Endogenous activation of mGlu5 metabotropic glutamate receptors supports self-renewal of cultured mouse embryonic stem cells.
Cultured mouse embryonic stem (ES) cells maintained under undifferentiated conditions (i.e. grown in medium containing 15% FCS and leukemia inhibitory factor--LIF) expressed mGlu5 metabotropic glutamate receptors. Activation of these receptors with quisqualate increased [Ca2+]i but only when cultures were deprived of extracellular glutamate, indicating that the receptor was saturated by the endogenous glutamate. Pharmacological blockade of mGlu5 receptors with 2-methyl-6-(phenylethynyl)pyridine (MPEP) or antisense-induced knock-down of mGlu5 receptors decreased the expression of the two main transcription factors that sustain ES cell self-renewal, i.e. Oct-4 and Nanog, as assessed by real-time PCR and immunoblotting. Exposure of ES cell cultures to MPEP also reduced alkaline phosphatase activity, a marker of undifferentiated ES cells. These data support a critical role for mGlu receptors in early development showing that mGlu5 receptors are expressed by ES cells and their activation sustains ES cell self-renewal in culture.
Cappuccio
Irene
I
Department of Human Physiology and Pharmacology, University of Rome La Sapienza, Piazzale Aldo Moro 5, 00185 Rome, Italy.
Spinsanti
Paola
P
Porcellini
Antonio
A
Desiderati
Francesca
F
De Vita
Teresa
T
Storto
Marianna
M
Capobianco
Loredana
L
Battaglia
Giuseppe
G
Nicoletti
Ferdinando
F
Melchiorri
Daniela
D
D000410:Alanine Transaminase / Q000494:pharmacology; D000818:Animals; D015152:Blotting, Northern / Q000379:methods; D015153:Blotting, Western / Q000379:methods; D001921:Brain / Q000166:cytology; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004347:Drug Interactions; D004622:Embryo, Mammalian; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists; D005434:Flow Cytometry / Q000379:methods; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000502:physiology*; D018698:Glutamic Acid / Q000032:analysis; D015850:Interleukin-6 / Q000494:pharmacology; D053605:Leukemia Inhibitory Factor; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D016318:Quisqualic Acid / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013234:Stem Cells / Q000187:drug effects / Q000502:physiology*; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D014316:Tritium / Q000378:metabolism
D018690:Excitatory Amino Acid Agonists; D018691:Excitatory Amino Acid Antagonists; D015850:Interleukin-6; D053605:Leukemia Inhibitory Factor; C507838:Lif protein, mouse; D020319:Oligodeoxyribonucleotides, Antisense; D011725:Pyridines; D012333:RNA, Messenger; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate; D014316:Tritium; D018698:Glutamic Acid; C121465:6-methyl-2-(phenylethynyl)pyridine; D016318:Quisqualic Acid; D000410:Alanine Transaminase; D002118:Calcium; D013936:Thymidine