Pathogenesis of methanol-induced craniofacial defects in C57BL/6J mice.
BACKGROUND
Methanol administered to C57BL/6J mice during gastrulation causes severe craniofacial dysmorphology. We describe dysmorphogenesis, cell death, cell cycle assessment, and effects on development of cranial ganglia and nerves observed following administration of methanol to pregnant C57BL/6J mice on gestation day (GD) 7.


METHODS
Mice were injected (i.p.) on GD 7 with 0, 2.3, 3.4, or 4.9 gm/kg methanol, split into two doses. In embryos of mice treated with 0 or 4.9 gm/kg methanol, we used histology and LysoTracker red staining on GD 8 0 hr through GD 8 18 hr to examine cell death and dysmorphogenesis, and we also evaluated cell-cycle distribution and proliferation using flow cytometry (FCM) and BrdU immunohistochemistry. On GD 10, we evaluated the effect of GD 7 exposure to 0, 2.3, 3.4, or 4.9 gm/kg methanol on cranial ganglia and nerve development using neurofilament immunohistochemistry.


RESULTS
Methanol treatment on GD 7 resulted in reduced mesenchyme surrounding the fore- and midbrain, and in the first branchial arches, by GD 8 12 hr. There were disruptions in the forebrain neuroepithelium and optic pit. Neural crest cell emigration from the mid- and hindbrain region was reduced in methanol-exposed embryos. Methanol had no apparent effect on BrdU incorporation or cell-cycle distribution on GD 8. Cell death was observed in the hindbrain region along the path of neural crest migration and in the trigeminal ganglion on GD 8 18 hr. Development of the cranial ganglia and nerves was adversely affected by methanol. Development of ganglia V, VIII, and IX was decreased at all dosage levels; ganglion VII was reduced at 3.4 and 4.9 gm/kg, and ganglion X was reduced at 4.9 gm/kg.


CONCLUSIONS
These results suggest that gastrulation-stage methanol exposure affects neural crest cells and the anterior mesoderm and neuroepithelium. Cell death was evident in areas of migrating neural crest cells, but only at time points after methanol was cleared from the embryo, suggesting an indirect effect on these cells. Birth Defects Research (Part A), 2004. Published 2004 Wiley-Liss, Inc.
Degitz
Sigmund J
SJ
United States Environmental Protection Agency, Research Triangle Park, North Carolina, USA. degitz.sigmund@epa.gov
Zucker
Robert M
RM
Kawanishi
Clinton Y
CY
Massenburg
Gwen S
GS
Rogers
John M
JM
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D003391:Cranial Nerves / Q000002:abnormalities / Q000187:drug effects; D019465:Craniofacial Abnormalities / Q000139:chemically induced* / Q000503:physiopathology; D005260:Female; D005724:Ganglia / Q000002:abnormalities / Q000187:drug effects; D000432:Methanol / Q000633:toxicity*; D051379:Mice; D008810:Mice, Inbred C57BL; D012997:Solvents / Q000633:toxicity*
D012997:Solvents; D000432:Methanol