Micrograph of a peripheral nerve, with the neuromuscular endplates stained in red. The nerve-cell mitochondria were imaged with a fluorescent redox sensor (green in the cytoplasm, yellow at the endplates).
Scientists at two German universities have developed a hybrid microscopic imaging technique to investigate real-time signals within mitochondria, the organelles that perform vital tasks within living cells (Nature Medicine, doi:10.1038/nm.3520).
Specifically, researchers from Ludwig Maximilian University of Munich and the Technical University of Munich wanted to trace the reduction/oxidation, or redox, signals mediated by reactive oxygen species inside cells. Cells need a certain amount of these molecules in order to function, but high levels can cause structural damage to the mitochondria.
To get living nerve tissue, the team obtained transgenic mice carrying a biosensor that could be tagged with a specific kind of green fluorescent protein sensitive to redox signals. The researchers imaged nerve and muscle tissue samples from the mice with a wide-field microscope equipped with a polychromator that rapidly switched between two excitation wavelengths of the protein, 408 and 488 nm. Applying hydrogen peroxide to the tissue stimulated redox signals.
Combining confocal and two-photon excitation microscopy enabled the scientists to track the redox signals and mitochondrial calcium currents in response to exterior stimuli, such as the severance of a nerve. When an axon is cut, its mitochondria send oxidation signals in a wave up the fiber. The team also imaged spontaneous contractions of mitochondria during changes in their redox state.