Refinement of embolic stroke model in rats: Effect of post-embolization anesthesia duration on arterial blood pressure, cerebral edema and mortality

Abstract

Background: Injection of a clot into the internal carotid artery is an experimental model of ischemic stroke that is considered to closely mimic embolic stroke in humans. In this model, the common carotid artery typically remains temporarily occluded to permit time for stabilization of the clot in the middle cerebral artery. However, the associated lengthening of the anesthesia duration could affect arterial blood pressure and stroke outcome.

New method: We refined the model by examining how increasing isoflurane anesthesia duration from 30 to 60 min after clot embolization affects mortality, infarct volume, edema, blood-brain barrier permeability, and the 8-h post-ischemic time course of blood pressure, which has not been reported previously in this model.

Results: We found that arterial pressure increased after discontinuing anesthesia in both embolized groups and that the increase was greater than in the corresponding non-embolized sham-operated rats. At 24 h, the group with 60-min post-ischemia anesthesia exhibited greater brain water content and a greater ipsilateral-to-contralateral ratio of extravasated Evans blue dye. Mortality was greater in the 60-min group, but infarct volume among survivors was not different from that in the 30-min anesthesia group.

Comparison with existing methods: This study refines the embolic stroke model by demonstrating the importance of minimizing the duration of anesthesia after embolization.

Conclusions: These data indicate that early discontinuation of isoflurane anesthesia after clot embolization permits an earlier hypertensive response that limits edema formation and mortality without significantly affecting infarct volume in survivors, thereby decreasing the required number of animals.

Keywords: Animal model of stroke; Embolic stroke; Isoflurane anesthesia; Middle cerebral artery occlusion; Rat.

Figure 1

Box-whisker plots of infarct volume in survivors (n = 54) and non-survivors (n = 16) among Group 1 rats subjected to 30 min of anesthesia after embolization and in survivors (n = 34) and non-survivors (n = 36) among Group 2 rats subjected to 60 min of anesthesia after embolization. *p < 0.05 from corresponding survivor group by the Mann-Whitney rank sum test.

Figure 2

Time course of mean arterial pressure over 480 min after clot embolization (time = 0 min) with continued anesthesia for an additional 30 min (A; n = 11) or 60 min (B; n = 11). Corresponding surgical sham groups received an additional 30 min (A; n = 8) or 60 min (B; n = 8) of anesthesia. Vertical lines indicate cessation of anesthesia for sham and clot embolization groups. An additional control group (n = 6) with a short duration of anesthesia ending at time = 0 min is also shown in A and B. Data are expressed as mean ± 95% confidence interval; *p < 0.05 from control group; †p <0.05 from corresponding anesthesia group after sham surgery; ‡p < 0.05 from 30 min anesthesia group at the same time after embolization.

Figure 3

Evan’s blue dye concentration in cerebral hemispheres contralateral (left) and ipsilateral (right) to sham surgery or to clot embolization followed by 30 (n = 6) or 60 (n = 6) min of anesthesia. The dye was injected at 4 h after surgery/embolization, and tissue was analyzed at 6 h. Because there was no difference in sham groups exposed to 30 or 60 min of additional anesthesia, these groups were combined into a single sham group (n = 12) for statistical purposes. Data are expressed as mean ± 95% confidence interval; *p < 0.05 from sham.

Figure 4

Water content of cerebral hemispheres contralateral (left) and ipsilateral (right) to sham surgery or clot embolization followed by an additional 30 or 60 min of anesthesia (sham surgery, n = 8 per group; embolization, n = 11 per group). Measurements were made at 8 h after surgery/embolization in the same rats that underwent blood pressure monitoring. A control group (n = 6) with a short duration of anesthesia is also shown. Data are expressed as mean ± 95% confidence interval; *p < 0.05 from corresponding anesthesia group after sham surgery; #p < 0.05 from 30 min anesthesia group after clot embolization.