Muscarinic receptor-mediated phosphorylation of cyclic AMP response element binding protein in human neuroblastoma cells.
This study describes the effect of signalling through muscarinic acetylcholine receptors on two transcription factors implicated in long-term synaptic plasticity and memory formation, EGR1 and the cyclic AMP response element binding protein (CREB). In SK-N-SH neuroblastoma cells, treatment with the cholinergic agonist carbachol led to maximal induction of EGR1 1 h after stimulation. This was preceded by the phosphorylation of CREB, which peaked as early as 5 minutes after carbachol treatment. The levels of both EGR1 and phosphorylated CREB (pCREB) slowly decayed over 4-8 h. CREB phosphorylation and EGR1 induction showed similar sensitivity to carbachol concentration, with EC(50) values in the range of 1-10 microM, and the changes in both transcription factors were blocked by the muscarinic antagonist atropine. As has been described elsewhere, EGR1 induction was dependent on activation of p42/44 MAP kinase, as it was blocked by the MEK inhibitor U0126. However, CREB phosphorylation by carbachol was largely unaffected by MAP kinase blockade. As both CREB phosphorylation and EGR1 induction have been linked to long-term potentiation and some forms of memory consolidation, these results may implicate CREB and EGR1 in independent or partially independent cholinergic signalling pathways involved in memory processes.
Department of Pharmacology, Faculty of Medicine and Health Science, University of Auckland, Auckland, New Zealand.
Jeffrey M
JM
Greenwood
Michael
M
Dragunow
D001285:Atropine / Q000494:pharmacology; D015153:Blotting, Western; D002217:Carbachol / Q000494:pharmacology; D018679:Cholinergic Agonists / Q000494:pharmacology; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D051766:Early Growth Response Protein 1; D006801:Humans; D017874:Immediate-Early Proteins; D007150:Immunohistochemistry; D017774:Long-Term Potentiation / Q000502:physiology; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D018727:Muscarinic Antagonists / Q000494:pharmacology; D009447:Neuroblastoma / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D009473:Neuronal Plasticity / Q000502:physiology; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism; D014407:Tumor Cells, Cultured
D018679:Cholinergic Agonists; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D018727:Muscarinic Antagonists; D011976:Receptors, Muscarinic; D014157:Transcription Factors; D001285:Atropine; D002217:Carbachol; C482128:MAP2K2 protein, human; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases