Toxic effects of maternal ethanol (E) intake on the fetus are well documented both in animal models and humans. Central among the responses to in vivo E exposure are neuro-developmental deficits. A source of this is E-related neuron death that is causally connected to decreased GSH in fetal brain. In vivo and in vitro supporting studies illustrate that E generates striking inhibition of the two primary sources of fetal neuron Cys, the control point for GSH synthesis. These Cys sources are the EAAC1 Cys transporter and the transsulfuration pathway (TSP), which internally synthesizes Cys. E-inhibits both protein and mRNA expression of EAAC1 and cystathionine-?-lyase (CSE), a rate-limiting enzyme in TSP. In vivo and in vitro data support the mechanism underlying this being disruption of the calcineurin/NFAT signaling pathway which is linked to craniofacial and neurological phenotypes in Down's and Fetal Alcohol Syndrome. Hypothesis: The ability of Nrf2/ARE neuroprotective pathways to fully prevent E-induced death of fetal cerebral cortical neurons is impaired by an E-related perturbation of cellular Cys. This is due to E inhibiting activities of the excitatory amin acid transporter (EAAC1) (Aim 1) and CSE (Aim 2). This E-inhibition of EAAC1 and CSE protein is due to decreased gene expressions caused by disruption of the NFAT signaling pathway (Aim 3).
Specific Aim 1 will test the hypothesis that a mechanism underlying the inability of Nrf2/ARE interactions to provide complete protection from E-induced damage of fetal neuron GSH homeostasis is impaired Cys internalization by the EAAC1 transporter. Experiments will elucidate mechanisms underlying E effects on regulation and control of EAAC1 and its maintenance of fetal neuron GSH. Experiments will develop augmentation of neuronal Cys/GSH homeostasis as a rescue strategy.
Specific Aim 2 will test the hypothesis that the second and functionally connected component of E impairment of neuron GSH homeostasis is reduced synthesis of Cys by the transsulfuration pathway. The mechanism underlying this is an E-related decrease in cystathionine-?-lyase expression and function. These experiments will address mechanisms underlying E effects on regulation and control points of the TSP, consequences of this, the role of E inhibition of CSE in TSP function, its role in maintenance of fetal neuron GSH homeostasis, and compensatory crosstalk between EAAC1 Cys transport and the Cys synthesis pathway, TSP.
Specific Aim 3 will test the hypothesis that E-inhibition of EAAC1 and CSE protein is due to decreased gene expressions caused by disruption of the NFAT signaling pathway. Experiments will elucidate the role of NFAT signaling which has been linked to craniofacial and neurological phenotypes in Fetal Alcohol Syndrome. Focus is on NFAT3 in E induced transcriptional down regulation of EAAC1 and CSE. This occurs despite EAAC1 and CSE being targets of Nrf2/ARE signaling which is activated by E. Experiments will develop the enhancement of NFAT3 as a rescue strategy.

Public Health Relevance

The data from this project will advance our understanding of how alcohol can damage the developing brain by causing the death of neurons. The cellular pathways involved in this neuron death will be clarified and this will allow us to develop new clinical treatments for Fetal Alcohol Spectrum Disorders.

National Institute of Health (NIH)
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Research Project (R01)
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Neurotoxicology and Alcohol Study Section (NAL)
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Gao, Peter
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Texas Tech University
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Patel, Dhyanesh; Rathinam, Marylatha; Jarvis, Courtney et al. (2018) Role for Cystathionine ? Lyase (CSE) in an Ethanol (E)-Induced Lesion in Fetal Brain GSH Homeostasis. Int J Mol Sci 19:
Patel, Dhyanesh; Mahimainathan, Lenin; Narasimhan, Madhusudhanan et al. (2017) Ethanol (E) Impairs Fetal Brain GSH Homeostasis by Inhibiting Excitatory Amino-Acid Carrier 1 (EAAC1)-Mediated Cysteine Transport. Int J Mol Sci 18:
Riar, Amanjot Kaur; Narasimhan, Madhusudhanan; Rathinam, Mary Latha et al. (2016) Ethanol induces cytostasis of cortical basal progenitors. J Biomed Sci 23:6
Riar, Amanjot Kaur; Narasimhan, Madhusudhanan; Rathinam, Mary Latha et al. (2014) Ethanol-induced transcriptional activation of programmed cell death 4 (Pdcd4) is mediated by GSK-3? signaling in rat cortical neuroblasts. PLoS One 9:e98080
Narasimhan, Madhusudhanan; Riar, Amanjot Kaur; Rathinam, Mary Latha et al. (2014) Hydrogen peroxide responsive miR153 targets Nrf2/ARE cytoprotection in paraquat induced dopaminergic neurotoxicity. Toxicol Lett 228:179-91
Narasimhan, Madhusudhanan; Rathinam, Marylatha; Riar, Amanjot et al. (2013) Programmed cell death 4 (PDCD4): a novel player in ethanol-mediated suppression of protein translation in primary cortical neurons and developing cerebral cortex. Alcohol Clin Exp Res 37:96-109
Narasimhan, Madhusudhanan; Rathinam, Marylatha; Patel, Dhyanesh et al. (2012) Astrocytes Prevent Ethanol Induced Apoptosis of Nrf2 Depleted Neurons by Maintaining GSH Homeostasis. Open J Apoptosis 1:
Kruman, Inna I; Henderson, George I; Bergeson, Susan E (2012) DNA damage and neurotoxicity of chronic alcohol abuse. Exp Biol Med (Maywood) 237:740-7
Narasimhan, Madhusudhanan; Patel, Dhyanesh; Vedpathak, Dhanashree et al. (2012) Identification of novel microRNAs in post-transcriptional control of Nrf2 expression and redox homeostasis in neuronal, SH-SY5Y cells. PLoS One 7:e51111
Fowler, Anna-Kate; Hewetson, Aveline; Agrawal, Rajiv G et al. (2012) Alcohol-induced one-carbon metabolism impairment promotes dysfunction of DNA base excision repair in adult brain. J Biol Chem 287:43533-42

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