Importance of normothermia control in investigating delayed neuronal injury in a mouse global ischemia model

Abstract

This study aims to establish a mouse global cerebral ischemia model in which the physiological parameter measurements and neuronal injury evaluations are conducted in the same group of animals and to identify the effect of post-ischemic core temperature (CT) on the outcome of neuronal injury. Global ischemia was induced by 12-min bilateral common carotid artery occlusion followed by 7 days of reperfusion in C57BL/6 mice. Immediately after occlusion, mice were randomly assigned to be kept in environments of different temperatures [25 degrees C (room temperature, group 1), 33-34 degrees C for 2h (group 2), and 33-34 degrees C for 24h (group 3)] before being returned to their home cages. We found that in group 1, CT declined to approximately 32 degrees C after ischemia and then recovered at 24h post-ischemia; in group 2, CT remained at the pre-ischemia level during the first 2h, declined after the mice were returned to room temperature, and recovered at 24h post-ischemia; and in group 3, CT remained constant at the pre-ischemia level throughout the reperfusion period. The number of surviving neurons in a sector of the hippocampal CA1 region was significantly lower in all ischemic groups than in the sham controls, but the number was significantly higher in group 1 than that in groups 2 or 3 (P<0.05). We observed that CT declines initially but recovers spontaneously at 24h post-ischemia. Early post-ischemic hypothermia impacts the delayed neuronal injury, suggesting that tight temperature control immediately following ischemia is important to obtain the most reproducible neuronal damage in mouse models of cerebral global ischemia.

Fig. 1

Changes in mean arterial blood pressure (MABP) and relative cerebral blood flow (CBF) in each hemisphere in mice subjected to 12-min global cerebral ischemia followed by reperfusion. MABP increased and CBF decreased during bilateral common carotid artery occlusion.

Fig. 2

Head and core temperature, MABP, and CBF in each hemisphere were recorded in mice subjected to 12-min global cerebral ischemia followed by reperfusion. The respective head (A) and core (B) temperatures were not different among groups during or early after ischemia. (C) The MABP was significantly higher in animals subjected to global cerebral ischemia than in sham-operated animals. (D) Upon initiation of ischemia, CBF decreased to a level significantly lower than that of sham-operated animals and then returned to near baseline after reperfusion. *P < 0.01 compared to sham-operated animals.

Fig. 3

Characterization of core body temperature in mice subjected to 12-min global cerebral ischemia and then exposed to a warm temperature-controlled chamber for different lengths of time. (A) Core temperature decreased to about 32 °C but spontaneously recovered 24 h post-ischemia in mice that were kept at room temperature after ischemia. (B) In mice exposed to a 33–34 °C chamber for 2 h, core temperature remained constant for 2 h, decreased to about 32 °C upon exposure to room-temperature air, and then spontaneously recovered 24 h post-ischemia. (C) Core temperature remained consistently at 37 °C in mice that were kept in a temperature-controlled chamber for 24 h before being returned to room temperature.

Fig. 4

Neuronal viability in the hippocampal CA1 region of mice subjected to 12-min global cerebral ischemia and then exposed to a warm temperature-controlled chamber for different lengths of time. Neuronal survival in the CA1 region was significantly decreased in mice subjected to 12-min global cerebral ischemia followed by 7 days of reperfusion compared to sham-operated mice. However, neuronal survival was significantly higher in animals kept at room temperature (25 °C) than in animals that were kept in a 33–34 °C chamber for 2 or 24 h after ischemia. Results are shown as mean ± SD. **P < 0.01 vs. sham-operated mice, ^#P < 0.05 vs. animals kept at room temperature.