The importance of hormone therapy in affording protection against the sequelae of global ischemia in postmenopausal women remains controversial. Global ischemia arising during cardiac arrest or cardiac surgery causes highly selective, delayed death of hippocampal CA1 neurons. Exogenous estradiol ameliorates global ischemia-induced neuronal death and cognitive impairment in male and female rodents. However, the molecular mechanisms by which estrogens intervene in global ischemia-induced apoptotic cell death are unclear. Here we show that estradiol acts via the classical estrogen receptors, the IGF-I receptor, and the ERK/MAPK signaling cascade to protect CA1 neurons in ovariectomized female rats and gerbils. We demonstrate that global ischemia promotes early dephosphorylation and inactivation of ERK1 and the transcription factor cAMP-response element binding protein (CREB), subsequent down-regulation of the antiapoptotic protein Bcl-2, a known gene target of estradiol and CREB, and activation of caspase-3. Estradiol treatment increases basal phosphorylation of both ERK1 and ERK2 in hippocampal CA1 and prevents ischemia-induced dephosphorylation and inactivation of ERK1 and CREB, down-regulation of Bcl-2 and activation of the caspase death cascade. Whereas ERK/MAPK signaling is critical to CREB activation and neuronal survival, the impact of estradiol on Bcl-2 levels is ERK independent. These findings support a model whereby estradiol acts via the classical estrogen receptors and IGF-I receptors, which converge on activation of ERK/MAPK signaling and CREB to promote neuronal survival in the face of global ischemia.

Transient global ischemia is a neuronal insult that induces delayed cell death. A hallmark event in the early. These findings reveal a novel mechanism by which ischemic insults disrupt the functional integrity of the outer mitochondrial membrane and implicate ⌬N-BCL-xL and VDAC in the large, Zn 2ϩ -dependent mitochondrial channels observed in post-ischemic hippocampal mitochondria.

Transient global ischemia induces selective, delayed neuronal death in the hippocampal CA1 and delayed cognitive deficits. Estrogen treatment ameliorates hippocampal injury associated with global ischemia. Although much is known about the impact of estrogen on neuronal survival, relatively little is known about its impact on functional outcome assessed behaviorally. We investigated whether long-term estradiol (21-day pellets implanted 14 days prior to ischemia) or acute estradiol (50 microg infused into the lateral ventricles immediately after ischemia) attenuates ischemia-induced cell loss and improves visual and spatial working memory in ovariectomized female rats. Global ischemia significantly impaired visual and spatial memory, assessed by object recognition and object placement tests at 6-9 days. Global ischemia did not affect locomotion, exploration, or anxiety-related behaviors, assessed by an open-field test at 6 days. Long-term estradiol prevented the ischemia-induced deficit in visual working memory, maintaining normal performance in tests with retention intervals of up to 1 h. Long-term estradiol also prevented ischemia-induced deficits in spatial memory tests with short (1 and 7 min), but not longer (15 min), retention intervals. Acute estradiol significantly improved visual memory assessed with short retention intervals, but did not prevent deficits in spatial memory. Acute estradiol significantly increased the number of surviving CA1 neurons, assessed either at 7 days after ischemia or after the completion of behavioral testing 9 days after ischemia. In contrast, chronic estradiol did not reduce CA1 cell death 9 days after ischemia. Thus, long-term estradiol at near physiological levels and acute estradiol administered after ischemic insult improve functional recovery after global ischemia. These findings have important implications for intervention in the neurological sequellae associated with global ischemia.

Neuronal NMDA receptors (NMDARs) colocalize with postsynaptic density protein-95 (PSD-95), a putative NMDAR anchoring protein and core component of the PSD, at excitatory synapses. PKC activation and PSD-95 expression each enhance NMDAR channel opening rate and number of functional channels at the cell surface. Here we show in Xenopus oocytes that PSD-95 and PKC potentiate NMDA gating and trafficking in a nonadditive manner. PSD-95 and PKC each enhance NMDA channel activity, with no change in single-channel conductance, reversal potential or mean open time. PSD-95 and PKC each potentiate NMDA channel opening rate ( k β ) and number of functional channels at the cell surface (N), as indicated by more rapid current decay and enhanced charge transfer in the presence of the open channel blocker MK-801. PSD-95 and PKC each increase NMDAR surface expression, as indicated by immunofluorescence. PKC potentiates NMDA channel function and NMDAR surface expression to the same final absolute values in the absence or presence of PSD-95. Thus, PSD-95 partially occludes PKC potentiation. We further show that Ser-1462, a putative phosphorylation target within the PDZ-binding motif of the NR2A subunit, is required for PSD-95-induced potentiation and partial occlusion of PKC potentiation. Coimmunoprecipitation experiments with cortical neurons in culture indicate that PKC activation promotes assembly of NR2 with NR1, and that the newly assembled NMDARs are not associated with PSD-95. These findings predict that synaptic scaffolding proteins and protein kinases convergently modulate NMDAR gating and trafficking at synaptic sites.

Global ischemia arising during cardiac arrest or cardiac surgery causes highly selective, delayed death of hippocampal CA1 neurons. Exogenous estradiol ameliorates global ischemia-induced neuronal death and cognitive impairment in male and female rodents. However, the molecular mechanisms by which a single acute injection of estradiol administered after the ischemic event intervenes in global ischemia-induced apoptotic cell death are unclear. Here we show that acute estradiol acts via the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling cascade to protect CA1 neurons in ovariectomized female rats. We demonstrate that global ischemia promotes early activation of glycogen synthase kinase-3β (GSK3β) and forkhead transcription factor of the O class (FOXO)3A, known Akt targets that are related to cell survival, and activation of caspase-3. Estradiol prevents ischemia-induced dephosphorylation and activation of GSK3β and FOXO3A, and the caspase death cascade. These findings support a model whereby estradiol acts by activation of PI3K/Akt signaling to promote neuronal survival in the face of global ischemia.