• Energy Research

AbstractEthanol exposure during the early stages of embryonic development can lead to a range of morphological and behavioral differences termed fetal alcohol spectrum disorders (FASDs). In a zebrafish model, we have shown that acute ethanol exposure at 8–10 hr postfertilization (hpf), a critical time of development, produces birth defects similar to those clinically characterized in FASD. Dysregulation of the Sonic hedgehog (Shh) pathway has been implicated as a molecular basis for many of the birth defects caused by prenatal alcohol exposure. We observed in zebrafish embryos that shh expression was significantly decreased by ethanol exposure at 8–10 hpf, while smo expression was much less affected. Treatment of zebrafish embryos with SAG or purmorphamine, small molecule Smoothened agonists that activate Shh signaling, ameliorated the severity of ethanol‐induced developmental malformations including altered eye size and midline brain development. Furthermore, this rescue effect of Smo activation was dose dependent and occurred primarily when treatment was given after ethanol exposure. Markers of Shh signaling (gli1/2) and eye development (pax6a) were restored in embryos treated with SAG post‐ethanol exposure. Since embryonic ethanol exposure has been shown to produce later‐life neurobehavioral impairments, juvenile zebrafish were examined in the novel tank diving test. Our results further demonstrated that in zebrafish embryos exposed to ethanol, SAG treatment was able to mitigate long‐term neurodevelopmental impairments related to anxiety and risk‐taking behavior. Our results indicate that pharmacological activation of the Shh pathway at specific developmental timing markedly diminishes the severity of alcohol‐induced birth defects.

BackgroundGenetic factors influence the physical and neurobehavioral manifestations of prenatal alcohol exposure (PAE). Animal models allow the investigation of specific genes that confer vulnerability to, or protection from, birth defects associated with fetal alcohol spectrum disorders (FASDs). The objective of the present experiments was to determine if genetic alterations in the Sonic Hedgehog (Shh) signaling pathways affect the vulnerability to PAE‐induced skeletal defects involving the forelimbs and/or hindlimbs.MethodWild‐type C57BL/6J female mice were bred with males in which one copy of the Shh or Gli2 genes had been knocked out, to produce litters with both wild‐type (+/+) and heterozygous (+/−) embryos. Alcohol doses (two injections of 2.9 g/kg, 4 hours apart) or vehicles were administered starting at gestational day (GD) 9.25, 9.5, or 9.75, a critical exposure time for inducing limb defects. Limb defects were examined at GD 17 using a dysmorphology scale based on abnormalities ranging from increased interdigital spacing to the deletion of multiple fingers and the ulna.ResultsAlcohol treatment caused a high incidence of forelimb defects, particularly on the right side, that was higher in Shh+/− and Gli2+/− fetuses compared to wild‐type fetuses. Dysmorphology scores were also significantly higher in the Shh+/− and Gli2+/− mice.ConclusionsThese results extend previous findings demonstrating enhanced sensitivity to PAE‐induced craniofacial dysmorphology and support the hypothesis that genetic alterations in the Shh signaling pathway influences the vulnerability to alcohol‐induced birth defects. Moreover, these results emphasize the importance of understanding the interactions between genes and prenatal exposure to alcohol or other teratogens. Birth Defects Research 109:860–865, 2017. © 2017 Wiley Periodicals, Inc.