Fetal alcohol spectrum disorder is common (affects ~2% of all live births) and is a major cause of mental dysfunction. Of the many negative effects that ethanol has on the developing nervous system, the most severe is neuronal death. It is severe because this loss is permanent;with the exception of a couple of sites, post-mitotic neurons in the CNS are not replaced by newly generated ones. Indeed, after the generation of neurons is complete, the nervous system has to decide if and whether damaged neurons can be repaired. If not, they must be eliminated. A key molecule in this decision process is the oncoprotein p53. p53 is critical for maintaining neuronal integrity and resiliency. We will test the hypothesis that developmental exposure to ethanol alters neuronal survival and DNA repair through p53-dependent activities. In the developing nervous system, neuronal death is a natural and critical process. This death appears to be apoptotic and to involve p53. Our preliminary data and the work of others on developing cerebral cortex show that exposure to ethanol during the period of naturally occurring neuronal death causes a dramatic and transient increase in both active caspase 3 expression and terminal uridylated nick-end labeling (TUNEL). On the other hand, our novel data also show that this pattern does not coincide with the ultimate loss of cortical neurons either in time or space. The implication is that ethanol causes DNA fragmentation, but this degradation may not be obligatory for apoptosis. Instead, it may reveal DNA repair mechanisms. p53 is a key player in DNA repair. Three complementary aims will be addressed using p53 deficient mice and cells. (1) Vulnerability of cortical neurons to ethanol will be addressed in long- and short-term in vivo studies. Long-term studies will determine the ethanol-induced loss of neurons in cortical layers occurring in the deficient mice. Complementary studies will examine short-term changes in the expression of presumed "death markers" (active caspase3 immunoexpression and TUNEL) in cortical layers and the timing of the neuronal loss. (2) The acute genomic responses of p53 deficient mice and cultured neural stem cells to ethanol will be determined. We will identify changes in the expression of transcripts involved in apoptosis and DNA repair. In addition, we will determine the epigenetic effects of ethanol on the silencing of genes through hyper-methylation. (3) Securin is a protein that is regulated by p53 and is pivotal for DNA repair. Preliminary microarray and immunocytochemical data show that it is profoundly affected by ethanol. Thus, we will examine the effects of ethanol on cells deficient of securin in vitro and in vivo after transplantation into layers that are apparently susceptible and refractory to ethanol. These studies will explore two dovetailed responses that developing neurons have to ethanol: DNA repair, and failing that, apoptotic death. We will address critical questions. For example, what defines the susceptibility of a young neuron to ethanol? Can neurons be manipulated to reduce their ethanol vulnerability? Thus, the studies explore two new targets of ethanol, p53 and securin, that are critical for the neuronal survival and integrity.

Public Health Relevance

Fetal alcohol spectrum disorder affects an estimated 2% of all live births in the United States. One common target of alcohol toxicity is differentiating cells, particularly those that are newly integrating into the complex environment of the developing brain. The present study will explore two complementary mechanisms by which alcohol-induced defects result (a) from ethanol-induced neuronal death and (b) from ethanol-altered DNA repair.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
5R01AA006916-29
Application #
8660005
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Hereld, Dale
Project Start
Project End
Budget Start
Budget End
Support Year
29
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Upstate Medical University
Department
Other Basic Sciences
Type
Schools of Medicine
DUNS #
City
Syracuse
State
NY
Country
United States
Zip Code
13210
Mooney, S M; Miller, M W (2011) Role of neurotrophins on postnatal neurogenesis in the thalamus: prenatal exposure to ethanol. Neuroscience 179:256-66
Hicks, Steven D; Miller, Michael W (2011) Effects of ethanol on transforming growth factor ?1-dependent and -independent mechanisms of neural stem cell apoptosis. Exp Neurol 229:372-80
Mooney, Sandra M; Miller, Michael W (2010) Prenatal exposure to ethanol affects postnatal neurogenesis in thalamus. Exp Neurol 223:566-73
Hicks, Steven D; Middleton, Frank A; Miller, Michael W (2010) Ethanol-induced methylation of cell cycle genes in neural stem cells. J Neurochem 114:1767-80
Lindke, Amanda L; Middleton, Frank A; Miller, Michael W (2010) Regulating the availability of transforming growth factor ýý1 in B104 neuroblastoma cells. Exp Neurol 225:123-32
Miller, Michael W; Hu, Huaiyu (2009) Lability of neuronal lineage decisions is revealed by acute exposures to ethanol. Dev Neurosci 31:50-7
Powrozek, Teresa A; Miller, Michael W (2009) Ethanol affects transforming growth factor beta1-initiated signals: cross-talking pathways in the developing rat cerebral wall. J Neurosci 29:9521-33
Mooney, Sandra M; Miller, Michael W (2009) Vulnerability of macaque cranial nerve neurons to ethanol is time- and site-dependent. Alcohol 43:323-31
Siegenthaler, Julie A; Tremper-Wells, Barbara A; Miller, Michael W (2008) Foxg1 haploinsufficiency reduces the population of cortical intermediate progenitor cells: effect of increased p21 expression. Cereb Cortex 18:1865-75
Siegenthaler, Julie A; Miller, Michael W (2008) Generation of Cajal-Retzius neurons in mouse forebrain is regulated by transforming growth factor beta-Fox signaling pathways. Dev Biol 313:35-46

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