Measurement of gamma-enolase release, a new method for selective quantification of neurotoxicity independently from glial lysis.
We have developed a sensitive enzymatic-immunoassay to quantify the level of gamma-enolase (a specific neuronal enzyme) which is released from cultured cells after exposure to various toxins. We show that this method can estimate selectively neuronal cell death without significantly interfering with glial cell death. Indeed, no gamma-enolase is released when glial cells are killed with free-radical producing agents. Experiments comparing the levels of neuronal cell death induced by NMDA or free-radical producing drugs, performed either by measuring gamma-enolase release or using the classical fluorescein diacetate method, yielded similar results. In addition to selectively follow neuronal death in a mixed population of neurons and glial cells, this method provides a way of determining the cell death kinetics from a single culture dish, since enolase can be measured on small samples taken from the culture medium. Finally, we propose these two methods as being complementary and useful neuronal and other cellular death indexes and also to understand the complex problem of glial influence on neuronal survival or death.
Lafon-Cazal
M
M
Centre CNRS-INSERM de Pharmacologie-Endocrinologie, Montpellier, France.
Bougault
I
I
Steinberg
R
R
Pin
J P
JP
Bockaert
J
J
D000818:Animals; D015415:Biomarkers; D016923:Cell Death / Q000187:drug effects*; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology*; D007118:Immunoassay / Q000379:methods; D007700:Kinetics; D008773:Methylphenazonium Methosulfate / Q000633:toxicity*; D051379:Mice; D016202:N-Methylaspartate / Q000633:toxicity*; D009457:Neuroglia / Q000166:cytology / Q000187:drug effects*; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D009498:Neurotoxins / Q000633:toxicity*; D010751:Phosphopyruvate Hydratase / Q000032:analysis* / Q000378:metabolism; D013997:Time Factors; D014969:Xanthine Oxidase / Q000633:toxicity*
D015415:Biomarkers; D009498:Neurotoxins; D008773:Methylphenazonium Methosulfate; D016202:N-Methylaspartate; D014969:Xanthine Oxidase; D010751:Phosphopyruvate Hydratase