The Role of Calpail-Mediated Spectrin Proteolysis in Traumatically Induced Axonal Injury

Abstract

In animals and man, traumatic brain injury (TBI) results in axonal injury (AI) that contributes to morbidity and mortality. Such injured axons show progressive change leading to axonal disconnection. Although several theories implicate calcium in the pathogenesis of AI, experimental studies have failed to confirm its pivotal role. To explore the contribution of Ca2+-induced proteolysis to axonal injury, this study was undertaken in an animal model of TBI employing antibodies targeting both calpain-mediated spectrin proteolysis (CMSP) and focal neurofilament compaction (NFC), a marker of intra-axonal cytoskeletal perturbation, at 15-120 minutes (min) postinjury. Light microscopy (LM) revealed that TBI consistently evoked focal, intra-axonal CMSP that was spatially and temporally correlated with NFC. These changes were seen at 15 min postinjury with significantly increasing number of axons demonstrating CMSP immunoreactivity over time postinjury. Electron microscopy (EM) demonstrated that at 15 min postinjury CMSP was confined primarily to the subaxolemmal network. With increasing survival (30-120 min) CMSP filled the axoplasm proper. These findings provide the first direct evidence for focal CMSP in the pathogenesis of generalized/diffuse AI. Importantly, they also reveal an initial subaxolemmal involvement prior to induction of a more widespread axoplasmic change indicating a spatial-temporal compartmentalization of the calcium-induced proteolytic process that may be amenable to rapid therapeutic intervention.

Gel-to-gel phase transition may occur in mammalian cells: Mechanism of formation of ?dark? (compacted) neurons

“…For axons, traumatic ultrastructural (neurofilament) compaction has been assumed by other authors (e.g. Buki et al, 1999) to be a morphological manifestation of calpain-mediated spectrin proteolysis, initiated by an uncontrolled influx of Ca ++ through the axolemma, perturbed focally by some traumatically generated intracerebral shearing force.…”

“…In accordance with this assumption, calpain-mediated spectrin proteolysis in compacted axons could be detected only in rats that survived for at least 15 min. after an in vivo head injury (Buki et al, 1999).…”

Wallerian-like axonal degeneration in the optic nerve after excitotoxic retinal insult: an ultrastructural study

“…It has been proposed that the decreased ATP and mitochondrial formation, usually seen with necrotic cell death, results in energy-dependent pump failure at active nodal sites causing ionic imbalance, focal cytoskeletal dissolution and neurofilament compaction; loss of membranous Ca 2+ -ATPase pump causing Ca 2+ influx induce calpain-mediated proteolysis of the subaxolemmal proteins which results in the formation of nodal blebs [61,62]. This proteolytic activity spreads to involve the entire nodal axoplasm [63] results in focal axonal swellings with variable amount of cytoskeletal disruption. Studies have also shown that proteolyic degradation of sidearms of neurofilaments results in their axoplasmic aggregation [64].…”

Pathophysiology of Cerebral Ischemia and Brain Trauma: Similarities and Differences

“…Several studies in cerebral ischemia and trauma have also implicated calcium-induced calpain-mediated proteolysis in the pathogenesis (Bartus et al, 1995;Buki et al, 1999a;Buki et al, 1999b;Hong et al, 1994;Posmantur et al, 1997;Saatman et al, 1996) of injury. The substrates for calpain effects have been shown to include many cytoskeletal proteins, membrane proteins, and various regulatory and signaling proteins.…”