Prenatal alcohol exposure (PAE) is the leading known cause of mental retardation and birth defects. Its teratogenicity originates, in part, through its initiation of apoptosis in critical cell populations. Under previous NIAAA support, we developed a chick embryo model of PAE and showed that ethanol causes the selective apoptosis of the neural crest, an embryonic cell population containing craniofacial and neuronal precursors. Here, we hypothesize that ethanol induces neural crest apoptosis by activating the phospholipase C-dependent release of intracellular Ca2+. This hypothesis is tested using pharmacologic approaches, taking advantage that we can locally target agonists and antagonists of Ca2+ signaling pathways to the premigratory neural crest in ovo, and thus test their ability to attenuate ethanol's effects upon the embryo. Experiments in this proposal test three sub-hypotheses: 1) Acute ethanol induces neural crest apoptosis by stimulating the release of intracellular Ca2+. The requirement for Ca2+ release is tested through the use of fluorescent Ca2+ indicators, Ca2+ ionophores and chelators, and the direct assay of downstream, Ca2+-dependent signaling proteins (CaM kinase, calcineurin). 2) Ethanol mobilizes intracellular Ca2+ and induces apoptosis by activating inositol triphosphate (IP3) receptors. Pharmacologic agonists and antagonists test that the ethanol-released Ca2+ originates from IP3-mediated stores; this IP3 release will be quantified directly. Potential contributions of ryanodine receptors and extracellular Ca2+ also are tested. 3) Ethanol's activation of Phospholipase C (PLC) is responsible for the Ca2+ release and neural crest apoptosis. PLC activation is measured by direct assay and by targeted inhibitors. Contributions of diacylglycerol, receptor-mediated tyrosine kinases, and receptor-mediated G proteins are investigated. these results address the molecular mechanism underlying the neural crest's sensitivity to ethanol, and, thus, contribute to understanding the basis for ethanol's teratogenicity. Results also may provide possible explanations for genetic-based variations in fetal responses to prenatal ethanol exposure.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Research Project (R01)
Project #
5R01AA011085-10
Application #
6890379
Study Section
Alcohol and Toxicology Subcommittee 4 (ALTX)
Program Officer
Foudin, Laurie L
Project Start
1996-07-01
Project End
2007-03-31
Budget Start
2005-05-01
Budget End
2007-03-31
Support Year
10
Fiscal Year
2005
Total Cost
$310,489
Indirect Cost
Name
University of Wisconsin Madison
Department
Nutrition
Type
Schools of Earth Sciences/Natur
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Berres, Mark E; Garic, Ana; Flentke, George R et al. (2017) Transcriptome Profiling Identifies Ribosome Biogenesis as a Target of Alcohol Teratogenicity and Vulnerability during Early Embryogenesis. PLoS One 12:e0169351
Huebner, Shane M; Blohowiak, Sharon E; Kling, Pamela J et al. (2016) Prenatal Alcohol Exposure Alters Fetal Iron Distribution and Elevates Hepatic Hepcidin in a Rat Model of Fetal Alcohol Spectrum Disorders. J Nutr 146:1180-8
Garic, Ana; Berres, Mark E; Smith, Susan M (2014) High-throughput transcriptome sequencing identifies candidate genetic modifiers of vulnerability to fetal alcohol spectrum disorders. Alcohol Clin Exp Res 38:1874-82
Palmer, Jessica A; Poenitzsch, Ashley M; Smith, Susan M et al. (2012) Metabolic biomarkers of prenatal alcohol exposure in human embryonic stem cell-derived neural lineages. Alcohol Clin Exp Res 36:1314-24
Garic, Ana; Flentke, George R; Amberger, Ed et al. (2011) CaMKII activation is a novel effector of alcohol's neurotoxicity in neural crest stem/progenitor cells. J Neurochem 118:646-57
Smith, Susan M; Flentke, George R; Kragtorp, Katherine A et al. (2011) Ethanol exposure during the early first trimester equivalent impairs reflexive motor activity and heightens fearfulness in an avian model. Alcohol 45:57-63
Wolff, Garen S; Chiang, Po Jen; Smith, Susan M et al. (2007) Epidermal growth factor-like growth factors prevent apoptosis of alcohol-exposed human placental cytotrophoblast cells. Biol Reprod 77:53-60
Garic-Stankovic, Ana; Hernandez, Marcos; Flentke, George R et al. (2006) Structural constraints for alcohol-stimulated Ca2+ release in neural crest, and dual agonist/antagonist properties of n-octanol. Alcohol Clin Exp Res 30:552-9
Kilburn, Brian A; Chiang, Po Jen; Wang, Jun et al. (2006) Rapid induction of apoptosis in gastrulating mouse embryos by ethanol and its prevention by HB-EGF. Alcohol Clin Exp Res 30:127-34