Increased calpain I-mediated proteolysis, and preferential loss of dephosphorylated NF200, following traumatic spinal cord injury.
We investigated the hypothesis that the Ca2+-activated protease calpain is involved in the pathophysiology of spinal cord injury, and is linked to the proteolytic degradation of cytoskeletal proteins. We report here that levels of calpain I (mu-calpain)-mediated spectrin breakdown products are increased by 15 min post-injury, with peak levels reached by 2 h post-injury. The dephosphorylated form of the neurofilament protein NF200 is substantially lost over the same time-period. A 35-g compressive injury was applied to the midthoracic rat spinal cord for 1 min, and animals were killed at 15 min, 1, 2, 4, 8, 16, and 24 h post-injury. Calpain I-mediated spectrin breakdown products accumulated post-injury, with peak levels reached at 2 h. Secondly, we have demonstrated a progressive loss of the 200,000 mol. wt neurofilament protein NF200, a cytoskeletal calpain substrate, which began within 1-2 h post-injury. Densitometric analyses confirmed that loss of NF200 is a substrate-specific phenomenon, since (i) dephosphorylated NF200 was preferentially lost while phosphorylated NF200 was relatively spared, and (ii) actin, which is not a substrate for calpain, was relatively spared following spinal cord injury. Finally, we demonstrated calpain I-mediated spectrin breakdown within NF200-positive neuronal processes post-injury. We conclude that the accumulation of spectrin breakdown products is temporally and spatially correlated with loss of dephosphorylated NF200 after spinal cord injury.
Schumacher
P A
PA
Playfair Neuroscience Unit and The Toronto Hospital Research Institute, University of Toronto, The Toronto Hospital, Ontario, Canada.
Eubanks
J H
JH
Fehlings
M G
MG
D000818:Animals; D001369:Axons / Q000473:pathology / Q000648:ultrastructure; D002154:Calpain / Q000378:metabolism*; D005260:Female; D007150:Immunohistochemistry; D016900:Neurofilament Proteins / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D017208:Rats, Wistar; D013049:Spectrin / Q000378:metabolism; D013116:Spinal Cord / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D013119:Spinal Cord Injuries / Q000378:metabolism* / Q000473:pathology; D013997:Time Factors
D016900:Neurofilament Proteins; C051774:neurofilament protein H; D013049:Spectrin; D002154:Calpain