Hypothermia revisited: Impact of ischaemic duration and between experiment variability

Abstract

To assess the true effect of novel therapies for ischaemic stroke, a positive control that can validate the experimental model and design is vital. Hypothermia may be a good candidate for such a positive control, given the convincing body of evidence from animal models of ischaemic stroke. Taking conditions under which substantial efficacy had been seen in a meta-analysis of hypothermia for focal ischaemia in animal models, we undertook three randomised and blinded studies examining the effect of hypothermia induced immediately following the onset of middle cerebral artery occlusion on infarct volume in rats (n = 15, 23, 264). Hypothermia to a depth of 33℃ and maintained for 130 min significantly reduced infarct volume compared to normothermia treatment (by 27-63%) and depended on ischaemic duration (F(3,244) = 21.242, p < 0.05). However, the protective effect varied across experiments with differences in both the size of the infarct observed in normothermic controls and the time to reach target temperature. Our results highlight the need for sample size and power calculations to take into account variations between individual experiments requiring induction of focal ischaemia.

Keywords: Ischaemic stroke; animal models; hypothermia; ischaemic duration; positive control.

Figure 1.

Effect of hypothermia treatment on infarct size after tMCAo in the SHR. (a) Experiment 1. Hypothermia reduced ischaemic damage by 63%. (b) Experiment 2. Hypothermia had a reduced protective effect when performed by surgery team 2. (c) Experiment 3. Hypothermia reduced ischaemic damage compared to normothermic controls for 60, 75, 90 and 120 min tMCAo. All data presented as mean ± standard deviation. Individual animals are represented by each data point. Each shape/colour represents a different surgeon. #p < 0.001. *p < 0.05.

Figure 2.

Behavioural deficit assessed using a five-point neuroscore at 24 h after stroke. Box plots of total score at 24 h (a, d, g – Experiments 1, 2, 3). Proportion of animals with each score broken into 0.5 unit increments (b, e, h – Experiments 1, 2, 3). No animal scored greater than 3. Proportion of animals with deficits in forelimb flexion (c, f, i – Experiments 1, 2, 3). #p < 0.001. *p < 0.05.