Cell type-specific changes in Wnt signaling and neuronal differentiation in the developing mouse cortex after prenatal alcohol exposure during neurogenesis

Abstract

Fetal Alcohol Spectrum Disorder (FASD) encompasses an array of effects of prenatal alcohol exposure (PAE), including physical abnormalities and cognitive and behavioral deficits. Disruptions of cortical development have been implicated in multiple PAE studies, with deficits including decreased progenitor proliferation, disrupted neuronal differentiation, aberrant radial migration of pyramidal neurons, and decreased cortical thickness. While several mechanisms of alcohol teratogenicity have been explored, how specific cell types in the brain at different developmental time points may be differentially affected by PAE is still poorly understood. In this study, we used single nucleus RNA sequencing (snRNAseq) to investigate whether moderate PAE from neurulation through peak cortical neurogenesis induces cell type-specific transcriptomic changes in the developing murine brain. Cluster analysis identified 25 neuronal cell types, including subtypes of radial glial cells (RGCs), intermediate progenitor cells (IPCs), projection neurons, and interneurons. Only Wnt-expressing cortical hem RGCs showed a significant decrease in the percentage of cells after PAE, with no cell types showing PAE-induced apoptosis as measured by caspase expression. Cell cycle analysis revealed only a subtype of RGCs expressing the downstream Wnt signaling transcription factor Tcf7l2 had a decreased percentage of cells in the G2/M phase of the cell cycle, suggesting decreased proliferation in this RGC subtype and further implicating disrupted Wnt signaling after PAE at this early developmental timepoint. An increased pseudotime score in IPC and projection neuron cell types indicated that PAE led to increased or premature differentiation of these cells. Biological processes affected by PAE included the upregulation of pathways related to synaptic activity and neuronal differentiation and downregulation of pathways related to chromosome structure and the cell cycle. Several cell types showed a decrease in Wnt signaling pathways, with several genes related to Wnt signaling altered by PAE in multiple cell types. As Wnt has been shown to promote proliferation and inhibit differentiation at earlier stages in development, the downregulation of Wnt signaling may have resulted in premature neuronal maturation of projection neurons and their intermediate progenitors. Overall, these findings provide further insight into the cell type-specific effects of PAE during early corticogenesis.

Risk and Resilience Variants in the Retinoic Acid Metabolic and Developmental Pathways Associated with Risk of FASD Outcomes

“…Interestingly, a greater number of resilience than risk alleles, 16 vs 11, were found in these pathways, compared to those found in the RA metabolism pathway. This was a surprising result given that animal models have shown these pathways are directly impacted by PAE, and when perturbed can exacerbate PAE teratogenesis [80,81]. Although these PAEimpacted pathways can be viewed as independent of each other, they are co-dependent and function through defined signaling gradients [82].…”

Effects of prenatal alcohol exposure on the olfactory system development

“…In addition, it is necessary to study in more detail the effects of PAE on the structure and function of regions involved in olfactory processing other than the OB, such as olfactory epithelium and olfactory cortex. Furthermore, while diverse in vivo and in vitro studies have elucidated various signaling pathways affected by PAE during brain development (Hashimoto-Torii et al, 2011;Mohammad et al, 2020;Fischer et al, 2021;Salem et al, 2021;Sambo et al, 2022), this type of research has so far been insufficient for the olfactory system. Understanding the molecules and pathways affected by PAE in the developing olfactory system could shed light on potential mechanisms underlying the etiology of abnormal sense of smell.…”

The Effects of Extrinsic and Intrinsic Factors on Neurogenesis

“…There is clear evidence that excessive alcohol consumption extensively damages multiple organ systems, especially the CNS, as chronic alcohol use leads to permanent cognitive decline and motor deficits [109]. In vitro and in vivo studies have shown that prenatal alcohol exposure impairs the development of the newly formed cortex in the developing fetus [110,111], and chronic alcohol exposure reduces neurogenesis and enhances cell death in adolescents; these changes may not be alleviated until a substantial period of time after alcohol cessation [112,113]. High-dose alcohol exposure in young adult rats suppresses hippocampal neurogenesis, including by decreasing the number of type-1 NSCs and type-2a NPCs in the DG subgranular zone and FOS+ granule cells, and reduces synaptic plasticity [114].…”