Glutamate receptor requirement for neuronal death from anoxia-reoxygenation: An in vitro model for assessment of the neuroprotective effects of estrogens

1. Previous studies demonstrated that estrogens, specifically 17 beta-estra diol, the potent, naturally occurring estrogen, are neuroprotective in a va riety of models including glutamate toxicity. The aim of the present study is twofold: (1) to assess the requirement for glutamate receptors in neuron al cell death associated with anoxia-reoxygenation in three cell types, SK- N-SH and MT-22 neuronal cell lines and primary rat cortical neuronal cultur es, and (2) to evaluate the neuroprotective activity of both 17 beta-estrad iol and its weaker isomer, 17 alpha-estradiol, in both anoxia-reoxygenation and glutamate toxicity. 2. SK-N-SH and MT-22 cell lines, both of which lack NMDA receptors as asses sed by MK-801 binding assays, were resistant to both anoxia-reoxygenation a nd glutamate-induced cell death. In contrast, primary rat cortical neurons, which exhibit both NMDA and AMPA receptors, were sensitive to brief period s of exposure to anoxia-reoxygenation or glutamate. As such, there appears to be an obligatory requirement for NMDA and/or AMPA receptors in neuronal cell death resulting from brief periods of anoxia followed by reoxygenation . 3. Using primary rat cortical neurornal cultures, we evaluated the neuropro tective activity of 17 beta-estradiol (1.3 or 133 nM) and 17 alpha-estradio l (133 nM) in both anoxia-reoxygenation and excitotoxicity models of cell d eath. We found that the 133 nM but not the 1.3 nM dose of the potent estrog en, 17 beta-estradiol, protected 58.0, 57.5, and 85.3% of the primary rat c ortical neurons from anoxia-reoxygenation, glutamate, or AMPA toxicity, res pectively, and the 133 nM dose of the weak estrogen, 17 alpha-estradiol, pr otected 74.6, 81.7, and 85.8% of cells from anoxia-reoxygenation, glutamate , or AMPA toxicity, respectively. These data demonstrate that pretreatment with estrogens can attenuate glutamate excitotoxicity and that this protect ion is independent of the ability of the steroid to bind the estrogen recep tor.