Ischemic Preconditioning and Brain Tolerance: Temporal Histological and Functional Outcomes, Protein Synthesis Requirement, and Interleukin-1 Receptor Antagonist and Early Gene Expression

Abstract

Background and Purpose —A short duration of ischemia (ie, ischemic preconditioning [PC]) can provide significant brain protection to subsequent ischemic events (ie, ischemic tolerance [IT]). The present series of studies was conducted to characterize the temporal pattern of a PC paradigm, to systematically evaluate the importance of protein synthesis in PC-induced IT, and to explore candidate gene expression changes associated with IT. Methods —Temporary middle cerebral artery occlusion (MCAO) (10 minutes) was used for PC. Various periods of reperfusion (ie, 2, 6, and 12 hours and 1, 2, 7, 14, and 21 days) were allowed after PC and before permanent MCAO (PMCAO) (n=7 to 9 per group) to establish IT compared with non-PC (sham-operated) rats (n=22). Infarct size, forelimb and hindlimb motor function, and cortical perfusion (laser-Doppler flowmetry; n=9 per group) were measured after PMCAO. The effects of the protein synthesis inhibitor cycloheximide administered just before PC (n=13 to 17) or administered long after PC but just before PMCAO (n=7 to 8) on IT were also determined. Interleukin-1 receptor antagonist mRNA (reverse transcriptase and polymerase chain reactions [n=20] and Northern analysis [n=50]) and protein expression (immunohistochemistry [n=16]) after PC and early response gene expression (Northern analysis [n=16]) after PMCAO in PC animals were determined. Results —Hemispheric infarct was significantly ( P <0.01) reduced only if PC was performed 1 day (decreased 58.4%), 2 days (decreased 58.1%), or 7 days (decreased 59.4%) before PMCAO. PC significantly ( P <0.01) reduced neurological deficits (similar to reductions in infarct size). Cycloheximide eliminated ischemic PC–induced IT effects on both brain injury and neurological deficits if administered before PC ( P <0.05) but not if administered long after PC but before PMCAO. PC did not produce any significant brain injury, alter cortical blood flow after PMCAO, or produce contralateral cortical neuroprotection. Interleukin-1 receptor antagonist mRNA and protein expression were increased significantly ( P <0.01) only during the IT period. PC rats also exhibited a significant ( P <0.01) reduction in c- fos and zif268 mRNA expression after PMCAO. Conclusions —PC is a powerful inducer of ischemic brain tolerance as reflected by preservation of brain tissue and motor function. PC induces IT that is dependent on de novo protein synthesis. New protein(s) that occurs at the PC brain site 1 to 7 days after PC contributes to the neuroprotection. Those proteins that are produced after the more severe PMCAO in PC animals apparently do not contribute to IT. The PC-induced IT is also associated with increased expression of the neuroprotective protein interleukin-1 receptor antagonist and a reduced postischemic expression of the early response genes c- fos and zif268.