title: Thiol oxidation and altered NR2B/NMDA receptor functions in in vitro and in vivo pilocarpine models: implications for epileptogenesis.
abstract: Hippocampal sclerosis, the main pathological sign of chronic temporal lobe epilepsy (TLE), is associated with oxidative injury, altered N-methyl d-aspartate receptor (NMDAR) stoichiometry, and loss of hippocampal neurons. However, the mechanisms that drive the chronic progression of TLE remain elusive. Our previous studies have shown that NADPH oxidase activation and ERK 1/2 phosphorylation are required for the up-regulation of the predominantly pre-synaptic NR2B subunit auto-receptor in both in vitro and in vivo pilocarpine (PILO) models of TLE. To provide further understanding of the cellular responses during the early-stages of hyper excitability, we investigated the role of oxidative damage and altered NR2B functions. In rat primary hippocampal cultures, we found that N-acetylcysteine (NAC) prevented PILO-mediated thiol oxidation, apoptosis, cell death and NR2B subunit over-expression. Interestingly, NAC did not block thiol oxidation when added to the neurons 6h after the PILO exposure, suggesting that disulfide formation could rapidly become an irreversible phenomenon. Moreover, NAC pre-treatment did not prevent PILO-induced NR2A subunit over-expression, a critical event in hippocampal sclerosis. Pre-treatment with the highly specific NR2B subunit inhibitor, ifenprodil, partially decreased PILO-mediated thiol oxidation and was not effective in preventing apoptosis and cell death. However, if acutely administered 48h after PILO exposure, ifenprodil blocked glutamate-induced aberrant calcium influx, suggesting the crucial role of NR2B over-expression in triggering neuronal hyper-excitability. Furthermore, ifenprodil treatment was able to prevent NR2A subunit over-expression by means of ERK1/2 phosphorylation. Our findings indicate oxidative stress and NR2B/NMDA signaling as promising therapeutic targets for co-treatments aimed to prevent chronic epilepsy following the seizure onset.
authors: Pittsburgh Institute for Neurodegenerative Diseases and Department of Neurology, University of Pittsburgh, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA; Ri.MED Foundation, 10, Piazza Sett'Angeli Palermo, Italy. Electronic address: rdimaio@hs.pitt.edu.
authors: Roberto
authors: R
authors: Di Maio
authors: Pittsburgh Institute for Neurodegenerative Diseases and Department of Neurology, University of Pittsburgh, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA; Department of Genetics, Erasmus MC, Rotterdam, The Netherlands. Electronic address: p.g.mastroberadino@erasmusmc.nl.
authors: Pier G
authors: PG
authors: Mastroberardino
authors: Pittsburgh Institute for Neurodegenerative Diseases and Department of Neurology, University of Pittsburgh, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA. Electronic address: xhcz2@pitt.edu.
authors: Xiaoping
authors: X
authors: Hu
authors: Pittsburgh Institute for Neurodegenerative Diseases and Department of Neurology, University of Pittsburgh, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA. Electronic address: lmm129@pitt.edu.
authors: Laura M
authors: LM
authors: Montero
authors: Pittsburgh Institute for Neurodegenerative Diseases and Department of Neurology, University of Pittsburgh, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA. Electronic address: jgreena@pitt.edu.
authors: J Timothy
authors: JT
authors: Greenamyre
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004195:Disease Models, Animal; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004833:Epilepsy, Temporal Lobe / Q000188:drug therapy / Q000503:physiopathology*; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D006624:Hippocampus / Q000187:drug effects / Q000503:physiopathology*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D008297:Male; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D018696:Neuroprotective Agents / Q000494:pharmacology; D010084:Oxidation-Reduction / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D010862:Pilocarpine; D010880:Piperidines / Q000494:pharmacology; D017207:Rats, Sprague-Dawley; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism*
chemical_list: D018691:Excitatory Amino Acid Antagonists; C120997:NR2A NMDA receptor; C121001:NR2B NMDA receptor; D018696:Neuroprotective Agents; D010880:Piperidines; D016194:Receptors, N-Methyl-D-Aspartate; D013438:Sulfhydryl Compounds; D010862:Pilocarpine; D016291:Dizocilpine Maleate; C010739:ifenprodil; D000111:Acetylcysteine