Intracellular Free Zinc During Cardiac Excitation-contraction Cycle: Calcium and Redox Dependencies

Aims: Zinc exists in biological systems as bound and histochemically reactive free Zn(2+). It is an essential structural constituent of many proteins, including enzymes from cellular signalling pathways, in which it functions as a signalling molecule. In cardiomyocytes at rest, Zn(2+) concentration is in the nanomolar range. Very little is known about precise mechanisms controlling the intracellular distribution of Zn(2+) and its variations during cardiac function.

Methods And Results: Live-cell detection of intracellular Zn(2+) has become feasible through the recent development of Zn(2+)-sensitive and -selective fluorophores able to distinguish Zn(2+) from Ca(2+). Here, in freshly isolated rat cardiomyocytes, we investigated the rapid changes in Zn(2+) homeostasis using the Zn(2+)-specific fluorescent dye, FluoZin-3, in comparison to Ca(2+)-dependent fluo-3 fluorescence. Zn(2+) sparks and Zn(2+) transients, in quiescent and electrically stimulated cardiomyocytes, respectively, were visualized in a similar manner to known rapid Ca(2+) changes. Both Zn(2+) sparks and Zn(2+) transients required Ca(2+) entry. Inhibiting the sarcoplasmic reticulum Ca(2+) release or increasing the Ca(2+) load in a low-Na(+) solution suppressed or increased Zn(2+) movements, respectively. Mitochondrial inhibitors slightly reduced both Zn(2+) sparks and Zn(2+) transients. Oxidation by H₂O₂ facilitated and acidic pH inhibited the Ca(2+)-dependent Zn(2+) release.

Conclusion: It is proposed that Zn(2+) release during the cardiac cycle results mostly from intracellular free Ca(2+) increase, triggering production of reactive oxygen species that induce changes in metal-binding properties of metallothioneins and other redox-active proteins, aside from ionic exchange on these proteins.