FASD Inhibition of ASPH-Notch Mediates Adolescent Cerebral White Matter Pathology-Potential Utility of Non-invasive Extracellular Vesicle Assays

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Long-term effects of fetal alcohol spectrum disorder (FASD) on the CNS vary with exposure dose, duration, and timing. Despite decades of research, we still lack efficient means of detecting and therapeutically targeting the damage. However, progress may have been limited by our failure to attend to the full spectrum of brain pathology linked to neurobehavioral dysfunction. For example, besides neurons, alcohol exposures across the lifespan, but especially during development, exert toxic/metabolic effects that impact the integrity of white matter (WM) myelin, axons, and oligodendrocytes (OLs), yet the mechanisms are poorly understood. Dysfunction or loss of OLs compromises the structure and function of myelin, impairing axonal conductivity. Preliminary studies showed that developmental exposures to ethanol reduce the expression of mature myelin-associated genes and proteins, in part due to impaired maturation of oligoprogenitor cells and/or survival of mature OLs. Attendant hypomyelination and dysmyelination render WM axons vulnerable to toxic/metabolic injury and degeneration with loss of synaptic connections. This competing renewal R37 application builds on our previous gains that mechanistically linked ethanol-impaired insulin and IGF-1 signaling through PI3K-Akt and downstream pathways to deficits in neuronal survival and migration during development. Another critical success of those experiments was that they revealed how ethanol inhibition of aspartyl-asparaginyl-β-hydroxylase (ASPH) and Notch impair survival, maturation and migration of immature CNS neurons. However, empirical observations and subsequent preliminary studies led to the hypothesis that WM OLs may be similarly impaired by ethanol via disruption of Akt-mTOR-mTORC1/2 which alters the composition and integrity of WM myelin. We now propose to extend these gains by determining how ethanol inhibits insulin/IGF-Akt-mTOR-mTORC1/2 signaling and reduces ASPH-Notch activation in relation to WM structure and function during postnatal and adolescent development. Aim 1 will delineate mechanisms of long term FASD WM and OL pathologies by examining developmental stage-associated impairments in signaling through insulin/IGF-1-PI3K-Akt-mTOR-mTORC1/2, and ASPH-Notch activation. Aim 2 will assess FASD dose- dependent inhibition of sulfatide (ST) in relation to impairments in OL maturation; in many CNS diseases, ST depletion in WM mediates cognitive-motor deficits. Aim 3 stems from preliminary studies showing that ethanol’s effects on WM sphingolipid profiles, including STs, are detectable in peripheral blood circulating extracellular vesicles (EVs). Therefore, we propose to examine the utility of MALDI Mass Spectrometry (MS) EV assays for non-invasive detection of WM pathology and responses to treatments that restore ASPH-Notch pathways needed for OL/WM maturation and function. Our hypotheses will be addressed with in vivo, slice culture, and isolated OL models, quantitative immunohistochemistry, electron microscopy, multiplex ELISAs, mRNA analyses, and MALDI-MS. The research is novel, mechanistic, robust, transparent, and inclusive of dose effects and both sexes. The prospects for extending this research to humans via non-invasive MALDI EV assays are extremely high.