Role of TPEN in Amyloid-β25-35-Induced Neuronal Damage Correlating with Recovery of Intracellular Zn2+ and Intracellular Ca2+ Overloading

The overproduction of neurotoxic amyloid-ss (A ss) peptides in the brain is a hallmark of Alzheimer's disease ( AD). To determine the role of intracellular zinc ion (iZn(2+)) dysregulation in mediating A ss-related neurotoxicity, this study aimed to investigate whether N, N, N', N'-tetrakis (2-pyridylmethyl) ethylenediamine (TPEN), a -Zn2+-specific chelator, could attenuate A ss 25-35-induced neurotoxicity and the underlying mechanism. We used the 3-(4, 5-dimethyl-thiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay to measure the viability of primary hippocampal neurons. We also determined intracellular -Zn2+ and -Ca2+ concentrations, mitochondrial and lysosomal functions, and intracellular reactive oxygen species (ROS) content in hippocampal neurons using live-cell confocal imaging. We detected L-type voltage-gated calcium channel currents (L-ICa) in hippocampal neurons using the whole-cell patch-clamp technique. Furthermore, we measured the mRNA expression levels of proteins related to the iZn2+ buffer system (ZnT-3, MT-3) and voltage-gated calcium channels (Cav1.2, Cav1.3) in hippocampal neurons using RT-PCR. The results showed that TPEN attenuated A ss 25-35-induced neuronal death, relieved the A ss 25-35-induced increase in intracellular -Zn2+ and -Ca2+ concentrations; reversed the A ss 25-35-induced increase in ROS content, the A ss 25-35-induced increase in the L-ICa peak amplitude at different membrane potentials, the A ss 25-35-induced the dysfunction of the mitochondria and lysosomes, and the A ss 25-35-induced decrease in ZnT-3 and MT-3 mRNA expressions; and increased the Cav1.2 mRNA expression in the hippocampal neurons. These results suggest that TPEN, the -Zn2+-specific chelator, attenuated A ss 25-35-induced neuronal damage, correlating with the recovery of intracellular -Zn2+ and modulation of abnormal -Ca2+-related signaling pathways.