The interplay of intracellular calcium and zinc ions in response to electric field stimulation in primary rat cortical neurons in vitro

Recent pharmacological studies demonstrate a role for zinc (Zn(2+)) in shaping intracellular calcium (Ca(2+)) dynamics and vice versa in excitable cells including neurons and cardiomyocytes. Herein, we sought to examine the dynamic of intracellular release of Ca(2+) and Zn(2+) upon modifying excitability of primary rat cortical neurons using electric field stimulation (EFS) in vitro. We show that exposure to EFS with an intensity of 7.69 V/cm induces transient membrane hyperpolarization together with transient elevations in the cytosolic levels of Ca(2+) and Zn(2+) ions. The EFS-induced hyperpolarization was inhibited by prior treatment of cells with the K(+) channel opener diazoxide. Chemical hyperpolarization had no apparent effect on either Ca(2+) or Zn(2+). The source of EFS-induced rise in Ca(2+) and Zn(2+) seemed to be intracellular, and that the dynamic inferred of an interplay between Ca(2+) and Zn(2+) ions, whereby the removal of extracellular Ca(2+) augmented the release of intracellular Ca(2+) and Zn(2+) and caused a stronger and more sustained hyperpolarization. We demonstrate that Zn(2+) is released from intracellular vesicles located in the soma, with major co-localizations in the lysosomes and endoplasmic reticulum. These studies further support the use of EFS as a tool to interrogate the kinetics of intracellular ions in response to changing membrane potential in vitro.