Figure 1. Maternal, paternal, and dual parental alcohol exposures each induce changes in offspring craniofacial patterning.(A) We used a 2× 2 factorial design to contrast craniofacial development among the offspring of maternal (MatExp), paternal (PatExp), and dual parental (DualExp) alcohol exposures. Using geometric morphometrics, followed by canonical variate analysis and multivariate ANOVA, we identified treatment-specific shifts in the profiles of the gestational day 16.5 face (analysis of right profile male offspring shown in B, n=~ 48).(C) Representative wire diagram demonstrating the shift in facial landmarks of a PatExp male head along principal component 1. The blue lines demarcate the individual sample, while the turquoise lines demarcate the average of the entire population.(D) χ2 analysis, followed by Holm-Bonferroni correction, revealed an increased incidence of microcephaly, with an increased proportion of male offspring exhibiting head sizes at or below the smallest 10th percentile of the control population. We used ANOVA, followed by Dunnett’s multiple comparison test, to examine established alcohol-related craniofacial phenotypes, including changes in (E) ocular size (right eye shown),(F) midfacial depth, and (G) snout-occipital distance (males, top; females, bottom; n= 30–50). Pearson correlation analysis, followed by Holm-Bonferroni correction, contrasted offspring (H) snout-occipital distance (males, top; females, bottom) and (I) normalized brain weights (males, left; females, right), with average parental daily EtOH dose (n= 30–50, see Supplemental Table 1). Data represent mean±SEM.* P< 0.05,** P< 0.01,