Activity-Dependent Neuroprotection and cAMP Response Element-Binding Protein (CREB): Kinase Coupling, Stimulus Intensity, and Temporal Regulation of CREB Phosphorylation at Serine 133

The dual nature of the NMDA receptor as a mediator of excitotoxic cell death and activity-dependent cell survival likely results from divergent patterns of kinase activation, transcription factor activation, and gene expression. To begin to address this divergence, we examined cellular and molecular signaling events that couple excitotoxic and nontoxic levels of NMDA receptor stimulation to activation of the cAMP response element-binding protein (CREB)/cAMP response element (CRE) pathway in cultured cortical neurons. Pulses (10 min) of NMDA receptor-mediated synaptic activity (nontoxic) triggered sustained (up to 3 h) CREB phosphorylation (pCREB) at serine 133. In contrast, brief stimulation with an excitotoxic concentration of NMDA (50 μm) triggered transient pCREB. The duration of pCREB was dependent on calcineurin activity. Excitotoxic levels of NMDA stimulated calcineurin activity, whereas synaptic activity did not. Calcineurin inhibition reduced NMDA toxicity and converted the transient increase in pCREB into a sustained increase. In accordance with these observations, sustained pCREB (up to 3 h) did not require persistent kinase pathway activity. The sequence of stimulation with excitotoxic levels of NMDA and neuroprotective synaptic activity determined which stimulus exerted control over pCREB duration. Constitutively active and dominant-negative CREB constructs were used to implicate CREB in synaptic activity-dependent neuroprotection against NMDA-induced excitotoxicity. Together these data provide a framework to begin to understand how the neuroprotective and excitotoxic effects of NMDA receptor activity function in an antagonistic manner at the level of the CREB/CRE transcriptional pathway.

[1]  N. Sousa,et al.  Ionotropic and metabotropic glutamate receptor mediation of glucocorticoid-induced apoptosis in hippocampal cells and the neuroprotective role of synaptic N-methyl-d-aspartate receptors , 2003, Neuroscience.

[2]  H. Enslen,et al.  Differential activation of CREB by Ca2+/calmodulin-dependent protein kinases type II and type IV involves phosphorylation of a site that negatively regulates activity. , 1994, Genes & development.

[3]  G. Bustos,et al.  BDNF gene transcripts in mesencephalic neurons and its differential regulation by NMDA , 1998, Neuroreport.

[4]  M. Catsicas,et al.  Rapid onset of neuronal death induced by blockade of either axoplasmic transport or action potentials in afferent fibers during brain development , 1992, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[5]  M. Dichter,et al.  Properties of inhibitory and excitatory synapses between hippocampal neurons in very low density cultures , 1994, Synapse.

[6]  R. Balázs,et al.  N-methyl-d-aspartate promotes the survival of cerebellar granule cells in culture , 1988, Neuroscience.

[7]  J. Morán,et al.  Mechanisms of cell death by deprivation of depolarizing conditions during cerebellar granule neurons maturation , 2003, Neurochemistry International.

[8]  D. Storm,et al.  Phosphorylation of cAMP Response Element-Binding Protein in Hippocampal Neurons as a Protective Response after Exposure to Glutamate In Vitro and Ischemia In Vivo , 2001, The Journal of Neuroscience.

[9]  T. Soderling,et al.  Characterization of Ca2+/calmodulin-dependent protein kinase IV. Role in transcriptional regulation. , 1994, The Journal of biological chemistry.

[10]  Michael J. O'Donovan,et al.  The effects of excitatory amino acids and their antagonists on the generation of motor activity in the isolated chick spinal cord. , 1987, Brain research.

[11]  D. Choi,et al.  Quantitative determination of glutamate mediated cortical neuronal injury in cell culture by lactate dehydrogenase efflux assay , 1987, Journal of Neuroscience Methods.

[12]  S. Paul,et al.  N-methyl-D-aspartate receptor-mediated neuroprotection in cerebellar granule cells requires new RNA and protein synthesis. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[13]  A. N. van den Pol,et al.  GABAB receptor-mediated inhibition of GABAA receptor calcium elevations in developing hypothalamic neurons. , 1998, Journal of neurophysiology.

[14]  G. Crabtree.  Generic Signals and Specific Outcomes Signaling through Ca2+, Calcineurin, and NF-AT , 1999, Cell.

[15]  K. Deisseroth,et al.  L-type calcium channels and GSK-3 regulate the activity of NF-ATc4 in hippocampal neurons , 1999, Nature.

[16]  L. Kaczmarek,et al.  Inducible cAMP Early Repressor, an Endogenous Antagonist of cAMP Responsive Element-Binding Protein, Evokes Neuronal Apoptosis In Vitro , 2003, The Journal of Neuroscience.

[17]  Steven Finkbeiner,et al.  Ca2+ Influx Regulates BDNF Transcription by a CREB Family Transcription Factor-Dependent Mechanism , 1998, Neuron.

[18]  Michael E. Greenberg,et al.  Coupling of the RAS-MAPK Pathway to Gene Activation by RSK2, a Growth Factor-Regulated CREB Kinase , 1996, Science.

[19]  D. Storm,et al.  Stimulation of cAMP response element (CRE)-mediated transcription during contextual learning , 1998, Nature Neuroscience.

[20]  R. Fisher,et al.  Nuclear and axonal localization of Ca2+/calmodulin-dependent protein kinase type Gr in rat cerebellar cortex. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[21]  L. Boxer,et al.  Induction of bcl-2 expression by phosphorylated CREB proteins during B-cell activation and rescue from apoptosis , 1996, Molecular and cellular biology.

[22]  Jean-Claude Martinou,et al.  Overexpression of BCL-2 in transgenic mice protects neurons from naturally occurring cell death and experimental ischemia , 1994, Neuron.

[23]  K. Deisseroth,et al.  Activity-dependent CREB phosphorylation: Convergence of a fast, sensitive calmodulin kinase pathway and a slow, less sensitive mitogen-activated protein kinase pathway , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[24]  P. Gluckman,et al.  The role of the cyclic AMP-responsive element binding protein (CREB) in hypoxic-ischemic brain damage and repair. , 1996, Brain research. Molecular brain research.

[25]  D. K. Berg,et al.  Voltage-Gated Channels Block Nicotinic Regulation of CREB Phosphorylation and Gene Expression in Neurons , 2001, Neuron.

[26]  G. Steinberg,et al.  Overexpression of Bcl-2 with herpes simplex virus vectors protects CNS neurons against neurological insults in vitro and in vivo , 1996, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[27]  A. N. van den Pol,et al.  GABA neurotransmission in the hypothalamus: developmental reversal from Ca2+ elevating to depressing , 1995, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[28]  A. Graybiel,et al.  Spatiotemporal Dynamics of CREB Phosphorylation: Transient versus Sustained Phosphorylation in the Developing Striatum , 1996, Neuron.

[29]  Philip R. Cohen,et al.  MSK1 is required for CREB phosphorylation in response to mitogens in mouse embryonic stem cells , 2000, FEBS letters.

[30]  K. Fukunaga,et al.  Activation of CA(2+)/calmodulin-dependent protein kinase IV in cultured rat hippocampal neurons. , 2000, Journal of neuroscience research.

[31]  R. Linden,et al.  Activation of NMDA receptors protects against glutamate neurotoxicity in the retina: evidence for the involvement of neurotrophins , 1999, Brain Research.

[32]  K. Obrietan,et al.  Embryonic hypothalamic expression of functional glutamate receptors , 1995, Neuroscience.

[33]  D. Ginty,et al.  A Dominant-Negative Inhibitor of CREB Reveals that It Is a General Mediator of Stimulus-Dependent Transcription of c-fos , 1998, Molecular and Cellular Biology.

[34]  P. Sassone-Corsi,et al.  Rsk-2 activity is necessary for epidermal growth factor-induced phosphorylation of CREB protein and transcription of c-fos gene. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[35]  L. Maffei,et al.  Patterned Vision Causes CRE-Mediated Gene Expression in the Visual Cortex through PKA and ERK , 2003, The Journal of Neuroscience.

[36]  A. Edelman,et al.  Phosphorylation and activation of Ca(2+)-calmodulin-dependent protein kinase IV by Ca(2+)-calmodulin-dependent protein kinase Ia kinase. Phosphorylation of threonine 196 is essential for activation. , 1995, The Journal of biological chemistry.

[37]  Mark Farrant,et al.  NMDA receptor subunits: diversity, development and disease , 2001, Current Opinion in Neurobiology.

[38]  Anirvan Ghosh,et al.  Identification of a Signaling Pathway Involved in Calcium Regulation of BDNF Expression , 1998, Neuron.

[39]  J. Barker,et al.  Rat hippocampal neurons in culture: voltage-clamp analysis of inhibitory synaptic connections. , 1984, Journal of neurophysiology.

[40]  P. Gass,et al.  Expression of activating transcription factor-2, serum response factor and cAMP/Ca response element binding protein in the adult rat brain following generalized seizures, nerve fibre lesion and ultraviolet irradiation , 1997, Neuroscience.

[41]  A. J. Bower,et al.  ROLE OF AFFERENTS IN THE DEVELOPMENT AND CELL SURVIVAL OF THE VERTEBRATE NERVOUS SYSTEM , 1998, Clinical and experimental pharmacology & physiology.

[42]  K. Fukunaga,et al.  Activation of CA2+/calmodulin‐dependent protein kinase IV in cultured rat hippocampal neurons , 2000, Journal of neuroscience research.

[43]  A. N. van den Pol,et al.  Early synaptogenesis in vitro: Role of axon target distance , 1998, The Journal of comparative neurology.

[44]  Y. Ben-Ari.  Excitatory Amino Acids and Neuronal Plasticity , 1990, Advances in Experimental Medicine and Biology.

[45]  J. Barker,et al.  Glutamate Acting at NMDA Receptors Stimulates Embryonic Cortical Neuronal Migration , 1999, The Journal of Neuroscience.

[46]  D. Storm,et al.  CRE‐mediated gene transcription in the peri‐infarct area after focal cerebral ischemia in mice , 2004, Journal of neuroscience research.

[47]  T. Sick,et al.  εPKC Is Required for the Induction of Tolerance by Ischemic and NMDA-Mediated Preconditioning in the Organotypic Hippocampal Slice , 2003, The Journal of Neuroscience.

[48]  S. Cohen-Cory.  The Developing Synapse: Construction and Modulation of Synaptic Structures and Circuits , 2002, Science.

[49]  M. Sheng,et al.  Developmentally Regulated NMDA Receptor-Dependent Dephosphorylation of cAMP Response Element-Binding Protein (CREB) in Hippocampal Neurons , 2000, The Journal of Neuroscience.

[50]  R. Nicoll,et al.  NMDA-receptor-dependent synaptic plasticity: multiple forms and mechanisms , 1993, Trends in Neurosciences.

[51]  Kortaro Tanaka,et al.  Phosphorylation of Cyclic Adenosine Monophosphate Response Element Binding Protein in Oligodendrocytes in the Corpus Callosum after Focal Cerebral Ischemia in the Rat , 2001, Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism.

[52]  I. Kameshita,et al.  Purification and characterization of a brain-specific multifunctional calmodulin-dependent protein kinase from rat cerebellum. , 1992, The Journal of biological chemistry.

[53]  Mark Ellisman,et al.  Persistent phosphorylation of cyclic amp responsive element-binding protein and activating transcription factor-2 transcription factors following transient cerebral ischemia in rat brain , 1999, Neuroscience.

[54]  T. Soderling,et al.  CaM-kinases: modulators of synaptic plasticity , 2000, Current Opinion in Neurobiology.

[55]  S. R. Datta,et al.  Cell survival promoted by the Ras-MAPK signaling pathway by transcription-dependent and -independent mechanisms. , 1999, Science.

[56]  L. Verburgt,et al.  Overexpression of Bcl-2 and mutations in p53 and K-ras in resected human non-small cell lung cancers. , 1996, American journal of respiratory cell and molecular biology.

[57]  S. Nakanishi.  Molecular diversity of glutamate receptors and implications for brain function. , 1992, Science.

[58]  A. N. van den Pol,et al.  Excitatory actions of GABA increase BDNF expression via a MAPK-CREB-dependent mechanism--a positive feedback circuit in developing neurons. , 2002, Journal of neurophysiology.

[59]  H. Bading,et al.  Extrasynaptic NMDARs oppose synaptic NMDARs by triggering CREB shut-off and cell death pathways , 2002, Nature Neuroscience.

[60]  D. Ginty,et al.  Function and Regulation of CREB Family Transcription Factors in the Nervous System , 2002, Neuron.

[61]  J. Hamada,et al.  CREB is required for acquisition of ischemic tolerance in gerbil hippocampal CA1 region , 2003, Journal of neurochemistry.

[62]  M. Dragunow,et al.  CREB Phosphorylation Promotes Nerve Cell Survival , 1999, Journal of neurochemistry.

[63]  C. M. Davenport,et al.  Mediation by a CREB family transcription factor of NGF-dependent survival of sympathetic neurons. , 1999, Science.

[64]  S. Paul,et al.  N-methyl-D-aspartate exposure blocks glutamate toxicity in cultured cerebellar granule cells. , 1992, Molecular pharmacology.

[65]  K. Deisseroth,et al.  CREB Phosphorylation and Dephosphorylation: A Ca2+- and Stimulus Duration–Dependent Switch for Hippocampal Gene Expression , 1996, Cell.

[66]  A. Craig,et al.  Clustering of Gephyrin at GABAergic but Not Glutamatergic Synapses in Cultured Rat Hippocampal Neurons , 1996, The Journal of Neuroscience.

[67]  C. Shatz,et al.  Synaptic Activity and the Construction of Cortical Circuits , 1996, Science.

[68]  J. Kornhauser,et al.  Nerve Growth Factor Activates Extracellular Signal-Regulated Kinase and p38 Mitogen-Activated Protein Kinase Pathways To Stimulate CREB Serine 133 Phosphorylation , 1998, Molecular and Cellular Biology.

[69]  Eric C. Griffith,et al.  Regulation of transcription factors by neuronal activity , 2002, Nature Reviews Neuroscience.