Estradiol reduction of ischemia-induced cerebral edema: Role of astrocyte aquaporin 4, sodium potassium chloride cotransporter, and sodium hydrogen exchanger

In the early hours of ischemic stroke, edema formation occurs in the presence of an intact blood-brain barrier (BBB). During this time Na, Cl and osmotically obliged water are secreted across the barrier from blood into brain and are rapidly taken up by astrocytes causing these cells to swell. Results reported by other labs suggest that astrocyte swelling may be due to ischemia-stimulated Na-K-Cl cotransporter (NKCC) and Na/H exchanger (NHE) activity. Furthermore, the aquaporin 4 (AQP4) water channel, found in high abundance in the perivascular membrane of astrocytes, is believed to play a role in edema formation and resolution. While recent studies have raised questions about the effect hormone replacement therapy has on stroke, there is significant evidence that estradiol acts as a neuroprotectant in ischemic stroke. Studies from our lab have shown that estradiol reduces edema formation and infarct volume, possibly by reducing ischemia-induced stimulation of NKCC and NHE in BBB endothelial cells. For the studies reported herein, we hypothesized that estradiol reduces edema formation by altering NKCC or NHE activity and/or by reducing the abundance of AQP4 found in cortical astrocytes. Initial experiments using 3-O-methyl-D-[3H]glucose as a marker for total water space showed that vasopressin (AVP), hypoxia, aglycemia and oxygen-glucose deprivation (OGD) all induced astrocyte swelling and that the NKCC and NHE both participated in the AVP- but not aglycemia-induced astrocyte swelling. Estradiol treatment abolished the AVP- and hypoxia-induced astrocyte-swelling but did not alter the cell-swelling effects induced by aglycemia or OGD. Western blot analyses revealed that estradiol treatment (100 nM for 7 days) reduced cortical astrocyte AQP4 protein abundance while AVP and aglycemia did not alter NKCC and NHE abundance. Using a 86 Rb flux assay, we found that although ischemic conditions appeared to increase NKCC activity in cultured astrocytes, this increase was not found to be significant. These findings suggest that ischemia-induced astrocyte swelling appears to contribute to edema formation and that estradiol reduces ischemia-induced edema by attenuating astrocyte swelling and that estradiol carries this out in part by inhibiting NKCC- and NHE-modulated swelling and reducing the abundance of AQP4 in cortical astrocytes.