Allopregnanolone effects on astrogliosis induced by hypoxia in organotypic cultures of striatum, hippocampus, and neocortex

Abstract

Perinatal asphyxia occurs in approximately 0.3% full-term newborn babies, and this percentage has not decreased despite medical advances. There are now evidences indicating that neurosteroids are important in neurodevelopment showing neuroprotective effects. We studied the potential protective effect of allopregnanolone (Allo) in vitro using organotypic cultures from neocortex, striatum, and hippocampus. Immunocytochemistry and confocal microscopy showed an increase of the glial fibrillary acidic protein (GFAP) signal in the studied brain areas after hypoxia. Western blot studies supported these results (hippocampus, 193%; neocortex, 306%; and striatum, 231%). Twenty-four-hour pretreatment with Allo showed different effects at the brain areas studied. In the hippocampus and the neocortex, 24-h pretreatment with Allo 5x10(-6) M showed to be neuroprotective as there was a significant decrease of the GFAP signal compared to control cultures exposed to hypoxia. Pretreatment with 5x10(-8) M Allo attenuated the astrogliosis response in the hippocampus and the neocortex in a nonsignificant way. Allo pretreatment at all doses did not show to affect the astrogliosis triggered by hypoxia in the striatum. Cell survival was analyzed by measuring LDH. After 1 h of hypoxia, all cultures showed a nonsignificant increase of LDH, which was greater after 24 h of hypoxia (hippocampus, 180%; striatum-cortex co-cultures, 140%). LDH levels have no changes by Allo pretreatment before hypoxia.