ABSTRACT
Tyrosine kinase phosphorylation plays an important role in the induction of long-term potentiation (LTP). Focal adhesion kinase (FAK) is a 125 kDa nonreceptor tyrosine kinase that shows decreased phosphorylation in fyn mutant mice, and Fyn plays a critical role in LTP induction. By examining the role of FAK involved in LTP induction in dentate gyrus in vivo with medial perforant path stimulation, we found that both FAK and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) phosphorylation were increased significantly 5 and 10 min after LTP induction, whereas cAMP-responsive element binding protein (CREB) phosphorylation was increased 40 min later. Transfection of the dominant-negative FAK mutant construct HA-FAK(Y397F) impaired LTP, whereas transfection of the constitutively activated form HA-FAK(Delta1-100) reduced the threshold for LTP induction. Transfection of HA-FAK(Delta1-100) by itself did not induce long-lasting potentiation. Further, transfection of the HA-FAK(Y397F) construct decreased FAK, MAPK/ERK, and CREB phosphorylation, and the inhibition of MAPK/ERK decreased CREB phosphorylation. Moreover, blockade of NMDA receptor (NMDAR) did not decrease FAK, MAPK/ERK, and CREB phosphorylation although LTP induction was blunted by NMDAR antagonist. These biochemical changes were not associated with low-frequency stimulation either. Immunoprecipitation results revealed that tyrosine phosphorylation of NR2A and NR2B as well as the association of phosphorylated FAK with NR2A and NR2B was increased with LTP induction. These results together suggest that FAK is required, but not sufficient, for the induction of LTP in a NMDAR-independent manner and that MAPK/ERK and CREB are the downstream events of FAK activation. Further, FAK may interact with NR2A and NR2B to modulate LTP induction.
Subject(s)
Dentate Gyrus/cytology , Dentate Gyrus/enzymology , Long-Term Potentiation/physiology , Neurons/enzymology , Neurons/physiology , Protein-Tyrosine Kinases/physiology , Animals , Cyclic AMP Response Element-Binding Protein/metabolism , Cyclic AMP Response Element-Binding Protein/physiology , Dentate Gyrus/chemistry , Focal Adhesion Kinase 1 , Focal Adhesion Protein-Tyrosine Kinases , Male , Mitogen-Activated Protein Kinase 1/metabolism , Mitogen-Activated Protein Kinase 1/physiology , Mitogen-Activated Protein Kinase 3 , Mitogen-Activated Protein Kinases/metabolism , Mitogen-Activated Protein Kinases/physiology , Mutagenesis, Site-Directed , Neurons/chemistry , Phosphorylation , Plasmids , Precipitin Tests , Protein-Tyrosine Kinases/genetics , Protein-Tyrosine Kinases/metabolism , Proto-Oncogene Proteins/deficiency , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/physiology , Proto-Oncogene Proteins c-fyn , Rats , Rats, Mutant Strains , Rats, Sprague-Dawley , Reaction Time/genetics , Reaction Time/physiology , Receptors, Metabotropic Glutamate/physiology , Receptors, N-Methyl-D-Aspartate/physiology , Sensory Thresholds/physiology , Synaptosomes/metabolism , TransfectionABSTRACT
By using differential display PCR, we have identified 98 cDNA fragments from the rat dorsal hippocampus that are expressed differentially between the fast learners and slow learners in the water maze learning task. One of these cDNA fragments encodes the rat serum- and glucocorticoid-inducible kinase (sgk) gene. Northern blot analysis revealed that the sgk mRNA level was approximately 4-fold higher in the hippocampus of fast learners than slow learners. In situ hybridization results indicated that sgk mRNA level was increased markedly in CA1, CA3, and dentate gyrus of hippocampus in fast learners. Transient transfection of the sgk mutant DNA to the CA1 area impaired, whereas transfection of the sgk wild-type DNA facilitated water maze performance in rats. These results provide direct evidence that enhanced sgk expression facilitates memory consolidation of spatial learning in rats. These results also elucidate the molecular mechanism of glucocorticoid-induced memory facilitation in mammals.
