Nuclear factor kappaB activation by muscarinic receptors in astroglial cells: effect of ethanol.
Activation of muscarinic receptors leads to proliferation of astroglial cells and this effect is inhibited by ethanol. Among the intracellular pathways involved in the mitogenic action of muscarinic agonists, activation of the atypical protein kinase C zeta (PKC zeta) appears to be of most importance, and is also affected by low ethanol concentrations. PKC zeta has been reported to activate nuclear factor kappaB (NF-kappaB), a transcription factor that has been shown to play an important role in cell proliferation. The aim of this study was, therefore, to determine whether muscarinic receptors would activate NF-kappaB in astroglial cells, whether such activation would play a role in the mitogenic action of muscarinic agonists, and whether it would represent a possible target for ethanol. Carbachol activated NF-kappaB in human 1321N1 astrocytoma cells, as evidenced by translocation of the p65 subunit of NF-kappaB to the nucleus, phosphorylation and degradation of IkappaBalpha in the cytosol, and increase NF-kappaB binding to DNA. Carbachol also induced translocation of p65 to the nucleus in primary rat astrocytes. Carbachol-induced NF-kappaB activation was mediated by the M3 subtype of muscarinic receptors and appeared to involve Ca(2+) mobilization and activation of PKC epsilon and PKC zeta, but not PI3-kinase and mitogen-activated protein kinase. The NF-kappaB peptide inhibitor SN50, but not the inactive peptide SN50M, strongly inhibited carbachol-induced astrocytoma cells proliferation and p65 translocation to the nucleus. Increased DNA synthesis was also antagonized by the IkappaBalpha kinase inhibitor BAY 11-7082. Ethanol (25-100 mM) inhibited the translocation of p65 and the binding of NF-kappaB to DNA in both 1321N1 astrocytoma cells and primary rat cortical astrocytes. Together, these results suggest that activation of NF-kappaB by muscarinic receptors in astroglial cells is important for carbachol-induced DNA synthesis and that ethanol-mediated inhibition of cell proliferation may be due in part to inhibition of NF-kappaB activation.
Guizzetti
M
M
Department of Environmental and Occupational Health Sciences, University of Washington, 4225 Roosevelt Way Northeast 100, Seattle, WA 98105, USA. marinag@u.washington.edu
Bordi
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Dieguez-Acuña
F J
FJ
Vitalone
A
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Madia
F
F
Woods
J S
JS
Costa
L G
LG
D001253:Astrocytes / Q000187:drug effects* / Q000378:metabolism; D001254:Astrocytoma; D001285:Atropine / Q000494:pharmacology; D015153:Blotting, Western / Q000379:methods; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D022082:Cellular Structures / Q000187:drug effects / Q000378:metabolism; D002492:Central Nervous System Depressants / Q000494:pharmacology*; D002614:Chelating Agents / Q000494:pharmacology; D018679:Cholinergic Agonists / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology*; D005703:Gallamine Triethiodide / Q000494:pharmacology; D006801:Humans; D018727:Muscarinic Antagonists / Q000494:pharmacology; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D018733:Nicotinic Antagonists / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000378:metabolism*; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D014316:Tritium / Q000378:metabolism
D002492:Central Nervous System Depressants; D002614:Chelating Agents; D018679:Cholinergic Agonists; D004791:Enzyme Inhibitors; D018727:Muscarinic Antagonists; D016328:NF-kappa B; D018733:Nicotinic Antagonists; D010880:Piperidines; D011976:Receptors, Muscarinic; D014316:Tritium; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D000431:Ethanol; D004533:Egtazic Acid; D001285:Atropine; C042375:4-diphenylacetoxy-1,1-dimethylpiperidinium; D002217:Carbachol; D037342:Pertussis Toxin; D005703:Gallamine Triethiodide; D013936:Thymidine