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

Brain Res. 2007 Jun 2;1151:20-31. doi: 10.1016/j.brainres.2007.03.020. Epub 2007 Mar 12.


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.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Aging / physiology*
  • Animals
  • Calcium / metabolism
  • Calcium Channels, L-Type
  • Cell Survival / drug effects
  • Cells, Cultured
  • Dizocilpine Maleate / pharmacology
  • Embryo, Mammalian
  • Female
  • Gene Expression / drug effects
  • Glutamic Acid / toxicity
  • Hippocampus / cytology*
  • L-Lactate Dehydrogenase / metabolism
  • Membrane Potentials / drug effects
  • Membrane Potentials / physiology
  • N-Methylaspartate / pharmacology
  • Neurons / drug effects
  • Neurons / physiology*
  • Neuroprotective Agents / pharmacology
  • Patch-Clamp Techniques / methods
  • Pregnancy
  • Rats
  • Rats, Sprague-Dawley
  • Receptors, N-Methyl-D-Aspartate / metabolism*


  • Calcium Channels, L-Type
  • NR2A NMDA receptor
  • Neuroprotective Agents
  • Receptors, N-Methyl-D-Aspartate
  • Glutamic Acid
  • N-Methylaspartate
  • Dizocilpine Maleate
  • L-Lactate Dehydrogenase
  • Calcium