Hypothermia differentially increases extracellular signal-regulated kinase and stress-activated protein kinase/c-Jun terminal kinase activation in the hippocampus during reperfusion after asphyxial cardiac arrest.

Mitogen-activated protein kinases are signal transduction mediators that have been implicated in cell survival and cell death. This study characterized the activation of pathways in the hippocampus during reperfusion after global cerebral ischemia, as well as the influence of a regimen of hypothermia that reduces ischemic cell death in the hippocampus. Circulatory arrest was induced in rats by 8 min of asphyxia. Relative levels of phosphorylated and total extracellular signal-regulated kinase, stress-activated protein kinase/c-Jun N-terminal kinase and p38 mitogen-activated protein kinase were measured in the hippocampus after 6, 12 or 24h of reperfusion using immunoblotting. Asphyxia induced a progressive increase in phosphorylated extracellular signal-regulated kinase and stress-activated protein kinase/c-Jun N-terminal kinase, but no change in phosphorylated p38 mitogen-activated protein kinase. Induction of mild hypothermia (33 degrees C) during reperfusion increased extracellular signal-regulated kinase phosphorylation and produced a smaller increase in stress-activated protein kinase/c-Jun N-terminal kinase phosphorylation at 24h. Hypothermia did not alter extracellular signal-regulated kinase activation in rats not subjected to ischemia. Extracellular signal-regulated kinase activation was associated with an increase in phosphorylation of the mitogen-activated protein kinase kinase 1/2, and was inhibited by administration of the specific mitogen-activated protein kinase kinase 1/2 inhibitor SL327. Immunohistochemical staining showed an increase in active extracellular signal-regulated kinase in the CA1, CA2, CA3 and dentate gyrus regions of the hippocampus after ischemia and reperfusion. In contrast, active stress-activated protein kinase/c-Jun N-terminal kinase immunoreactivity was most intense in the CA3 and dentate gyrus regions. These data demonstrate that both extracellular signal-regulated kinase and stress-activated protein kinase/c-Jun N-terminal kinase pathways are activated during the first 24h of reperfusion after global cerebral ischemia, and that hypothermia increases the activation of extracellular signal-regulated kinase relative to stress-activated protein kinase/c-Jun N-terminal kinase. Thus, an increase in extracellular signal-regulated kinase activation may be associated with improved neuronal survival after ischemic injury.

Hypothermia during reperfusion after asphyxial cardiac arrest improves functional recovery and selectively alters stress-induced protein expression.

This study examined whether prolonged hypothermia induced 1 hour after resuscitation from asphyxial cardiac arrest would improve neurologic outcome and alter levels of stress-related proteins in rats. Rats were resuscitated from 8 minutes of asphyxia resulting in cardiac arrest. Brain temperature was regulated after resuscitation in three groups: normothermia (36.8 degrees C x 24 hours), immediate hypothermia (33 degrees C x 24 hours, beginning immediately after resuscitation), and delayed hypothermia (33 degrees C x 24 hours, beginning 60 minutes after resuscitation). Mortality and neurobehavioral deficits were improved in immediate and delayed hypothermia rats relative to normothermia rats. Furthermore, both immediate and delayed hypothermia improved neuronal survival in the CA1 region of the hippocampus assessed at 14 days. In normothermia rats, the 70-kDa heat shock protein (Hsp70) and 40-kDa heat shock protein (Hsp40) were increased within 12 hours after resuscitation in the hippocampus. Delayed hypothermia attenuated the increase in Hsp70 levels in the hippocampus but did not affect Hsp70 induction in the cerebellum. Hippocampal expression of Hsp40 was not affected by hypothermia. These data indicate that prolonged hypothermia during later reperfusion improves neurologic outcome after experimental global ischemia and is associated with selective changes in the pattern of stress-induced protein expression.

Geldanamycin provides posttreatment protection against glutamate-induced oxidative toxicity in a mouse hippocampal cell line.

The benzoquinoid ansamycin geldanamycin interferes with many cell signaling pathways and is currently being evaluated as an anticancer agent. The main intracellular target of geldanamycin is the 90-kDa heat shock protein, hsp90. In this report we demonstrate that geldanamycin is effective at preventing glutamate-induced oxidative toxicity in the HT22 mouse hippocampal cell line, even if given 4 h after glutamate treatment. Geldanamycin prevents glutamate-induced internucleosomal DNA cleavage in the HT22 cells but does not reverse the depletion of glutathione levels brought about by glutamate treatment. Both anabolic and catabolic effects are generated by geldanamycin treatment of HT22 cells, as evidenced by the induction of hsp70 expression and degradation of c-Raf-1 protein, respectively. Thus, geldanamycin may provide an effective strategy for manipulating signaling pathways in neuronal cells that use hsp90 as they proceed through a programmed cell death pathway in response to oxidative stress.

Normoxic ventilation during resuscitation and outcome from asphyxial cardiac arrest in rats.

The formation of reactive oxygen species during reperfusion is one trigger for neuronal injury after global cerebral ischemia. Because formation of reactive oxygen species requires delivery of molecular oxygen to ischemic tissue, restricting inspired oxygen during reperfusion may decrease neurological damage. This study examined whether ventilation with room air rather than pure oxygen during resuscitation would improve neurological recovery after cardiac arrest in rats. Adult, male rats were subjected to 8 min of asphyxia resulting in cardiac arrest. During resuscitation, rats were ventilated either with hyperoxia (FiO2 = 1.0) or normoxia (FiO2 = 0.21, room air). Neurobehavioral deficits were scored daily for 72 h after resuscitation, after which brains were collected for histology. Normoxia decreased arterial oxygen content. Other physiological parameters and mortality did not differ between groups. All surviving rats exhibited behavioral and histological signs of brain damage. Neurological deficit scores did not differ between normoxia and hyperoxia conditions at any time point. The number of ischemic neurons in the hippocampus also did not differ between groups. These data indicate neither benefit nor detriment of reducing inspired oxygen concentration during resuscitation from asphyxial cardiac arrest in rats.