Group I metabotropic glutamate receptor inhibition selectively blocks a prolonged Ca²⁺ elevation associated with age-dependent excitotoxicity

It has been recognized for some years that a prolonged Ca²⁺ elevation that is predictive of impending cell death develops in cultured neurons following excitotoxic insult. In addition, neurons exhibit enhanced sensitivity to excitotoxic insult with increasing age in culture. However, little is known about the processes that selectively regulate the post-insult Ca²⁺ elevation and therefore, it remains unclear whether it is associated specifically with age-dependent toxicity. Here, we tested the hypothesis that a group I metabotropic glutamate receptor antagonist selectively modulates the prolonged Ca²⁺ elevation in direct association with its protective effects against excitotoxicity. Rat hippocampal cultures of two ages (8-9 and 21-28 days in vitro) were exposed to a 5-min glutamate insult (400 μM in younger and 10 μM in older cultures) sufficient to kill >50% of the neurons, and were treated with vehicle or the specific group I metabotropic glutamate receptor antagonist 1-aminoindan-1,5-dicarboxylic acid (AIDA; 1 mM), throughout and following the insult. Neuronal survival was quantified 24 h after insult. In parallel studies, neurons of similar age in culture were imaged ratiometrically with a confocal microscope during and for 60 min after the glutamate insult. A large post-insult Ca²⁺ elevation was present in older but not most younger neurons. The N-methyl-D-aspartate receptor antagonist, MK-801, blocked the Ca²⁺ elevation both during and following the insult. In contrast, AIDA blocked only the post-insult prolonged Ca²⁺ elevation in older neurons. Moreover, AIDA was neuroprotective in older but not younger cultures. From these results we suggest that the post-insult Ca²⁺ elevation is regulated differently from the Ca²⁺ elevation during glutamate insult and is modulated by group I metabotropic glutamate receptors. Further, the prolonged Ca²⁺ elevation appears to be directly linked to an age-dependent component of vulnerability.