Neurochemical Research

, Volume 32, Issue 12, pp 2210–2216 | Cite as

Direct Evidence for Calpain Involvement in Apoptotic Death of Neurons in Spinal Cord Injury in Rats and Neuroprotection with Calpain Inhibitor

  • Eric A. Sribnick
  • Denise D. Matzelle
  • Naren L. Banik
  • Swapan K. Ray
Original Paper


To demonstrate calpain involvement in neurodegeneration in rat spinal cord injury (SCI), we examined SCI segments for DNA fragmentation, neurons for calpain overexpression, neuronal death, and neuroprotection with calpain inhibitor (E-64-d). After the induction of SCI (40 g cm force) on T12, rats were treated within 15 min with vehicle (DMSO) or E-64-d. Sham animals underwent laminectomy only. Animals were sacrificed at 24 h, and five 1-cm long spinal cord segments were collected: two rostral (S1 and S2), one lesion (S3), and two caudal segments (S4 and S5). Agarose gel electrophoresis of DNA samples isolated from the SCI segments showed both random and internucleosomal DNA fragmentation indicating occurrence of necrosis as well as apoptosis mostly in the lesion, moderately in caudal, and slightly in rostral segments from SCI rats. Treatment of SCI rats with E-64-d (1 mg/kg) reduced DNA fragmentation in all segments. The lesion and adjacent caudal segments (S3 and S4) were further investigated by in situ double-immunofluorescent labelings that showed increase in calpain expression in neurons in SCI rats and decrease in calpain expression in SCI rats treated with E-64-d. In situ combined TUNEL and double-immunofluorescent labelings directly detected co-localization of neuronal death and calpain overexpressin in SCI rats treated with only vehicle while attenuation of neuronal death in SCI rats treated with E-64-d. Previous studies from our laboratory indirectly showed neuroprotective effect of E-64-d in SCI rats. Our current results provide direct in situ evidence for calpain involvement in neuronal death and neuroprotective efficacy of E-64-d in lesion and penumbra in SCI rats.


Calpain E-64-d Immunofluorescent labelings Neuronal death SCI 



This work was supported in part by R01 grants (NS-31622, NS-57811, and CA-91460) and a Medical Scientist Training Program (MSTP) grant (GM08716) from the National Institutes of Health, and also a Spinal Cord Injury Research Fund grants (SCIRF-0803 and SCIRF-1205) from the state of South Carolina.


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Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Eric A. Sribnick
    • 1
  • Denise D. Matzelle
    • 1
  • Naren L. Banik
    • 1
  • Swapan K. Ray
    • 1
  1. 1.Department of NeurosciencesMedical University of South CarolinaCharlestonUSA

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