Glia-conditioned medium induces de novo synthesis of tyrosine hydroxylase and increases dopamine cell survival by differential signaling pathways.
The mesencephalic astroglia-conditioned medium (GCM) greatly increases dopamine (DA) phenotype expression, and it also protects from spontaneous and toxin-induced cell death in midbrain cultures. In this study, we have investigated the signaling pathways implicated in those effects. Genistein at 5 microM, an inhibitor of tyrosine kinase receptors, and KT-5720, a protein kinase A inhibitor, blocked the GCM-induced effects on DA phenotype expression and DA cell survival but did not abolish the increased astrocytic (glial fibrillary acidic protein-positive; GFAP+) processes. We analyzed the role of phosphatidylinositol-3 kinase (PI-3K) on TH induction and cell survival, with the PI-3K inhibitors LY-294002 and wortmannin, and the role of the phosphorylation of mitogen-activated protein kinase (MAPK) with PD-98059, a p-ERK1/2 MAPK inhibitor. LY-294002 at 20-30 microM blocked the GCM-induced effects on TH expression and DA cell survival but did not abolish the increased astrocytic processes. PD-98059 at 20 and 40 microM blocked the GCM-induced effects on DA phenotype, cell survival, and GFAP expression. However, staurosporine at 10 nM, a protein kinase C inhibitor, only blocked the protective effects induced by GCM on midbrain cell apoptosis. The data presented herein show that tyrosine kinase receptors, cAMP-dependent protein kinase, PI-3K, and MAPK signaling pathways are implicated in de novo synthesis of TH+ cells induced by GCM as well as in DA cell apoptosis and that these effects are unrelated to increased GFAP expression. PKC inhibitors only abolished the GCM-induced effects on midbrain neuronal survival, suggesting that signaling pathways for DA phenotype expression and survival may be independent.
de Bernardo
S
S
Departamento de Investigación, Servicio de Neurobiología, Hospital Ramón y Cajal, Madrid, Spain.
Canals
S
S
Casarejos
M J
MJ
Rodriguez-Martin
E
E
Mena
M A
MA
D000818:Animals; D015153:Blotting, Western; D002452:Cell Count / Q000379:methods; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000494:pharmacology*; D004298:Dopamine / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004622:Embryo, Mammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D007150:Immunohistochemistry; D008636:Mesencephalon / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009457:Neuroglia / Q000187:drug effects* / Q000201:enzymology; D009474:Neurons / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology*; D014316:Tritium / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000096:biosynthesis*; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D005904:Glial Fibrillary Acidic Protein; D014316:Tritium; D005680:gamma-Aminobutyric Acid; D014446:Tyrosine 3-Monooxygenase; D020928:Mitogen-Activated Protein Kinases; D004298:Dopamine