The role of folic acid supplementation on exencephaly induced by maternal exposure to sodium valproate in the TO mouse

Abstract

Valproic acid (WA) is an anticonvulsant drug known to cause spina bifida in humans and exencephaly in animals. The Theiler Outbred (TO) mouse exhibits a low incidence (4.4%) of spontaneous exencephaly at birth. This frequency is enhanced by sodium valproate (VPA), an anticonvulsant drug. Earlier workers have reported that folic acid (FA) supplementation during periconceptional period prevents a substantial proportion of neural tube defects (NTD). The mechanism of preventive effect of FA is not known.
In addition to genetic factors, dose, timing and duration of supplementation during pregnancy may be important in protecting the embryos against occurrence of NTD resulting from teratogenic exposure. The objectives of this study were to determine the susceptible stages for the induction of NTD by maternal treatment with WA and to investigate how FA could rescue embryos from NTD. A single dose of 400 mg/kg of vaiproic acid was administered intraperitoneally (ip) to TO mouse on gestation day (GD) 7 or 8. This treatment resulted in significant increase in resorption, reduction in mean fetal weight, and exencephaly of live fetuses.
Exencephaly was found to be associated with several craniofacial malformations and hemorrhage. Alizarin red and alcian blue stained skeletal preparations disclosed the maxillary and mandibular hypoplasia, absence of skull vault, hemivertebrae,
longitudinal fusion of the vertebral arches and bodies, accessory ribs (cervical and lumbar), fusion of thoracic ribs, and several kinds of sternal malformations. There was a unique pattern of axial skeletal defects observed in both the exencephalic and nonexencephalic embryos in SPA treated animals. The GD 7 treated animals were susceptible for induction of cervical ribs and occipitalization of atlas vertebrae. The GD 8 group exhibited lumbar ribs and 8 sternal ribs instead of the usual 7. Nonexencephalic WA-treated embryos exhibited mandibular, maxillary hypoplasia, arched and cleft
palates, short or absent tail anomalies. During early stages of post-treatment period, the treated embryos were found to show considerable delay in elevation and fusion of neural folds. Most of them were also growth retarded.
Maternal plasma levels of folic acid and vitamin B12 were significantly lowered, with a single dose of 400 mg/kg of WA administration on GD 7 or 8. Administration of a single dose of 12 mg/kg of FA on GD 7 or 8 along with WA, resulted in an
improvement in the plasma FA and vitamin 13 12 levels. This dose did not reduce WAinduced NTD. Three doses of FA (4 mg/kg) and a single dose of WA given on GD 7 or 8 were found to reduce WA treatment related exencephaly and resorption on GD 7 but not on GD 8. In another experiment, daily administration of three doses of 4 mg/kg of FA from GD 5 through 10 was found to reduce significantly the WA-induced NTD.
These data suggest that a single high dose of FA administered at early somite stage does not prevent exencephaly. Three doses of FA administered daily from GD 5 through GD 10 restore and maintain plasma folate levels and significantly prevent exencephaly due to WA exposure in mouse embryos.