Different mechanisms of NMDA-mediated protection against neuronal apoptosis: a stimuli-dependent effect.
The mechanisms of protective effect of N-methyl-D-aspartate (NMDA) receptor stimulation on apoptosis of neurons at their early stage of development are poorly understood. In the present study, we investigated the effects of NMDA on staurosporine (St)- and low-potassium (LP)-evoked apoptotic cell death in primary cerebellar granule cell (CGC) cultures at 7 days in vitro (DIV). We found that NMDA (200 microM) attenuated the St (0.5 microM)- and LP (5 mM KCl)-induced neuronal cell death in 7 but not 12 DIV CGC as confirmed by LDH release and MTT reduction assays. Moreover, NMDA attenuated St-and LP-evoked DNA fragmentation and cytosolic apoptosis inducing factor (AIF) protein level but not caspase-3 activation induced by both pro-apoptotic factors. Neuroprotective effects of NMDA on St-induced apoptosis in CGC were attenuated by inhibitors of ERK/MAPK-signaling, PD 98059 and U0126 but not by NMDA receptor antagonists, AP-5 (100 microM) and MK-801 (1 microM) or by inhibitors of PI3-K/Akt pathway (LY 294002 and wortmannin). In contrast to staurosporine model of apoptosis, AP-5 and MK-801 but not inhibitors of PI3-K/Akt and MAPK/ERK1/2 prevented the NMDA-mediated neuroprotection in LP-induced apoptosis of CGC. In separate experiments, we observed also the anti-apoptotic action of NMDA on St (0.5 microM)- and salsolinol (250 microM)-evoked cell death in human neuroblastoma SH-SY5Y cells without its influence on caspase-3 activity, induced by these pro-apoptotic factors. These data indicate that neuroprotection evoked by NMDA in CGC strongly depends on used pro-apoptotic agent and could engage NMDA channel function or be connected with the activation of pro-survival MAPK/ERK1/2 pathway. It is also suggested that anti-apoptotic effects of NMDA is connected with inhibition of fragmentation of DNA via caspase-3-independent mechanism.
Jantas
Danuta
D
Department of Experimental Neuroendocrinology, Institute of Pharmacology, Polish Academy of Sciences, Krakow, Poland. jantas@if-pan.krakow.pl
Lason
Wladyslaw
W
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000502:physiology; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000502:physiology; D016202:N-Methylaspartate / Q000494:pharmacology / Q000502:physiology*; D009474:Neurons / Q000166:cytology* / Q000187:drug effects; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011188:Potassium / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors / Q000502:physiology; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology
D053148:Caspase 3; D007546:Isoquinolines; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D016202:N-Methylaspartate; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011188:Potassium; D051057:Proto-Oncogene Proteins c-akt; D016194:Receptors, N-Methyl-D-Aspartate; D019311:Staurosporine; C036617:salsolinol