title: Copper-uptake is critical for the down regulation of synapsin and dynamin induced by neocuproine: modulation of synaptic activity in hippocampal neurons.
abstract: Extracellular and intracellular copper and zinc regulate synaptic activity and plasticity, which may impact brain functionality and human behavior. We have found that a metal coordinating molecule, Neocuproine, transiently increases free intracellular copper and zinc levels (i.e., min) in hippocampal neurons as monitored by Phen Green and FluoZin-3 fluorescence, respectively. The changes in free intracellular zinc induced by Neocuproine were abolished by the presence of a non-permeant copper chelator, Bathocuproine (BC), indicating that copper influx is needed for the action of Neocuproine on intracellular Zn levels. Moreover, Neocuproine decreased the mRNA levels of Synapsin and Dynamin, and did not affect the expression of Bassoon, tubulin or superoxide dismutase (SOD). Western blot analysis showed that protein levels of synapsin and dynamin were also down regulated in the presence of Neocuproine and that these changes were accompanied by a decrease in calcium transients and neuronal activity. Furthermore, Neocuproine decreased the number of active neurons, effect that was blocked by the presence of BC, indicating that copper influx is needed for the action of Neocuproine. We finally show that Neocuproine blocks the epileptiform-like activity induced by bicuculline in hippocampal neurons. Collectively, our data indicates that presynaptic protein configuration and function of primary hippocampal neurons is sensitive to transient changes in transition metal homeostasis. Therefore, small molecules able to coordinate transition metals and penetrate the blood-brain barrier might modify neurotransmission at the Central Nervous System (CNS). This might be useful to establish therapeutic approaches to control the neuronal hyperexcitabiltity observed in brain conditions that are associated to copper dyshomeotasis such as Alzheimer's and Menkes diseases. Our work also opens a new avenue to find novel and effective antiepilepsy drugs based in metal coordinating molecules.
authors.affiliation: Department of Physiology and Membrane Biology, Shriners Hospital for Children Northern California, University of California at Davis School of Medicine California, USA.
authors.forename: Patricio A
authors.initials: PA
authors.lastname: Castro
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile ; Oxidation Biology Laboratory, The Florey Institute of Neuroscience and Mental Health, The University of Melbourne Parkville, Melbourne, Victoria, Australia.
authors.forename: Alejandra
authors.initials: A
authors.lastname: Ramirez
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile.
authors.forename: Fernando J
authors.initials: FJ
authors.lastname: Sepúlveda
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile.
authors.forename: Christian
authors.initials: C
authors.lastname: Peters
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile.
authors.forename: Humberto
authors.initials: H
authors.lastname: Fierro
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile.
authors.forename: Javier
authors.initials: J
authors.lastname: Waldron
authors.affiliation: Oxidation Biology Laboratory, The Florey Institute of Neuroscience and Mental Health, The University of Melbourne Parkville, Melbourne, Victoria, Australia.
authors.forename: Sandra
authors.initials: S
authors.lastname: Luza
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile.
authors.forename: Jorge
authors.initials: J
authors.lastname: Fuentealba
authors.affiliation: Laboratory of Molecular Physiology and Channelopathies, Departament de Ciències Experimentals i de la Salut, Universitat Pompeu Fabra Barcelona, Spain.
authors.forename: Francisco J
authors.initials: FJ
authors.lastname: Muñoz
authors.affiliation: Center for Biomedical Research and FONDAP Center for Genome Regulation, Universidad Andrés Bello Santiago, Chile.
authors.forename: Giancarlo V
authors.initials: GV
authors.lastname: De Ferrari
authors.affiliation: Oxidation Biology Laboratory, The Florey Institute of Neuroscience and Mental Health, The University of Melbourne Parkville, Melbourne, Victoria, Australia.
authors.forename: Ashley I
authors.initials: AI
authors.lastname: Bush
authors.affiliation: Laboratorio de Neurofisiología, Departamento de Fisiología, Facultad de Ciencias Biológicas, Universidad de Concepción Concepción, Chile.
authors.forename: Luis G
authors.initials: LG
authors.lastname: Aguayo
authors.affiliation: Oxidation Biology Laboratory, The Florey Institute of Neuroscience and Mental Health, The University of Melbourne Parkville, Melbourne, Victoria, Australia.
authors.forename: Carlos M
authors.initials: CM
authors.lastname: Opazo
keywords: copper; dynamin; epileptiform-like activity; hyperexcitability; neocuproine; synapsin; synaptic activity; zinc