Mechanisms of Choline's Improvement in Prenatal Alcohol Exposure

Univ Of North Carolina Chapel Hill, Chapel Hill NC

Investigators

Abstract

Prenatal alcohol exposure (PAE) alters gene expression and brain development and causes cognitive and behavioral impairment in humans and in animal models, and there is great interest in interventions that ameliorate these. One intervention showing efficacy is supplementation with the nutrient choline. Higher maternal choline status during pregnancy is positively associated with behavioral and cognitive improvements that persist throughout childhood, and both prenatal and postnatal choline treatment mitigate behavioral deficits in animal models of PAE and improve outcomes in people with Fetal Alcohol Spectrum Disorders. It also protects against a PAE-induced reduction in brain weight in mouse PAE models, but it is not known how choline improves brain development under PAE or if it affects crucial biological processes including cell proliferation and apoptosis. To gain insights into the potential mechanisms, we performed transcriptomics and recently reported that PAE alters the expression of multiple genes in the Hippo signaling pathway, which is crucial for brain development and growth due to its transcriptional regulation of cell proliferation. Complementary to this, in vitro studies in our lab show that PAE reduces proliferation and increases apoptosis of neural progenitor cells (NPCs), which can be regulated by the Hippo signaling pathway. Others have shown that this pathway is also influenced by maternal choline status, causing us to further investigate our transcriptomics analyses, to discover that choline supplementation during PAE reversed the impact of PAE on several Hippo components. In this proposal I hypothesize that the reductions in fetal brain size following PAE are due, in part, to increases in Hippo signaling that reduce the expansion of the neural progenitor cell population, and that choline improves brain development, in part, by normalizing those signals. These studies will generate novel insights into choline’s mechanism of action. I will test this using in vivo and in vitro techniques. I will use an established PAE mouse model where pregnant C57BL/6J mouse dams are exposed to alcohol and treated with choline, then fetal brains are collected at embryonic day (E)14.5. I will also manipulate Hippo signaling in a primary NPC culture model, followed by alcohol treatment in media having different choline concentrations. Experiments in Aim 1 use E14.5 fetal brains for anatomical analysis of cell proliferation and apoptosis and Hippo signaling components, then targeted qPCR and western blot to quantify the Hippo components that are dysregulated by PAE and mitigated by prenatal choline supplementation. Aim 2 uses transfected NPCs to induce gain- and loss-of-function of Hippo components to identify the role of Hippo pathway in alcohol’s effects on cell proliferation apoptosis and how choline remediates. This project fills a critical gap in understanding how prenatal choline supplementation mitigates the deficits in brain development caused by PAE and will help clarify the role of choline in normal brain development as well as the need for increased choline consumption during pregnancy to minimize the impact of stressors such as alcohol.