Apoptosis of bcl-x-deficient telencephalic cells in vitro.
bcl-x is a member of the bcl-2 gene family, which is expressed at high levels in the embryonic brain. The targeted disruption of bcl-x results in massive cell death of immature neurons in the developing mouse brain (Motoyama et al., 1995). bcl-x-deficient mice die around embryonic day 13 (E13), probably secondary to their inability to produce mature red blood cells. To determine whether the death of immature neurons in the bcl-x-deficient brain is cell autonomous, we examined primary telencephalic cell cultures from E12.5 homozygous mutant (bcl-x-/-), heterozygous mutant (bcl-x+/-), and wild-type (bcl-x+/+) mice. bcl-x-/- telencephalic cells cultured in 0.5 or 2.0% fetal calf serum (FCS)-containing medium for 48 hr showed increased apoptosis, defined by abnormal bisbenzamide staining and terminal-deoxytransferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL), and decreased numbers of microtubule-associated protein-2-immunoreactive neurons compared with bcl-x+/- and bcl-x+/+ cultures. Cycloheximide treatment of bcl-x-/- telencephalic cell cultures failed to prevent the increased cell death observed in low FCS-containing medium, suggesting a protein synthesis-independent apoptosis. There were no significant differences among bcl-x-/-, bcl-x+/-, and bcl-x+/+ telencephalic cells grown for 48 hr in 5% FCS-containing medium or in a chemically defined serum-free medium (ITS). bcl-x-/- neurons generated in ITS showed increased susceptibility to subsequent serum deprivation. These results indicate that bcl-x is important for both neuron maturation and survival.
Roth
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KA
Department of Pathology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Motoyama
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N
Loh
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DY
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002417:Cattle / Q000097:blood / Q000196:embryology; D002455:Cell Division; D003470:Culture Media; D016895:Culture Media, Serum-Free; D003513:Cycloheximide / Q000494:pharmacology; D005312:Fetal Blood; D051379:Mice; D009474:Neurons / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000172:deficiency*; D019253:Proto-Oncogene Proteins c-bcl-2; D013687:Telencephalon / Q000166:cytology / Q000378:metabolism*; D013997:Time Factors; D051020:bcl-X Protein
C494812:Bcl2l1 protein, mouse; D003470:Culture Media; D016895:Culture Media, Serum-Free; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051020:bcl-X Protein; D003513:Cycloheximide