Abstract
In the central nervous system, members of the Src family of tyrosine kinases (SFKs) are widely expressed and are abundant in neurons. The purpose of this study is to examine whether glycogen synthase-3 (GSK-3) is involved in SFK inhibitor-induced apoptosis. PP2 and SU6656, SFK inhibitors, increased apoptotic cell death with morphological changes that were characterized by cell shrinkage, chromatin condensation, or nuclear fragmentation. Moreover, both activation of caspase-9 and caspase-3 were accompanied by the cell death. GSK-3 inhibitors, such as alsterpaullone and SB216763, prevented the PP2-induced apoptosis. In addition, insulin-like growth factor-I prevented the PP2-induced cell death and PP2 inhibited phosphorylation of focal adhesion kinase (FAK). Phosphorylation of FAK on Tyr 576 by Src activates FAK. These results suggest that inhibition of SFK induces apoptosis possibly via blocking of FAK/phosphatidylinositol-3 kinase/Akt signaling pathway and activation of GSK-3 is involved in the cell death in rat cortical neurons.
Similar content being viewed by others
References
Alessi DR, James SR, Downes CP, Holmes AB, Gaffney PR, Reese CB, Cohen P (1997) Characterization of a 3-phosphoinositide-dependent protein kinase which phosphorylates and activates protein kinase Balpha. Curr Biol 7:261–269
Calalb MB, Polte TR, Hanks SK (1995) Tyrosine phosphorylation of focal adhesion kinase at sites in the catalytic domain regulates kinase activity: a role for Src family kinases. Mol Cell Biol 15:954–963
Choi DW, Koh JY, Peters S (1988) Pharmacology of glutamate neurotoxicity in cortical cell culture: attenuation by NMDA antagonists. J Neurosci 8:185–196
Cooper JA, MacAuley A (1988) Potential positive and negative autoregulation of p60c-src by intermolecular autophosphorylation. Proc Natl Acad Sci USA 85:4232–4236
Cross DA, Alessi DR, Cohen P, Andjelkovich M, Hemmings BA (1995) Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature 378:785–789
Crossthwaite AJ, Valli H, Williams RJ (2004) Inhibiting Src family tyrosine kinase activity blocks glutamate signalling to ERK1/2 and Akt/PKB but not JNK in cultured striatal neurones. J Neurochem 88:1127–1139
Dichter MA (1978) Rat cortical neurons in cell culture: culture methods, cell morphology, electrophysiology, and synapse formation. Brain Res 149:279–293
Girault JA, Costa A, Derkinderen P, Studler JM, Toutant M (1999) FAK and PYK2/CAKbeta in the nervous system: a link between neuronal activity, plasticity and survival? Trends Neurosci 22:257–263
Grimes CA, Jope RS (2001) The multifaceted roles of glycogen synthase kinase 3beta in cellular signaling. Prog Neurobiol 65:391–426
Hakem R, Hakem A, Duncan GS, Henderson JT, Woo M, Soengas MS, Elia A, de la Pompa JL, Kagi D, Khoo W, Potter J, Yoshida R, Kaufman SA, Lowe SW, Penninger JM, Mak TW (1998) Differential requirement for caspase 9 in apoptotic pathways in vivo. Cell 94:339–352
Hetman M, Cavanaugh JE, Kimelman D, Xia Z (2000) Role of glycogen synthase kinase-3beta in neuronal apoptosis induced by trophic withdrawal. J Neurosci 20:2567–2574
Hwang JY, Kim YH, Ahn YH, Wie MB, Koh JY (1999) N-Methyl-d-aspartate receptor blockade induces neuronal apoptosis in cortical culture. Exp Neurol 159:124–130
Ikonomidou C, Bosch F, Miksa M, Bittigau P, Vockler J, Dikranian K, Tenkova TI, Stefovska V, Turski L, Olney JW (1999) Blockade of NMDA receptors and apoptotic neurodegeneration in the developing brain. Science 283:70–74
Johnson D, Agochiya M, Samejima K, Earnshaw W, Frame M, Wyke J (2000) Regulation of both apoptosis and cell survival by the v-Src oncoprotein. Cell Death Differ 7:685–696
Kalia LV, Gingrich JR, Salter MW (2004) Src in synaptic transmission and plasticity. Oncogene 23:8007–8016
Kuida K, Zheng TS, Na S, Kuan C, Yang D, Karasuyama H, Rakic P, Flavell RA (1996) Decreased apoptosis in the brain and premature lethality in CPP32-deficient mice. Nature 384:368–372
Kuida K, Haydar TF, Kuan CY, Gu Y, Taya C, Karasuyama H, Su MS, Rakic P, Flavell RA (1998) Reduced apoptosis and cytochrome c-mediated caspase activation in mice lacking caspase 9. Cell 94:325–337
Lee M, Kim JY, Koh WS (2004) Apoptotic effect of PP2 a Src tyrosine kinase inhibitor, in murine B cell leukemia. J Cell Biochem 93:629–638
Linseman DA, Butts BD, Precht TA, Phelps RA, Le SS, Laessig TA, Bouchard RJ, Florez-McClure ML, Heidenreich KA (2004) Glycogen synthase kinase-3beta phosphorylates Bax and promotes its mitochondrial localization during neuronal apoptosis. J Neurosci 24:9993–10002
Morgan JC, Majors JE, Galileo DS (2000) Wild-type and mutant forms of v-src differentially alter neuronal migration and differentiation in vivo. J Neurosci Res 59:226–237
Pap M, Cooper GM (1998) Role of glycogen synthase kinase-3 in the phosphatidylinositol 3-kinase/Akt cell survival pathway. J Biol Chem 273:19929–19932
Stewart CE, Rotwein P (1996) Growth, differentiation, and survival: multiple physiological functions for insulin-like growth factors. Physiol Rev 76:1005–1026
Takadera T, Ohyashiki T (1997) Apoptotic cell death and caspase 3 (CPP32) activation induced by calcium ionophore at low concentrations and their prevention by nerve growth factor in PC12 cells. Eur J Biochem 249:8–12
Takadera T, Matsuda I, Ohyashiki T (1999) Apoptotic cell death and caspase-3 activation induced by N-methyl-d-aspartate receptor antagonists and their prevention by insulin-like growth factor I. J Neurochem 73:548–556
Takadera T, Sakamoto Y, Ohyashiki T (2004) NMDA receptor 2B-selective antagonist ifenprodil-induced apoptosis was prevented by glycogen synthase kinase-3 inhibitors in cultured rat cortical neurons. Brain Res 1020:196–203
Takadera T, Ishida A, Ohyashiki T (2006) Ketamine-induced apoptosis in cultured rat cortical neurons. Toxicol Appl Pharmacol 210:100–107
Thomas SM, Brugge JS (1997) Cellular functions regulated by Src family kinases. Annu Rev Cell Dev Biol 13:513–609
Thornberry NA, Rano TA, Peterson EP, Rasper DM, Timkey T, Garcia-Calvo M, Houtzager VM, Nordstrom PA, Roy S, Vaillancourt JP, Chapman KT, Nicholson DW (1997) A combinatorial approach defines specificities of members of the caspase family and granzyme B. Functional relationships established for key mediators of apoptosis. J Biol Chem 272:17907–17911
Wang YT, Salter MW (1994) Regulation of NMDA receptors by tyrosine kinases and phosphatases. Nature 369:233–235
Xia H, Nho RS, Kahm J, Kleidon J, Henke CA (2004) Focal adhesion kinase is upstream of phosphatidylinositol 3-kinase/Akt in regulating fibroblast survival in response to contraction of type I collagen matrices via a beta 1 integrin viability signaling pathway. J Biol Chem 279:33024–33034
Zhao WQ, Alkon DL, Ma W (2003) c-Src protein tyrosine kinase activity is required for muscarinic receptor-mediated DNA synthesis and neurogenesis via ERK1/2 and c-AMP-responsive element-binding protein signaling in neural precursor cells. J Neurosci Res 72:334–342
Acknowledgment
This study was supported in part by a special in-house research grant from Hokuriku University.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Takadera, T., Fujibayashi, M., Koriyama, Y. et al. Apoptosis Induced by Src-Family Tyrosine Kinase Inhibitors in Cultured Rat Cortical Cells. Neurotox Res 21, 309–316 (2012). https://doi.org/10.1007/s12640-011-9284-5
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12640-011-9284-5