Prenatal alcohol exposure triggers ceramide-induced apoptosis in neural crest-derived tissues concurrent with defective cranial development

Fetal alcohol syndrome (FAS) is caused by maternal alcohol consumption during pregnancy. The reason why specific embryonic tissues are sensitive toward ethanol is not understood. We found that in neural crest-derived cell (NCC) cultures from the first branchial arch of E10 mouse embryos, incubation with ethanol increases the number of apoptotic cells by fivefold. Apoptotic cells stain intensely for ceramide, suggesting that ceramide-induced apoptosis mediates ethanol damage to NCCs. Apoptosis is reduced by incubation with CDP-choline (citicoline), a precursor for the conversion of ceramide to sphingomyelin. Consistent with NCC cultures, ethanol intubation of pregnant mice results in ceramide elevation and increased apoptosis of NCCs in vivo. Ethanol also increases the protein level of prostate apoptosis response 4 (PAR-4), a sensitizer to ceramide-induced apoptosis. Prenatal ethanol exposure is concurrent with malformation of parietal bones in 20% of embryos at day E18. Meninges, a tissue complex derived from NCCs, is disrupted and generates reduced levels of TGF-β1, a growth factor critical for bone and brain development. Ethanol-induced apoptosis of NCCs leading to defects in the meninges may explain the simultaneous presence of cranial bone malformation and cognitive retardation in FAS. In addition, our data suggest that treatment with CDP-choline may alleviate the tissue damage caused by alcohol

Primary Cilia Are Not Required for Normal Canonical Wnt Signaling in the Mouse Embryo

Sonic hedgehog (Shh) signaling in the mouse requires the microtubule-based organelle, the primary cilium. The primary cilium is assembled and maintained through the process of intraflagellar transport (IFT) and the response to Shh is blocked in mouse mutants that lack proteins required for IFT. Although the phenotypes of mouse IFT mutants do not overlap with phenotypes of known Wnt pathway mutants, recent studies report data suggesting that the primary cilium modulates responses to Wnt signals.We therefore carried out a systematic analysis of canonical Wnt signaling in mutant embryos and cells that lack primary cilia because of loss of the anterograde IFT kinesin-II motor (Kif3a) or IFT complex B proteins (Ift172 or Ift88). We also analyzed mutant embryos with abnormal primary cilia due to defects in retrograde IFT (Dync2h1). The mouse IFT mutants express the canonical Wnt target Axin2 and activate a transgenic canonical Wnt reporter, BAT-gal, in the normal spatial pattern and to the same quantitative level as wild type littermates. Similarly, mouse embryonic fibroblasts (MEFs) derived from IFT mutants respond normally to added Wnt3a. The switch from canonical to non-canonical Wnt also appears normal in IFT mutant MEFs, as both wild-type and mutant cells do not activate the canonical Wnt reporter in the presence of both Wnt3a and Wnt5a.We conclude that loss of primary cilia or defects in retrograde IFT do not affect the response of the midgestation embryo or embryo-derived fibroblasts to Wnt ligands

Prenatal zinc treatment at the time of acute ethanol exposure limits spatial memory impairments in mouse offspring

Zinc (Zn) treatment given together with acute ethanol in early pregnancy has previously been demonstrated to protect against physical birth abnormalities in mice. The current study examined whether this Zn treatment (s.c. injection) can also prevent the more subtle cognitive impairments caused by ethanol exposure in early pregnancy. Pregnant C57BL/6J dams were injected with saline (0.85% wt/vol NaCl) or 25% ethanol (0.015 mL/g body weight) intraperitoneally at 0 and 4 h on gestational d (GD) 8. ZnSO4 (2.5 μg Zn/g at 0 h) treatment was administered by s.c. injection immediately following ethanol treatment. Offspring were randomly selected from litters for each of the three treatment groups and were tested at 55 and 70 d of age using a cross-maze water escape task for spatial learning and memory impairments consecutively. No differences were observed between treatments for the spatial learning task. However, young adult mice exposed to ethanol in utero demonstrated impaired spatial memory, with a decrease in correct trials and increased escape latency and incorrect entry measurements, compared with saline-treated controls. In comparison, offspring given s.c. Zn treatment at the time of ethanol exposure were not cognitively impaired, performing at the same level as control mice in the cross-maze escape task. These findings indicate that critically timed Zn administration can limit spatial memory impairments caused by ethanol exposure in early pregnancy.Brooke L. Summers, Allan M. Rofe and Peter Coyl