title: Increased vulnerability of hippocampal neurons with age in culture: temporal association with increases in NMDA receptor current, NR2A subunit expression and recruitment of L-type calcium channels.
abstract: Excessive glutamate (Glu) stimulation of the NMDA-R is a widely recognized trigger for Ca(2+)-mediated excitotoxicity. Primary neurons typically show a large increase in vulnerability to excitotoxicity with increasing days in vitro (DIV). This enhanced vulnerability has been associated with increased expression of the NR2B subunit or increased NMDA-R current, but the detailed age-courses of these variables in primary hippocampal neurons have not been compared in the same study. Further, it is not clear whether the NMDA-R is the only source of excess Ca(2+). Here, we used primary hippocampal neurons to examine the age dependence of the increase in excitotoxic vulnerability with changes in NMDA-R current, and subunit expression. We also tested whether L-type voltage-gated Ca(2+) channels (L-VGCCs) contribute to the enhanced vulnerability. The EC(50) for Glu toxicity decreased by approximately 10-fold between 8-9 and 14-15 DIV, changing little thereafter. Parallel experiments found that during the same period both amplitude and duration of NMDA-R current increased dramatically; this was associated with an increase in protein expression of the NR1 and NR2A subunits, but not of the NR2B subunit. Compared to MK-801, ifenprodil, a selective NR2B antagonist, was less effective in protecting older than younger neurons from Glu insult. Conversely, nimodipine, an L-VGCC antagonist, protected older but not younger neurons. Our results indicate that enhanced excitotoxic vulnerability with age in culture was associated with a substantial increase in NMDA-R current, concomitant increases in NR2A and NR1 but not NR2B subunit expression, and with apparent recruitment of L-VGCCs into the excitotoxic process.
authors: Department of Molecular and Biomedical Pharmacology, Chandler Medical Center, University of Kentucky, Lexington, KY 40536-0298, USA.
authors: Lawrence D
authors: LD
authors: Brewer
authors: Olivier
authors: O
authors: Thibault
authors: Jeanise
authors: J
authors: Staton
authors: Veronique
authors: V
authors: Thibault
authors: Justin T
authors: JT
authors: Rogers
authors: Gisela
authors: G
authors: Garcia-Ramos
authors: Susan
authors: S
authors: Kraner
authors: Philip W
authors: PW
authors: Landfield
authors: Nada M
authors: NM
authors: Porter
mesh_terms: D000375:Aging / Q000502:physiology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004622:Embryo, Mammalian; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D018698:Glutamic Acid / Q000633:toxicity; D006624:Hippocampus / Q000166:cytology*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016202:N-Methylaspartate / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D018696:Neuroprotective Agents / Q000494:pharmacology; D018408:Patch-Clamp Techniques / Q000379:methods; D011247:Pregnancy; D051381:Rats; D017207:Rats, Sprague-Dawley; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism*
chemical_list: D020746:Calcium Channels, L-Type; C120997:NR2A NMDA receptor; D018696:Neuroprotective Agents; D016194:Receptors, N-Methyl-D-Aspartate; D018698:Glutamic Acid; D016202:N-Methylaspartate; D016291:Dizocilpine Maleate; D007770:L-Lactate Dehydrogenase; D002118:Calcium