Indiana University-Purdue University Indianapolis (IUPUI) / Despite warnings about the dangers of drinking during pregnancy, little progress has been made in reducing alcohol drinking among women of childbearing age. Even after the recognition of pregnancy, 15% of women continue to drink, 3% of which admit to binge drinking. Because we cannot stop women from drinking during pregnancy, and many children with fetal alcohol spectrum disorders (FASD) are adopted, there is a significant need to develop postnatal interventions that can improve the long-term outcome of children adversely affected by prenatal alcohol exposure. This thesis aims to evaluate one promising new treatment in the rehabilitation or rescue of specific learning deficits long after the damage has occurred. The treatment evaluated herein (40µg D-NAP + 40µg D-SAL) has long been used in the prevention of the detrimental effects of long-term and binge-like alcohol exposures in rodent models of fetal alcohol syndrome and FASD. Until recently this peptide treatment had only been shown to be effective in preventing some of the consequences of alcohol exposure when administered concurrently with the prenatal alcohol exposure. A recent report by Incerti and colleagues (2010c), however, reported that these peptides could completely reverse a profound spatial learning deficit induced by one episode of a heavy binge-like alcohol exposure (5.9g.kg in a single intraperitoneal injection) on gestational day 8 (G8) in C57BL/6 mice. In that report, the peptide treatment was administered starting in late adolescence, beginning three days prior to and throughout water maze training, and the profound deficits in their alcohol-placebo group were completely eliminated in the alcohol-peptide group. There are currently no FDA-approved treatments for FASD. An effective treatment for the cognitive and behavioral dysfunctions suffered by the 1% of people born today could potentially improve the lives of millions of children and adults. The first aim of this thesis was to determine whether the peptide treatment could reverse the significant spatial learning deficits we have demonstrated in adult C57BL/6 mice given high-dose binge-like alcohol exposure (2.5 g/kg in each of two intraperitoneal injections separated by two hours) on postnatal day (P)7. When administered three days prior to and throughout water maze testing (P67-76), the peptide treatment had no effect on spatial learning. The second aim sought to determine whether the same peptide treatment could reverse water maze spatial learning deficits in G8 binge-like exposure models, as reported by Incerti et al. (2010c). For this analysis, the first study used a different binge-like alcohol exposure model that is more commonly used than that employed by the Incerti et al. (2010c) study, namely administration of 2.8g/kg in each of two intraperitoneal injections separated by four hours (Sulik et al., 1981). This model has been shown to produce high peak blood alcohol concentrations and neuroanatomical aberrations in the hippocampal formation and septal regions (Parnell et al., 2009), which have been implicated in learning and memory. Surprisingly, this G8 binge-like alcohol exposure failed to produce a spatial learning deficit, undermining the usefulness of this model in evaluating the peptide effects. In direct contrast to the outcomes of Incerti et al. (2010c), the G8 Webster alcohol exposure was also unable to produce any deficits in acquisition of spatial learning in the Morris water maze. Surprisingly, neither of the heavy binge-like alcohol exposures on G8 were able to produce spatial learning deficits in the Morris water maze. The binge-like alcohol exposure on P7 did yield the expected spatial learning deficit, but the peptide treatment was unsuccessful in recovering water maze learning. These findings fail to support oral administration of 40µg D-NAP and 40 µg D-SAL as a potential therapy for postnatal alcohol-induced spatial learning deficits in adult mice.

Indiana University-Purdue University Indianapolis (IUPUI) / Fetal alcohol spectrum disorders (FASD) is the leading neurodevelopment deficit in children born to women who drink alcohol during pregnancy. The hippocampus and cortex are among brain regions vulnerable to alcohol-induced neurotoxicity, and are key regions underlying the cognitive impairment, learning and memory deficits shown in FASD individuals. Hippocampal and cortical neuronal differentiation and maturation are highly influenced by both intrinsic transcriptional signaling and extracellular cues. Epigenetic mechanisms, primarily DNA methylation and histone modifications, are hypothesized to be involved in regulating key neural development events, and are subject to alcohol exposure. Alcohol is shown to modify DNA methylation and histone modifications through altering methyl donor metabolisms. Recent studies in our laboratory have shown that alcohol disrupted genome-wide DNA methylation and delayed early embryonic development. However, how alcohol affects DNA methylation in fetal hippocampal and cortical development remains elusive, therefore, will be the theme of this study. We reported that, in a dietary alcohol-intake model of FASD, prenatal alcohol exposure retarded the development of fetal hippocampus and cortex, accompanied by a delayed cellular DNA methylation program. We identified a programed 5-methylcytosine (5mC) and 5-hydroxylmethylcytosine (5hmC) cellular and chromatic re-organization that was associated with neuronal differentiation and maturation spatiotemporally, and this process was hindered by prenatal alcohol exposure. Furthermore, we showed that alcohol disrupted locus-specific DNA methylation on neural specification genes and reduced neurogenic properties of neural stem cells, which might contribute to the aberration in neurogenesis of FASD individuals. The work of this dissertation suggested an important role of DNA methylation in neural development and elucidated a potential epigenetic mechanism in the alcohol teratogenesis.