TY - JOUR
T1 - Ethanol impairs muscarinic receptor-induced neuritogenesis in rat hippocampal slices
T2 - Role of astrocytes and extracellular matrix proteins
AU - Giordano, Gennaro
AU - Guizzetti, Marina
AU - Dao, Khoi
AU - Mattison, Hayley A.
AU - Costa, Lucio G.
N1 - Funding Information:
This work was supported in part by grants R01AA08154 from NIAAA and P30ES07033 from NIEHS . We thank Dr. Andres Barria (Dept. of Physiology and Biophysics, University of Washington) for his help with the hippocampal slice preparation and the transfection, Dr. Pamela Lein (University of California at Davis) for helpful suggestions on preparation and culturing of hippocampal slices, and Mr. Glen MacDonald (Center for Human Developmental Disabilities, University of Washington), for his help with the confocal laser scanning microscope.
PY - 2011/12/1
Y1 - 2011/12/1
N2 - In an in vitro co-culture system of astrocytes and neurons, stimulation of cholinergic muscarinic receptors in astrocytes had been shown to cause neuritogenesis in hippocampal neurons, and this effect was inhibited by ethanol. The present study sought to confirm these earlier findings in a more complex system, in vitro rat hippocampal slices in culture. Exposure of hippocampal slices to the cholinergic agonist carbachol (1 mM for 24 h) induced neurite outgrowth in hippocampal pyramidal neurons, which was mediated by activation of muscarinic M3 receptors. Specifically, carbachol induced a >4-fold increase in the length of the longest neurite, and a 4-fold increase in the length of minor neurites and in the number of branches. Co-incubation of carbachol with ethanol (50 mM) resulted in significant inhibition of the effects induced by carbachol on all parameters measured. Neurite outgrowth in CNS neurons is dependent on various permissive factors that are produced and released by glial cells. In hippocampal slices carbachol increased the levels of two extracellular matrix protein, fibronectin and laminin-1, by 1.6-fold, as measured by Western blot. Co-incubation of carbachol with ethanol significantly inhibited these increases. Carbachol-induced increases in levels of extracellular matrix proteins were antagonized by a M3 muscarinic receptor antagonist. Furthermore, function-blocking fibronectin or laminin-1 antibodies antagonized the effect of carbachol on neurite outgrowth. These results indicate that in hippocampal slices stimulation of muscarinic M3 receptors induces neurite outgrowth, which is mediated by fibronectin and laminin-1, two extracellular matrix proteins released by astrocytes. By decreasing fibronectin and laminin levels ethanol prevents carbachol-induced neuritogenesis. These findings highlight the importance of glial-neuronal interactions as important targets in the developmental neurotoxicity of alcohol.
AB - In an in vitro co-culture system of astrocytes and neurons, stimulation of cholinergic muscarinic receptors in astrocytes had been shown to cause neuritogenesis in hippocampal neurons, and this effect was inhibited by ethanol. The present study sought to confirm these earlier findings in a more complex system, in vitro rat hippocampal slices in culture. Exposure of hippocampal slices to the cholinergic agonist carbachol (1 mM for 24 h) induced neurite outgrowth in hippocampal pyramidal neurons, which was mediated by activation of muscarinic M3 receptors. Specifically, carbachol induced a >4-fold increase in the length of the longest neurite, and a 4-fold increase in the length of minor neurites and in the number of branches. Co-incubation of carbachol with ethanol (50 mM) resulted in significant inhibition of the effects induced by carbachol on all parameters measured. Neurite outgrowth in CNS neurons is dependent on various permissive factors that are produced and released by glial cells. In hippocampal slices carbachol increased the levels of two extracellular matrix protein, fibronectin and laminin-1, by 1.6-fold, as measured by Western blot. Co-incubation of carbachol with ethanol significantly inhibited these increases. Carbachol-induced increases in levels of extracellular matrix proteins were antagonized by a M3 muscarinic receptor antagonist. Furthermore, function-blocking fibronectin or laminin-1 antibodies antagonized the effect of carbachol on neurite outgrowth. These results indicate that in hippocampal slices stimulation of muscarinic M3 receptors induces neurite outgrowth, which is mediated by fibronectin and laminin-1, two extracellular matrix proteins released by astrocytes. By decreasing fibronectin and laminin levels ethanol prevents carbachol-induced neuritogenesis. These findings highlight the importance of glial-neuronal interactions as important targets in the developmental neurotoxicity of alcohol.
KW - Ethanol
KW - Extracellular matrix
KW - Fibronectin
KW - Hippocampal slice
KW - Muscarinic receptors
KW - Neuritogenesis
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U2 - 10.1016/j.bcp.2011.08.014
DO - 10.1016/j.bcp.2011.08.014
M3 - Article
C2 - 21884684
AN - SCOPUS:80255129267
SN - 0006-2952
VL - 82
SP - 1792
EP - 1799
JO - Biochemical Pharmacology
JF - Biochemical Pharmacology
IS - 11
ER -