title: Nerve growth factor increases the sensitivity to zinc toxicity and induces cell cycle arrest in PC12 cells.
abstract: Zinc is a basic trace element that plays important roles in brain and, consequently, its homeostasis needs to be critically controlled. High zinc concentrations in the interneuron synaptic space may induce neuronal death through mechanisms still partially solved. Undifferentiated pheochromocytoma (PC12) cells have been used to study zinc toxicity. As these cells can be differentiated into neuronal-like cells, the results obtained from differentiated cultures are more useful to understand zinc toxicity in neurons. In this paper, we show by flow cytometry that nerve growth factor (NGF) induces PC12 cells differentiation characterized by cell cycle arrest in the G1/G0 phase, similarly to that observed in serum-deprived cultures. Zinc induces cell death in NGF-differentiated PC12 cultures with an EC(50) value of 143+/-14 microM, which reveals a higher sensitivity with respect to undifferentiated PC12 cultures (EC(50), 308+/-32 microM) and a similar response to that obtained in hippocampal neurons (134+/-12 microM). Thus, the differentiation process appeared responsible for such increase in sensitivity. To further support this tenet, when the NGF differentiation was impaired in presence of 10 microM MK-801, a selective blocker of the N-methyl-d-aspartate (NMDA) receptor that plays a role in the differentiation process, the higher sensitivity to zinc was reverted to an EC(50) value of 241+/-26 microM. Flow cytometry experiments showed that NGF-differentiated PC12 cells in presence of zinc were positive for propidium iodide but not for annexin-V labeling. These results, together with data from fluorescent labeling of nuclear fragmentation, caspase-3 activation, and reactive oxygen species generation, support the view that zinc toxicity in NGF-differentiated PC12 cells takes place mainly through a necrotic process.
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias, Universidad de Extremadura, Badajoz, Spain. fj171@unex.es
authors.forename: Francisco J
authors.initials: FJ
authors.lastname: Sánchez-Martín
authors.forename: Elvira
authors.initials: E
authors.lastname: Valera
authors.forename: Ilda
authors.initials: I
authors.lastname: Casimiro
authors.forename: Jaime M
authors.initials: JM
authors.lastname: Merino
mesh_terms: D000818:Animals; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects* / Q000502:physiology*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D016291:Dizocilpine Maleate / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism; D008668:Metallothionein / Q000378:metabolism; D009336:Necrosis / Q000139:chemically induced / Q000378:metabolism; D020932:Nerve Growth Factor / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D016716:PC12 Cells; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D014131:Trace Elements / Q000633:toxicity*; D019287:Zinc Sulfate / Q000633:toxicity*
chemical_list: D018691:Excitatory Amino Acid Antagonists; D017382:Reactive Oxygen Species; D014131:Trace Elements; D016291:Dizocilpine Maleate; D019287:Zinc Sulfate; D008668:Metallothionein; D020932:Nerve Growth Factor; C505410:Casp3 protein, rat; D053148:Caspase 3