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Journal of Neuroscience, Vol 16, 1753-1758, Copyright © 1996 by Society for Neuroscience
Apoptosis of bcl-x-deficient telencephalic cells in vitro
KA Roth, N Motoyama and DY Loh
Department of Pathology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
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.
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