10779/rcsi.10791338.v1
Beatrice D'Orsi
Beatrice
D'Orsi
Helena Bonner
Helena
Bonner
Liam P. Tuffy
Liam P.
Tuffy
Heiko Düssmann
Heiko
Düssmann
Ina Woods
Ina
Woods
Michael J. Courtney
Michael J.
Courtney
Manus W. Ward
Manus W.
Ward
Jochen HM Prehn
Jochen HM
Prehn
Calpains are downstream effectors of bax-dependent excitotoxic apoptosis.
Royal College of Surgeons in Ireland
2019
Animals
Apoptosis
Calpain
Cell Line
Tumor
Cells
Cultured
Dipeptides
Excitatory Amino Acid Agonists
Female
Hippocampus
Humans
Mice
129 Strain
Inbred C57BL
Knockout
N-Methylaspartate
Organ Culture Techniques
Pregnancy
bcl-2-Associated X Protein
Physiology
Medical Physics
2019-11-22 16:58:31
Journal contribution
https://repository.rcsi.com/articles/journal_contribution/Calpains_are_downstream_effectors_of_bax-dependent_excitotoxic_apoptosis_/10791338
<p>Excitotoxicity resulting from excessive Ca(2+) influx through glutamate receptors contributes to neuronal injury after stroke, trauma, and seizures. Increased cytosolic Ca(2+) levels activate a family of calcium-dependent proteases with papain-like activity, the calpains. Here we investigated the role of calpain activation during NMDA-induced excitotoxic injury in embryonic (E16-E18) murine cortical neurons that (1) underwent excitotoxic necrosis, characterized by immediate deregulation of Ca(2+) homeostasis, a persistent depolarization of mitochondrial membrane potential (Δψ(m)), and insensitivity to bax-gene deletion, (2) underwent excitotoxic apoptosis, characterized by recovery of NMDA-induced cytosolic Ca(2+) increases, sensitivity to bax gene deletion, and delayed Δψ(m) depolarization and Ca(2+) deregulation, or (3) that were tolerant to excitotoxic injury. Interestingly, treatment with the calpain inhibitor calpeptin, overexpression of the endogenous calpain inhibitor calpastatin, or gene silencing of calpain protected neurons against excitotoxic apoptosis but did not influence excitotoxic necrosis. Calpeptin failed to exert a protective effect in bax-deficient neurons but protected bid-deficient neurons similarly to wild-type cells. To identify when calpains became activated during excitotoxic apoptosis, we monitored calpain activation dynamics by time-lapse fluorescence microscopy using a calpain-sensitive Förster resonance energy transfer probe. We observed a delayed calpain activation that occurred downstream of mitochondrial engagement and directly preceded neuronal death. In contrast, we could not detect significant calpain activity during excitotoxic necrosis or in neurons that were tolerant to excitotoxic injury. Oxygen/glucose deprivation-induced injury in organotypic hippocampal slice cultures confirmed that calpains were specifically activated during bax-dependent apoptosis and in this setting function as downstream cell-death executioners.</p>