IMMUNOLOCALIZATION OF CALPAIN I-MEDIATED SPECTRIN DEGRADATION TO VULNERABLE NEURONS IN THE ISCHEMIC GERBIL BRAIN

Transient ischemia-induced perturbations in calcium homeostasis have b een proposed to lead to pathological activation of the cysteine protea se calpain I and subsequent delayed neuronal death in the CA1 region o f hippocampus. We report here on the design and characterization of an tibodies selective for calpain-generated fragments of brain spectrin, and their use for immunoblot and immunohistochemical analyses of calpa in activation following cerebral ischemia in the gerbil. Although spec trin was susceptible to degradation in vitro by many mammalian proteas es, only calpain degraded spectrin to generate fragments immunoreactiv e with the antibodies. Following 5 min of global ischemia, immunoreact ivity for calpain-degraded spectrin was rapidly (within 30 min) and ma rkedly elevated in the perikarya and dendrites of several populations of forebrain neurons. The rapid calpain activation was completely prev ented by the NMDA receptor antagonist MK-801. At later times postische mia, but prior to frank neuronal necrosis, calpain-degraded spectrin w as restricted to hippocampal area CA1 pyramidal neurons. Silver impreg nation histochemistry confirmed that neuronal damage was confined to a rea CA1. The results indicate that while nonpathological NMDA receptor stimulation can activate calpain, only those neurons showing sustaine d calpain activation are destined to die.