Activation of c-Jun NH2-terminal kinase 3 is mediated by the GluR6.PSD-95.MLK3 signaling module following cerebral ischemia in rat hippocampus.
Kainate receptor glutamate receptor 6 (GluR6) binds to the postsynaptic density protein 95 (PSD-95), which in turn anchors mixed lineage kinase 3 (MLK3) via SH3 domain in rat brain tissue. MLK3 subsequently activates c-Jun NH(2)-terminal kinase (JNK) via MAP kinase kinases (MKKs). We investigated the association of PSD-95 with GluR6 and MLK3, MLK3 autophosphorylation, the interaction of MLK3 with JNK3, and JNK3 phosphorylation following cerebral ischemia in rat hippocampus. Our results indicate that the GluR6.PSD-95.MLK3 complex peaked at 6 h of reperfusion. Furthermore, MLK3 autophosphorylation and the interaction of MLK3 with JNK3 occurred with the alteration of GluR6.PSD-95.MLK3 signaling module. To further prove whether JNK3 activation in ischemic hippocampus is mediated by GluR6.PSD-95.MLK3 signaling pathway, the AMPA/KA receptor antagonist 6,7-dinitroquinoxaline-2, (1H, 4H)-dione (DNQX), the GluR6 antagonist 6,7,8,9-Tetrahydro-5-nitro-1H-benz[g]indole-2,3-dione-3-oxime (NS102), the AMPA receptor antagonist 1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzo diazepine (GYKI52466), and the NMDA receptor antagonist ketamine were given to the rats 20 min prior to ischemia. Our findings indicate that both DNQX and NS102 significantly attenuated the association of PSD-95 with GluR6 and MLK3, MLK3 autophosphorylation, interaction of MLK3 with JNK3, and JNK3 phosphorylation, while GYKI52466 and ketamine had no effect. Moreover, administration of NS102 before cerebral ischemia significantly increased the number of the surviving hippocampal CA1 pyramidal cells at 5 days of reperfusion. Consequently, GluR6, one subunit of kainate receptor, plays a critical role in inducing JNK3 activation after ischemic injury.
Tian
Hui
H
School of Life Science, Xuzhou Normal University, 29 Shanghai Road of Tongshan district, Xuzhou, Jiangsu 2211162, PR China.
Zhang
Quan-Guang
QG
Zhu
Guang-Xuan
GX
Pei
Dong-Sheng
DS
Guan
Qiu-Hua
QH
Zhang
Guang-Yi
GY
D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002545:Brain Ischemia / Q000201:enzymology*; D000076122:Disks Large Homolog 4 Protein; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006624:Hippocampus / Q000201:enzymology*; D047468:Immunoprecipitation / Q000379:methods; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D020930:MAP Kinase Kinase Kinases; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D048057:Mitogen-Activated Protein Kinase 10 / Q000378:metabolism*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D018092:Kainic Acid Receptors / Q000378:metabolism*; D015424:Reperfusion / Q000379:methods; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013997:Time Factors; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11; D000097806:GluK2 Kainate Receptor
D000076122:Disks Large Homolog 4 Protein; D018691:Excitatory Amino Acid Antagonists; D047908:Intracellular Signaling Peptides and Proteins; D020930:MAP Kinase Kinase Kinases; D008565:Membrane Proteins; D048057:Mitogen-Activated Protein Kinase 10; D020929:Mitogen-Activated Protein Kinase Kinases; D018092:Kainic Acid Receptors; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11; D000097806:GluK2 Kainate Receptor; C501449:Dlg4 protein, rat