title: Validation of organotypical hippocampal slice cultures as an ex vivo model of brain ischemia: different roles of NMDA receptors in cell death signalling after exposure to NMDA or oxygen and glucose deprivation.
abstract: N-Methyl-D-aspartate receptors (NMDARs) are essential mediators of synaptic plasticity under normal physiological conditions. During brain ischemia, these receptors are excessively activated due to glutamate overflow and mediate excitotoxic cell death. Although organotypical hippocampal slice cultures are widely used to study brain ischemia in vitro by induction of oxygen and glucose deprivation (OGD), there is scant data regarding expression and functionality of NMDARs in such slice cultures. Here, we have evaluated the contribution of NMDARs in mediating excitotoxic cell death after exposure to NMDA or OGD in organotypical hippocampal slice cultures after 14 days in vitro (DIV14). We found that all NMDAR subunits were expressed at DIV14. The NMDARs were functional and contributed to cell death, as evidenced by use of the NMDAR antagonist MK-801 (dizocilpine). Excitotoxic cell death induced by NMDA could be fully antagonized by 10 μM MK-801, a dose that offered only partial protection against OGD-induced cell death. Very high concentrations of MK-801 (50-100 μM) were required to counteract cell death at long delays (48-72 h) after OGD. The relative high dose of MK-801 needed for long-term protection after OGD could not be attributed to down-regulation of NMDARs at the gene expression level. Our data indicate that NMDAR signaling is just one of several mechanisms underlying ischemic cell death and that prospective cytoprotective therapies must be directed to multiple targets.
authors.affiliation: Center for Molecular Biology and Neuroscience and Department of Anatomy, Institute of Basic Medical Sciences, University of Oslo, Norway.
authors.forename: Hanna
authors.initials: H
authors.lastname: Ahlgren
authors.forename: Kristi
authors.initials: K
authors.lastname: Henjum
authors.forename: Ole Petter
authors.initials: OP
authors.lastname: Ottersen
authors.forename: Elise
authors.initials: E
authors.lastname: Rundén-Pran
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002545:Brain Ischemia / Q000378:metabolism / Q000473:pathology*; D002417:Cattle; D016923:Cell Death / Q000187:drug effects; D002645:Chickens; D016291:Dizocilpine Maleate / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D005947:Glucose / Q000172:deficiency / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D008954:Models, Biological; D016202:N-Methylaspartate / Q000494:pharmacology*; D009498:Neurotoxins / Q000494:pharmacology; D009924:Organ Culture Techniques / Q000379:methods*; D010100:Oxygen / Q000494:pharmacology*; D021122:Protein Subunits / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D016194:Receptors, N-Methyl-D-Aspartate / Q000235:genetics / Q000378:metabolism*; D015203:Reproducibility of Results; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D009498:Neurotoxins; D021122:Protein Subunits; D012333:RNA, Messenger; D016194:Receptors, N-Methyl-D-Aspartate; D016202:N-Methylaspartate; D016291:Dizocilpine Maleate; D005947:Glucose; D010100:Oxygen