Ischaemic pre-conditioning in organotypic hippocampal slice cultures is inversely correlated to the induction of the 72 kDa heat shock protein (HSP72).

In vivo, preconditioning with a sublethal insult can confer resistance to normally lethal episodes of cerebral ischaemia. This phenomenon has been linked with the induction of the 72 kDa heat shock protein (HSP72), but this has not been clearly demonstrated in vitro. We have used organotypic hippocampal slice cultures to investigate whether tolerance to lethal ischaemia is dependent on HSP72. Cultures were maintained in vitro for 14 days, and neuronal damage assessed using propidium iodide fluorescence. Prolonged neuronal HSP72 upregulation occurred following exposure to 30 min ischaemia, 45 min hypoxia and 1 microM kainate, but not 1 microM NMDA or 20 min ischaemia, all sublethal insults. Preconditioning with ischaemia, kainate or hypoxia 24 h prior to lethal ischaemia (45 min) was not protective, and when the delay was increased to 48 h, damage in the CA3 pyramidal cell region was significantly increased compared to cultures exposed to 45 min ischaemia alone. Preconditioning with 20 min ischaemia had no effect on the severity of ischaemic damage. Preconditioning with 1 microM NMDA significantly reduced neuronal damage produced by either 45 or 60 min ischaemia when the delay between insults was 48 h. NMDA pre-treatment also prevented neurotoxicity produced by glutamate (5-10 mM) but not NMDA (10-30 microM). These data suggest that in vitro, the increased expression of HSP72 following some sublethal insults should be considered as a marker of cell stress prejudicial to the survival of neurones subsequently exposed to ischaemia, while tolerance can be produced through mechanisms independent of HSP72 induction.