Abstract
Using hippocampal primary cell cultures at 14 days in vitro (div), we have investigated actions of 17-beta estradiol (E; 10 nM) on the phosphorylation of CREB and on signaling pathways that regulate CREB phosphorylation. After demonstrating that 14 div is optimal for these studies, we examined the time course of E induction of CREB phosphorylation (pCREB) at serine residue 133. The induction of pCREB occurs as early as 1 h following E treatment, presumably via a mechanism involving an E-stimulated signal transduction system, which is sustained for at least 24 h but inhibited by 48 h. The early activity may represent an initial signal required for events leading to phosphorylation of CREB while the sustained signal may lead to CREB-mediated gene expression for cell survival and synapse formation. Furthermore, we examined the pathways for E action preceding pCREB induction by blocking three major kinases (protein kinase; mitogen activated protein kinase, MAPK; and calcium-calmodulin kinase II, CaMKII) upstream of pCREB. We found that E stimulates each pathway at 24 h and that phosphorylation of CREB is dependent on both MAPK and CaMK activities, but less dependent on the Akt pathway. Because CREB has been linked to E induction of excitatory spine synapses, we used a spine marker, spinophilin, to establish E effects on spine formation. Spinophilin expression was up-regulated in response to E and this effect was blocked by an inhibitor of (CaMKII). These studies demonstrate the central role played by CaMKII pathway in the actions of E on both transcriptional regulation and structural reorganization in neurons.
Publication types
-
Research Support, U.S. Gov't, P.H.S.
MeSH terms
-
Animals
-
Calcium-Calmodulin-Dependent Protein Kinase Type 2
-
Calcium-Calmodulin-Dependent Protein Kinases / antagonists & inhibitors
-
Calcium-Calmodulin-Dependent Protein Kinases / metabolism
-
Cell Differentiation / drug effects
-
Cell Differentiation / physiology
-
Cells, Cultured
-
Cyclic AMP Response Element-Binding Protein / drug effects
-
Cyclic AMP Response Element-Binding Protein / metabolism*
-
Dendrites / drug effects
-
Dendrites / metabolism
-
Dendrites / ultrastructure
-
Enzyme Inhibitors / pharmacology
-
Estrogens / metabolism*
-
Estrogens / pharmacology
-
Fetus
-
Genes, Regulator / drug effects
-
Genes, Regulator / genetics
-
Hippocampus / cytology
-
Hippocampus / embryology*
-
Hippocampus / metabolism*
-
Microfilament Proteins / metabolism
-
Mitogen-Activated Protein Kinase 1 / antagonists & inhibitors
-
Mitogen-Activated Protein Kinase 1 / metabolism
-
Nerve Tissue Proteins / metabolism
-
Neurons / drug effects
-
Neurons / metabolism*
-
Phosphorylation / drug effects
-
Protein Kinase Inhibitors
-
Protein Kinases / metabolism
-
Protein Serine-Threonine Kinases*
-
Proto-Oncogene Proteins / antagonists & inhibitors
-
Proto-Oncogene Proteins / metabolism
-
Proto-Oncogene Proteins c-akt
-
Rats
-
Reaction Time / drug effects
-
Reaction Time / physiology
-
Signal Transduction / drug effects
-
Signal Transduction / physiology
Substances
-
Cyclic AMP Response Element-Binding Protein
-
Enzyme Inhibitors
-
Estrogens
-
Microfilament Proteins
-
Nerve Tissue Proteins
-
Protein Kinase Inhibitors
-
Proto-Oncogene Proteins
-
neurabin
-
Protein Kinases
-
Akt1 protein, rat
-
Protein Serine-Threonine Kinases
-
Proto-Oncogene Proteins c-akt
-
Calcium-Calmodulin-Dependent Protein Kinase Type 2
-
Calcium-Calmodulin-Dependent Protein Kinases
-
Mitogen-Activated Protein Kinase 1