Limb remote ischemic postconditioning protects against focal ischemia in rats

Abstract

Remote ischemic postconditioning (RIP) refers to an ischemia conducted in a distant organ that protects against a prior ischemia in another organ. We tested whether RIP protects against focal ischemia in the rat brain. Stroke was generated by a permanent occlusion of the left distal middle cerebral artery combined with a 30-min occlusion of the bilateral common carotid arteries (CCA) in male rats. After CCA release, RIP was generated by three cycles of 15-min occlusion/15-min release of the left-hind femoral artery. The results showed that rapid RIP performed immediately after CCA release reduced infarction by 67% measured at 2 days after stroke. In addition, delayed RIP initiated as late as 3 h, but not 6 h, still robustly reduced infarction by 43% 2 days after stroke. RIP's protective effect was abolished by injecting the protein synthesis inhibitor, cycloheximide, and the afferent nerve blocker, capsaicin, suggesting that RIP blocks ischemic injury by modulating protein synthesis and nerve activity. Nevertheless, rapid RIP did not reduce infarction size 2 months after stroke while it ameliorated the outcome of the behavioral test. In conclusion, RIP attenuates brain injury after focal ischemia.

Fig. 1

Surgery protocols. Rats were divided into 5 groups. Group 1: control ischemia was induced by 30 min occlusion of the bilateral CCA combined with a permanent occlusion of the left MCA. Group 2: a corresponding isoflurane group was used as a control for group 3, for which rapid limb postconditioning was conducted in the left hind limb by 3 cycles of 15 min occlusion/reperfusion. For group 4 and 5, limb postconditioning was induced 3h and 6h after CCA release, respectively.

Fig. 2

Remote limb postconditioning reduced infarct size measured at 2d after stroke. A: Representative infarcts stained by TTC from each group. Rat brains were cut into 5 slices and stained by TTC. Infarct cortex of each slice was measured and normalized to non-ischemic cortex, and expressed as percentage. B: Bar graphs show the average infarct size of the 5 slices. Control, control ischemia; Isoflu, isoflurane of 90 min after reperfusion. * vs 6h, P=0.014; *** vs control, isoflu, P<0.001; ## vs control, isoflu, P<0.01.

Fig.3

The effects of capsaicin and cycloheximide on rapid remote postconditioning's protection. The infarct size of control group from Fig. 2 was re-portrayed with the rest of the 6 groups receiving drugs or vehicles plus rapid remote postconditioning after focal ischemia. Cap, capsaicin; cyclo, cycloheximide. * vs vehicle for cycloheximide, p=0.019; *** vs vehicle for capsaicin, P<0.001.

Fig. 4

Long-term protective effect of rapid remote postconditioning. A: Rapid remote postconditioning attenuated neurological deficits after stroke. * vs sham P<0.001, # vs control ischemia, P<0.001. B: Rapid remote postconditioning did not block ischemic injury measured at two months after stroke. The bar graph represents the average.