The profound and deleterious effects that ethanol exerts on the developing brain are well established, yet the mechanism(s) involved in its neurotoxic effects remain elusive. Receptors for neurotransmitters and growth factors and their cell signaling systems are increasingly recognized as playing relevant roles in various aspects o rain development. In the last few years we have pursued the hypothesis that the metabolism of phosphoinositides coupled to activation of acetylcholine muscarinic receptors may represent a relevant target for the developmental neurotoxicity of ethanol. Our studies have shown that ethanol exerts a dose-, age-, brain region- and neurotransmitter - specific inhibition of this biochemical response, and have demonstrated a strong correlation between disruption of muscarinic receptor - stimulated phosphoinositide metabolism during the brain growth spurt and long-lasting microencephaly. As proliferation of glial cells is a major event occurring during the in growth spurt, and following observations that acetylcholine can act as a mitogen in astrocytes, we have formulated the hypothesis that disruption of acetylcholine - driven glial cell proliferation may play a primary role in the developmental neurotoxicity of ethanol, and may provide a direct causal link between the observe inhibition of phosphoinositide metabolism and the ensuing microencephaly. The focus of the propose -experiments has shifted from investigations of the effects of ethanol in vivo or in vitro in brain slices or mixed cultures, to the study of its interactions with a selected glial cell population (astrocytes) in vitro. We believe that this shirt represents a logical progression toward a more comprehensive understanding of the sequence of event :hat may link our initial observations of the effects of ethanol on phosphoinositide metabolism to the observe neurotoxic endpoints, such as microencephaly.
The specific aims of this proposal are: 1. To investigate selected signal transduction pathways involved in the mitogenic effect of acetylcholine in cultured cortical astrocytes. These will include the hydrolysis of phosphoinositides and phosphatidylcholine, mobilization of intracellular calcium, activation of protein kinase C and induction of immediate - early - genes. 2. To investigate the molecular pathways involved in the observed inhibition by ethanol of the mitogenic action of acetylcholine in astrocytes.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Research Project (R01)
Project #
2R01AA008154-05A2
Application #
2000232
Study Section
Special Emphasis Panel (ZRG4-ALTX-3 (01))
Program Officer
Broman, Sarah H
Project Start
1991-09-27
Project End
2000-11-30
Budget Start
1996-12-01
Budget End
1997-11-30
Support Year
5
Fiscal Year
1997
Total Cost
Indirect Cost
Name
University of Washington
Department
Public Health & Prev Medicine
Type
Schools of Public Health
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Roqué, Pamela J; Guizzetti, Marina; Costa, Lucio G (2014) Synaptic structure quantification in cultured neurons. Curr Protoc Toxicol 60:12.22.1-32
Guizzetti, Marina; Kavanagh, Terrance J; Costa, Lucio G (2011) Measurements of astrocyte proliferation. Methods Mol Biol 758:349-59
Guizzetti, Marina; Moore, Nadia H; VanDeMark, Kathryn L et al. (2011) Muscarinic receptor-activated signal transduction pathways involved in the neuritogenic effect of astrocytes in hippocampal neurons. Eur J Pharmacol 659:102-7
Giordano, Gennaro; Guizzetti, Marina; Dao, Khoi et al. (2011) Ethanol impairs muscarinic receptor-induced neuritogenesis in rat hippocampal slices: Role of astrocytes and extracellular matrix proteins. Biochem Pharmacol 82:1792-9
Guizzetti, Marina; Moore, Nadia H; Giordano, Gennaro et al. (2010) Ethanol inhibits neuritogenesis induced by astrocyte muscarinic receptors. Glia 58:1395-406
Moore, Nadia H; Costa, Lucio G; Shaffer, Scott A et al. (2009) Shotgun proteomics implicates extracellular matrix proteins and protease systems in neuronal development induced by astrocyte cholinergic stimulation. J Neurochem 108:891-908
VanDeMark, Kathryn L; Guizzetti, Marina; Giordano, Gennaro et al. (2009) The activation of M1 muscarinic receptor signaling induces neuronal differentiation in pyramidal hippocampal neurons. J Pharmacol Exp Ther 329:532-42
VanDemark, Kathryn L; Guizzetti, Marina; Giordano, Gennaro et al. (2009) Ethanol inhibits muscarinic receptor-induced axonal growth in rat hippocampal neurons. Alcohol Clin Exp Res 33:1945-55
Guizzetti, Marina; Moore, Nadia H; Giordano, Gennaro et al. (2008) Modulation of neuritogenesis by astrocyte muscarinic receptors. J Biol Chem 283:31884-97
Guizzetti, Marina; Chen, Jing; Oram, John F et al. (2007) Ethanol induces cholesterol efflux and up-regulates ATP-binding cassette cholesterol transporters in fetal astrocytes. J Biol Chem 282:18740-9

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