Activation of mitogen-activated protein kinase by muscarinic receptors in astroglial cells: Role in DNA synthesis and effect of ethanol

• Published in: Glia 2001-08, Vol.35 (2), p.111-120
• Main Authors: Yagle, Kevin, Lu, Hailing, Guizzetti, Marina, Möller, Thomas, Costa, Lucio G.
• Format: Article
• Language: English
• Subjects: Animals; Astrocytes - cytology; Astrocytes - drug effects; Astrocytes - enzymology; Astrocytoma; astroglial cells; Biological and medical sciences; Brain - abnormalities; Brain - drug effects; Brain - physiopathology; Carbachol - pharmacology; Cell Division - drug effects; Cholinergic Agonists - pharmacology; DNA - biosynthesis; ethanol; Ethanol - pharmacology; Female; Fetal Alcohol Spectrum Disorders - enzymology; Fetal Alcohol Spectrum Disorders - physiopathology; Fundamental and applied biological sciences. Psychology; Humans; Isolated neuron and nerve. Neuroglia; Mitogen-Activated Protein Kinase 1 - drug effects; Mitogen-Activated Protein Kinase 1 - metabolism; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases - drug effects; Mitogen-Activated Protein Kinase Kinases - metabolism; Mitogen-Activated Protein Kinases - drug effects; Mitogen-Activated Protein Kinases - metabolism; mitogen-activated-protein kinase; Muscarinic Antagonists - pharmacology; muscarinic receptors; Pertussis Toxin; Pregnancy; protein kinase C; Protein Kinase C - drug effects; Protein Kinase C - metabolism; Rats; Receptors, Muscarinic - drug effects; Receptors, Muscarinic - metabolism; Tumor Cells, Cultured; Vertebrates: nervous system and sense organs; Virulence Factors, Bordetella - pharmacology
• ISSN: 0894-1491; 1098-1136
• DOI: 10.1002/glia.1076

Mitogen‐activated protein kinase (MAPK) can be phosphorylated by mitogens binding to G‐protein‐coupled receptors and is considered a major pathway involved in cell proliferation. In this study, we report on the activation of MAPK by muscarinic acetylcholine receptors in astroglial cells, namely the 1321N1 human astrocytoma cell line, primary rat cortical astrocytes, and fetal human astrocytes. Carbachol caused a rapid and transient phorphorylation of MAPK (ERK1/2) in all cell types, with an increase in MAPK activity, without changing the levels of MAPK proteins. Human astrocytoma cells were used to characterize the effect of carbachol on MAPK. Experiments with M2‐ and M3‐receptor subtype‐selective antagonists, and with pertussis toxin, indicated that the M3 subtype is responsible for activating MAPK in glial cells. Pretreatment of cells with the protein kinase C (PKC) inhibitor bisindolylmaleimide I, or downregulation of PKC by 24‐h treatment with the phorbol ester TPA inhibited carbachol‐induced MAPK activation. Additional experiments with PKC α‐ or PKC ϵ‐specific compounds indicated that the ϵ isozyme of PKC is primarily involved in MAPK activation by carbachol. Chelation of calcium also inhibited MAPK activation by carbachol. Two MEK (MAPK kinase) inhibitors inhibited carbachol‐induced DNA synthesis but only at concentrations that exceeded those sufficient to block carbachol‐induced MAPK activation. Ethanol (≤200 mM) had no effect on MAPK when present alone and did not affect carbachol‐induced MAPK activation under various experimental conditions, although it inhibits carbachol‐induced DNA synthesis at low concentrations (10–100 mM). These results suggest that activation of MAPK by carbachol may be necessary but not sufficient for its mitogenic effect in astroglial cells, and that does not represent a target for ethanol‐induced inhibition of DNA synthesis elicited by muscarinic receptors. GLIA 35:111–120, 2001. © 2001 Wiley‐Liss, Inc.