Effect of zinc and copper ions on cadmium-induced toxicity in rat cultured cortical neurons

Abstract

Background: Cadmium is a highly toxic heavy metal that is capable of accumulating in the body and causing neurodegeneration. However, the effect of other trace elements on Cd²⁺ toxicity is currently poorly understood. The aim of this work was to study the effect of Zn²⁺ and Cu²⁺ ions on cadmium-induced death of neurons in the cerebral cortex.

Methods: The work was performed on rat cortical primary cultures. The MTT test was used to determine the cytotoxicity effects. Analysis of intracellular Ca²⁺ concentration was assessed by the Fluo-4 AM calcium indicator that exhibit an increase in fluorescence upon binding Ca²⁺. MitoSOX Red (mitochondrial superoxide indicator) was used to measuring mitochondrial ROS content in live cells.

Results: In this article, we show that the administration of CdCl₂ (0.005-0.02 mM) for 48 h induced an increase in dose-dependent death rate of cultured cortical neurons. Mature neurons were more sensitive to the damaging effects of Cd²⁺ than immature ones. ZnCl₂ (0.01-0.03 mM) significantly protected neurons from this toxic effect. In contrast to ZnCl₂, CuCl₂ (0.01 mM) increased cadmium neurotoxicity. Using Fluo-4 AM, measurements of intracellular calcium ions demonstrated that 24 h-exposure to Cd²⁺ induced intensive increase in Fluo-4 fluorescence in neurons, which was significantly reduced by zinc ions. CuCl₂ increased the cadmium-induced Fluo-4 and MitoSOX Red fluorescence in neurons. The chelator of intracellular Ca²⁺ BAPTA significantly decreased Cd²⁺-induced intensive increase in Fluo-4 fluorescence in cells.

Conclusion: The data obtained by us indicate that Zn²⁺ and Cu²⁺ can affect the neurotoxicity of cadmium in different directions: Zn²⁺ weaken the violation of intracellular calcium homeostasis caused by cadmium, preventing cell death, while Cu²⁺ potentiate the increase in the level of free intracellular calcium induced by cadmium and the development of mitochondrial dysfunction with an increase in the production of free radicals in differentiated cultured neurons of the cerebral cortex, which ultimately stimulates cytotoxicity.

Keywords: Cadmium; Calcium; Copper; Neuroprotection; Neurotoxicity; Zinc.