MECHANICAL INJURY TO NEURONAL GLIAL CULTURES IN MICROPLATES - ROLE OFNMDA RECEPTORS AND PH IN SECONDARY NEURONAL CELL-DEATH/

In vitro models of traumatic injury are useful adjuncts to animal mode ls for studying mechanisms of post-traumatic cell death, Here we descr ibe a new in vitro model in which reproducible levels of injury are de livered by a punch device that produces 28 parallel cuts in individual wells of 96-well microplates, Cell loss is measured by LDH assay or q uantitative fluorometric assay for ethidium homodimer staining, Glial cultures show cell death restricted to the initial injury site, wherea s neuronal/glial cultures demonstrate substantial spread of cell loss over time, We used this model to examine the role of pH and NMDA recep tors in delayed post-traumatic injury, NMDA receptor blockade by dizoc ilpine (MK-801) or treatment with antisense oligodeoxynucleotides dire cted against NMDAR1 was neuroprotective. Decreased cell death was obse rved under acidic conditions whereas increased extracellular pH was as sociated with increased, MK-801 sensitive cell loss, Advantages of our model include: reproducible trauma induction; rapid measurements of c ell injury; and use of 96-well microplates which reduce time and cost, This model appears to be well-suited for the study of selected mechan isms of post-traumatic neuronal injury as well as for screening potent ial neuroprotective agents, (C) 1998 Wiley-Liss, Inc.