRET, fluorescence resonance energy transfer; ROI, region of interest; YC, yellow cameleon.


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