Ratiometric-pericam-mt, a novel tool to evaluate intramitochondrial zinc

Zn²⁺ can enter mitochondria and promote a plethora of physiological and patho-physiological effects. The issue of measuring changes in intramitochondrial levels is therefore critical. Past studies have employed fluorescent Zn²⁺ indicators, like Rhod-2 and RhodZin-3, however, the use of these probes is impaired by their extramitochondrial sequestration. In this study, we show that the ratiometric mitochondria-targeted pericam, RPmt, can be employed to detect changes of intramitochondrial free Zn²⁺ ([Zn²⁺]_m) levels. Using RPmt in neuronal and non neuronal cell lines we demonstrate that mitochondria can take up the cation mobilized from the cytosolic pool of protein-bound Zn²⁺ and that mitochondrial Zn²⁺ sequestration is largely mediated by the activity of the Ca²⁺ uniporter.