Acute Ischemic Injury of Astrocytes Is Mediated by Na-K-Cl Cotransport and not Ca2+Influx at a Key Point in White Matter Development

Cerebral palsy is a common birth disorder that frequently involves ischemic-type injury to developing white matter (WM). Dead glial cells are a common feature of this injury and here we describe a novel form of acute ischemic cell death in developing WM astrocytes. Ischemia, modeled by the withdrawal of oxygen and glucose, evoked [Ca 2+ ] i increases and cell death in astrocytes in post-natal day 10 (P10) rat optic nerve (RON). Removing extracellular Ca 2+ prevented increases in [Ca 2+ ] i but increased the amount of cell death. Astrocytes showed rapid [Na + ] i increases during ischemia and cell death was reduced to control levels by substitution of extracellular Na + or Cl or by perfusion with bumetanide, a selective Na-K-Cl cotransport (NKCC) blocker. Astrocytes showed marked swelling during ischemia in the absence of extracellular Ca 2+ , which was blocked by bumetanide. Raising the extracellular osmolarity to limit water uptake reduced ischemic astrocyte death to control levels. Ultrastructural examination showed that post-ischemic astrocytes had lost their processes and frequently were necrotic, effects partially prevented by bumetanide. At this point in development, therefore. NKCC activation in astrocytes during ischemia produces an osmo-regulatory challenge. Astrocytes can subsequently regulate their cell volume in a Ca 2+ -dependent fashion but this will require ATP hydrolysis and does not protect the cells against acute cell death.