Study On Cerebral Ischemic Tolerance Induced By Various Preconditioning And Their Combinations In Rats

Stroke is the 3rd leading cause of death and continues to be the leading cause of long-term disability, with very few options for effective treatment paradigms. The thrombolytic compound tissue plasminogen activator (t-PA) and edaravone were recognized as the effective therapy for stroke in humans. However, both of them are known to have a relatively short therapeutic window and are not suitable for many stroke patients. Are there any measures that can prevent brain from ischemic injuries in patients undergoing neurosurgical procedures intra-and pre-operatively? The concept of preconditioning and ischemic tolerance might enable us to find practical measures to prevent brain damage. In our laboratory, we found that non-ischemic preconditioning measures such as hyperbaric oxygen, isoflurane and electroacupuncture mimic ischemic preconditioning to induce neuroprotection against ischemia both in the brain and spinal cord. In neurosurgical procedures, temporary vessel occlusion might produce focal cerebral ischemia. If neurosurgical patients with possible temporary vessel clipping are preconditioned with hyperbaric oxygen, isoflurane and acupuncture, cerebral ischemic damage might be prevented. However, which preconditioning method is the most effective and how much should be administered for how long? Isoflurane and sevoflurane, which is the best choice? Do these different preconditioning methods could use in clinical safty? There were more substantial information about non-ischemic preconditionings must be collected before these can be advocated. Thus, the present study focused on the cerebral ischemia tolerance induced by different preconditioning and their combinations.Experiment 1 Rapid brain ischemic tolerance induced by isoflurane preconditioning compared with sevoflurane preconditioning in ratsAim: To investigate whether preconditioning with isoflurane and sevoflurane could induce rapid tolerance to focal cerebral ischemia in rats, and to compare their neuroprotective effects against cerebral ischemic injury. Methods: Forty-two male SD rats weighing 280-320g were randomly assigned into 3 groups (n=14 each group). Animals in Control, ISO and SEVO groups received preconditioning with 97%O2, 1 minimum alveolar concentration (MAC, 1.5%) isoflurane in 97%O2 and 1 MAC(2.4%) sevoflurane in 97%O2 for 1 h respectively. Four rats of each group were randomly assigned to monitor mean artery pressure (MAP) and artery blood gas analysis at the onset of and at the end of preconditioning. At 1h after preconditioning, the other rats were subjected to the right middle cerebral artery occlusion(MCAO)for 2 h, followed by reperfusion for 72 h. The neurological behavior scores (NBS) were assessed at 24 h, 48 h, and 72 h after reperfusion. The percentage of infarct volume was determined at 72 h after reperfusion. Results: The NBS of ISO group and SEVO group at 24 h, 48 h and 72 h after reperfusion were significantly higher than that of Control group (P<0.05). The percentage of infarct volume in ISO group and SEVO group were (31.88±3.66)% and (38.47±2.31)%, which were significantly smaller than in Control group [(48.55±2.18)%, P =0.001 and P=0.032 respectively]. Both NBS and infarct volume were of no significant difference between ISO group and SEVO group. Conclusion: Preconditioning with 1MAC isoflurane or 1MAC sevoflurane could induce rapid brain ischemic tolerance in rats subjected to focal cerebral ischemia, but the neuroprotective effects were similar between these two inhalational anesthetics.Experiment 2 Combination of isoflurane and remote ischemic preconditioning reduces cerebral injury in rats subjected to focal cerebral ischemiaAim: To investigate whether the combination of isoflurane and remote ischemic preconditioning could reduce cerebral injury in rats subjected to focal cerebral ischemia. Methods: 70 rats were randomly assigned to 7 groups: A, control animals only underwent focal cerebral ischemia for 2h followed by reperfusion for 72 h; B and C, rats underwent right femoral artery occlusion/reperfusion for 1 cycle and 3 cycles of 15 minutes respectively just before 2hMCAO; D and E, rats were exposed to 0.75% and 1.5% isoflurane for 1 hour respectively 24 hour before MCAO; F and G were to study the protective effect of the combination of group B plus D, C plus E. The neurological behavior scores (NBS) were assessed at 24 h, 48 h, and 72 h after reperfusion. The percentage of infarct volume was determined at 72 h after reperfusion. Results: No difference was found in physiologic variables values. Animals in A, B, D showed severe neurological damage at 24h, 48h, 72h after reperfusion and C, E, F, G got higher NDS compare with A, B, D. The infarct volume in C, E, F, G [35.61%±5%, 37.04%±3%, 34.54%±4%, 36.97%±3%, respectively]were also smaller than A, B, D [48.56%±2%, 49.83%±5%,40.49%±3%, respectively], but F and G had no statistical significance. Conclusion: Preconditioning with 0.75% of ISO or 1 cycle RIPC had no neuroprotective effects when use alone. However, preconditioning with 0.75% of ISO plus 1 cycle RIPC could reduce cerebral injury in rats subjected to focal cerebral ischemia with improved neurological outcome and reduced brain infarct volume. Those results indicated that preconditioning with ISO in combination with RIPC had a synergistic effect on focal cerebral ischemia in rats.Experiment 3 Comparion of the neuroprotective effects induced by different preconditiong combination with RIPC in ratsAim: To investigate whether the combination of HBO plus RIPC, and EA plus RIPC could reduce cerebral injury as ISO plus RIPC in rats subjected to focal cerebral ischemia. And to find which combination could exerts stronger neuroprotective effects. Methods: 70 rats were randomly assigned to 7 groups: A, Control, animals only underwent focal cerebral ischemia for 2h followed by reperfusion for 72h;B,3 cycles 15min RIPC, rats underwent right femoral artery occlusion/reperfusion 3 cycles of 15 minutes just before MCAO;C, ISO plus RIPC, 24h before MCAO rats were exposed to 0.75% isoflurane for 1h, 1 cycle 15 min RIPC was given just before MCAO;D, HBO plus RIPC, 24h before MCAO rats were preconditioned with HBO(100%O2, 2.5ATA, 1h), 1 cycle 15 min RIPC was given just before MCAO; E, rats were preconditioned with HBO(100%O2, 2.5ATA, 1h) 24h before MCAO; F,EA plus RIPC, EA at the Baihui acupoint for 30min 24h before MCAO, 1 cycle 15 min RIPC was given just before MCAO;G, EA at the Baihui acupoint for 30min 24h before MCAO. The neurological behavior scores (NBS) were assessed at 24 h, 48 h, and 72 h after reperfusion. The percentage of infarct volume was determined at 72 h after reperfusion. Result:Animals in groups B,C,D,F,G showed better neurological function compared with control group, the infarct volume in groups B,C,D,F,G [22.13%±3%, 22.75%±4%, 34.42%±3%, 33.21%±3%, 35.60%±4%, respectively] were also smaller than control group [50.72%±2%]. Group B showed strongest neuroprotective effects. The neuroprotective effects between B and C were no significant difference, but group D showed smaller protective effects compare with group B. Conclusion: Preconditioning with ISO in combination with RIPC or HBO in combination with RIPC could induce cerebral ischemia tolerance in rats by reducing brain infarct volume and improving neurological function. EA preconditioning, which could induced cerebral ischemia tolerance itself, didn't show stronger neuroprotective effects when used together with RIPC.Conclusion1. Preconditioning with 1MAC ISO or 1MAC SEVO could induce rapid brain ischemic tolerance in rats subjected to focal cerebral ischemia, and the neuroprotective effects were similar between these two inhalational anesthetics.2. Preconditioning with 0.75% of ISO, single HBO and 1 cycle RIPC couldn't induce brain ischemic tolerance in rats, but 0.75% of ISO plus 1 cycle RIPC or HBO plus 1 cycle RIPC could reduce cerebral injury in rats respectively indicating that preconditioning with ISO or HBO in combination with RIPC had a synergistic effect on focal cerebral ischemia in rats.3. 1.5% ISO plus 3 cycles RIPC didn't show stronger neuron protective effects than 0.75% ISO plus 1 cycle RIPC,indicating that the synergistic effect has ceiling effect.4. ISO plus RIPC showed better neuron protective effects in all the combination groups of experiment 3.