Caspase inhibitors reduce the apoptotic but not necrotic component of kainate injury in primary murine cortical neuronal cultures.
Excitotoxicity has been demonstrated to play a major role in ischemic neuronal injury. While the necrotic component of excitotoxicity has been well demonstrated, apoptosis has also been shown to play a role. We sought to quantitate and modulate the apoptotic component of kainate-induced injury. Experiments were performed in mouse primary cortical neuronal cultures after three or 10 days in vitro. Cell death was assessed by Hoechst/propidium iodide staining and cell counting. Apoptosis was further confirmed with inhibition by caspase inhibitors. Exposure of three-day old neurons to 100 microM kainate produced an injury in which 56% +/- 0.9% of cells showed apoptotic nuclei and 13.5% +/- 2.0% showed necrotic nuclei. After 10 days in vitro neurons were more easily injured by kainate, but the cell death had primarily necrotic characteristics. Inhibition of both caspases 1 and 3 significantly reduced the apoptotic injury of 3-day old neurons. Neither reduced the necrotic component. Inhibition of protein synthesis with cycloheximide was also effective in reducing the apoptotic injury without affecting the necrotic injury. Kainate injury causes both apoptosis and necrosis, with the injury depending on both the dose of kainate and the age of the culture. The apoptotic component can be selectively reduced by caspase inhibition or cycloheximide.
Glassford
Alexander
A
Department of Anesthesia, Stanford University School of Medicine, Stanford, CA, USA.
Lee
Jong-Eun
JE
Xu
Lijun
L
Giffard
Rona G
RG
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002545:Brain Ischemia / Q000188:drug therapy / Q000201:enzymology* / Q000503:physiopathology; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects / Q000201:enzymology* / Q000503:physiopathology; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D005333:Fetus; D007608:Kainic Acid / Q000494:pharmacology; D051379:Mice; D009336:Necrosis; D009410:Nerve Degeneration / Q000188:drug therapy / Q000378:metabolism* / Q000503:physiopathology; D009474:Neurons / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D018696:Neuroprotective Agents / Q000494:pharmacology; D009498:Neurotoxins / Q000494:pharmacology
D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D018696:Neuroprotective Agents; D009498:Neurotoxins; D020169:Caspases; D007608:Kainic Acid