Calpain activation and apoptosis in motor neurons of cultured adult mouse spinal cord.
Calpain, a Ca2+-dependent cysteine protease, has been implicated in neuronal apoptosis following spinal cord injury. In this study, activation of calpain was investigated in motor neurons of adult spinal cord slices from the mouse, using a cell-permeable fluorogenic calpain substrate and Western blotting. Calpain was rapidly activated in the motor neurons of excised spinal cord slices and calpain activity was observed both in the cytoplasm and the nuclei. In these neurons, nuclear and chromatin condensation were pronounced. Both calpain inhibitor VI and EGTA (ethyleneglycol-bis(beta-aminoethyl ether) N' ,N' ,N' ,N' -tetraacetic acid) inhibited calpain activation and subsequent appearance of apoptotic nuclei. In contrast, the general caspase inhibitor Z-VAD.fmk had no effect. Calpain activation was also observed in the slices by Western blotting using an antibody to 150-kD calpain-cleaved alpha-fodrin fragment. These results show that calpain is rapidly activated in injured motor neurons and imply that this activation could be responsible for execution of caspase-independent apoptosis in injured adult motor neurons.
Momeni
Hamid Reza
HR
Department of Biology, Faculty of Science, Arak University, Arak, Iran. h-momeni@araku.ac.ir
Azadi
Seifollah
S
Kanje
Martin
M
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004396:Coloring Agents; D004533:Egtazic Acid / Q000494:pharmacology; D004789:Enzyme Activation / Q000502:physiology; D005260:Female; D006023:Glycoproteins / Q000494:pharmacology; D007150:Immunohistochemistry; D007202:Indicators and Reagents; D051379:Mice; D008840:Microfilament Proteins / Q000737:chemistry; D009046:Motor Neurons / Q000502:physiology* / Q000648:ultrastructure; D011419:Propidium; D013116:Spinal Cord / Q000166:cytology* / Q000648:ultrastructure; D016707:Tissue Fixation; D013049:Spectrin
D002154:Calpain; D002352:Carrier Proteins; D002614:Chelating Agents; D004396:Coloring Agents; D004533:Egtazic Acid; D006023:Glycoproteins; D007202:Indicators and Reagents; D008840:Microfilament Proteins; D011419:Propidium; D013049:Spectrin; C039304:calpain inhibitors