Astrocytes protect neurons from ethanol-induced oxidative stress and apoptotic death.
Ethanol induces oxidative stress in cultured fetal rat cortical neurons and this is followed by apoptotic death, which can be prevented by normalization of cell content of reduced glutathione (GSH). Because astrocytes can play a central role in maintenance of neuron GSH homeostasis, the following experiments utilized cocultures of neonatal rat cortical astrocytes and fetal cortical neurons to determine if astrocytes could protect neurons from ethanol-mediated apoptotic death via this mechanism. In cortical neurons cultured in the absence of astrocytes, ethanol (2.5 and 4 mg/ml; 6-, 12-, and 24-hr exposures) decreased trypan blue exclusion and the MTT viability measures by up to 45% (P < 0.05), increased levels of reactive oxygen species (ROS) by up to 81% (P < 0.05), and decreased GSH within 1 hr of treatment by 49 and 51% for 2.5 and 4 mg/ml, respectively (P < 0.05). This was followed by onset of apoptotic cell death as determined by increased Annexin V binding and DNA fragmentation by 12 hr of ethanol exposure. Coculturing neurons with astrocytes prevented GSH depletion by 2.5 mg/ml ethanol, whereas GSH content was increased over controls in neurons exposed to 4 mg/ml ethanol (by up to 341%; P < 0.05). Ethanol generated increases in neuron ROS and apoptosis; decreases in viability were also prevented by coculture. Astrocytes were largely insensitive to ethanol, using the same measures. Only exposure to 4.0 mg/ml ethanol decreased GSH content in astrocytes, concomitant with a 204% increase in GSH efflux (P < 0.05). These studies illustrate that astrocytes can protect neurons from ethanol-mediated apoptotic death and that this may be related to maintenance of neuron GSH.
Watts
Lora Talley
LT
Department of Pharmacology, The University of Texas Health Science Center, San Antonio, TX, USA.
Rathinam
Mary Latha
ML
Schenker
Steven
S
Henderson
George I
GI
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001253:Astrocytes / Q000166:cytology* / Q000378:metabolism*; D002450:Cell Communication / Q000502:physiology*; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D002492:Central Nervous System Depressants / Q000494:pharmacology; D004396:Coloring Agents / Q000493:pharmacokinetics; D000431:Ethanol / Q000494:pharmacology; D005260:Female; D063647:Fetal Alcohol Spectrum Disorders / Q000378:metabolism / Q000473:pathology; D005978:Glutathione / Q000378:metabolism; D006706:Homeostasis / Q000187:drug effects / Q000502:physiology; D009474:Neurons / Q000166:cytology* / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D011247:Pregnancy; D051381:Rats; D017207:Rats, Sprague-Dawley; D014343:Trypan Blue / Q000493:pharmacokinetics
D002492:Central Nervous System Depressants; D004396:Coloring Agents; D000431:Ethanol; D005978:Glutathione; D014343:Trypan Blue