Neuroprotective peptides prevent the alcohol-induced alteration in GABA-A3 which plays a role in cleft lip and palate in fetal alcohol syndrome

Abstract

Prenatal alcohol exposure affects 1/200 live births in US and results in cranio-facial dysmorphology including cleft lip and palate and long-term learning disabilities. The GABA receptor subunit GABA-Aβ3 is critical during embryogenesis for neural tube and palate development. The GABA-Aβ3 knock-out results in cleft palate and neonatal death that were rescued by introduction of the β3-subunit transgene. We have shown that prenatal alcohol exposure downregulated GABA-Aβ3 in the embryo. In a mouse model for fetal alcohol syndrome (FAS), neuroprotective peptides prevented alcohol-induced fetal death, growth restriction and learning deficits. Our objective was to determine if the peptides' prevention of alcohol-induced complications includes GABA-Aβ3. Timed, pregnant C57BL6/J mice were treated on gestational day 8 with either alcohol (0.03 mL/g) or placebo or alcohol+peptides (NAP+SAL 20μg). Embryos were harvested at 24h (E9) and 10 days (E18) after treatment. Using samples from at least 3 litters per time-point, calibrator-normalized relative real time PCR was performed using primers for GABA-Aβ3 with GAPDH standardization. Statistical analysis included ANOVA and Fisher PLSD. Twenty-four hours after treatment, the alcohol-induced decrease in GABA-Aβ3 was not prevented by the pretreatment with the peptides (P=0.8). However, at E18, the alcohol induced GABA-Aβ3 down regulation was prevented by the peptides (P=0.01) with levels higher than controls (P=0.007). Prenatal treatment with neuroprotective peptides prevented the alcohol-induced decline and resulted in a significant elevation in GABA-Aβ3 ten days after alcohol exposure. Since palate formation continues through gestational day 18, the increase in GABA-Aβ3 expression may be an attempt to overcompensate for alcohol's effects and may play a role in the prevention of cleft lip and palate. These findings suggest that NAP and SAL may be beneficial for prevention of cleft palate in FAS and in other pathologic conditions.