Therapeutic hypothermia protects against ischemia-induced impairment of synaptic plasticity following juvenile cardiac arrest in sex-dependent manner

Abstract

Pediatric cardiac arrest (CA) often leads to poor neurologic outcomes, including deficits in learning and memory. The only approved treatment for CA is therapeutic hypothermia, although its utility in the pediatric population remains unclear. This study analyzed the effect of mild therapeutic hypothermia after CA in juvenile mice on hippocampal neuronal injury and the cellular model of learning and memory, termed long-term potentiation (LTP). Juvenile mice were subjected to cardiac arrest and cardiopulmonary resuscitation (CA/CPR) followed by normothermia (37°C) and hypothermia (30°C, 32°C). Histological injury of hippocampal CA1 neurons was performed 3days after resuscitation using hematoxylin and eosin (H&E) staining. Field excitatory post-synaptic potentials (fEPSPs) were recorded from acute hippocampal slices 7days after CA/CPR to determine LTP. Synaptic function was impaired 7days after CA/CPR. Mice exposed to hypothermia showed equivalent neuroprotection, but exhibited sexually dimorphic protection against ischemia-induced impairment of LTP. Hypothermia (32°C) protects synaptic plasticity more effectively in females, with males requiring a deeper level of hypothermia (30°C) for equivalent protection. In conclusion, male and female juvenile mice exhibit equivalent neuronal injury following CA/CPR and hypothermia protects both males and females. We made the surprising finding that juvenile mice have a sexually dimorphic response to mild therapeutic hypothermia protection of synaptic function, where males may need a deeper level of hypothermia for equivalent synaptic protection.

Keywords: cardiac arrest; global ischemia; hypothermia; pediatric; synaptic function; synaptic plasticity.

Fig. 1

Ischemia impairs synaptic plasticity in juvenile mice. (A) Example fEPSP traces from control mice and mice 7 days after CA where red indicates pre-TBS trace and black indicates post-TBS trace. (B) Time course of fEPSP slope (mean±SEM) from control male mice (black circle), control female mice (white square), male (blue circle) and female (red square) 7 days after CA/CPR. Arrow indicates timing of theta-burst stimulation (40 pulses). (C) Quantification of change in fEPSP slope after 60 min following TBS normalized to baseline, set to 100%. ^*p<0.05 compared with control mice. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 2

Juvenile CA/CPR does not affect probability of release or intrinsic excitability. (A) Quantification of the paired-pulse ratio of the control and 7 days after CA/CPR in each sex. (B) Input–output curve showing normalized fEPSP slope plotted against stimulus intensity (fEPSP slope is normalized to the maximum of each recording). Data are presented as mean±SEM.

Fig. 3

Mild therapeutic hypothermia significantly reduces CA1 neuronal injury in juvenile mice. Representative photomicrographs of hippocampal CA1 neurons from male (A) and female (B) juvenile mice exposed to normothermic cardiac arrest at 37 °C and cardiac arrest followed by 30 min of mild hypothermia (32 °C) in male (C) and female (D) mice. (E) Quantification of ischemic CA1 neurons 3 days after CA/CPR (mean±SEM), indicating a significant reduction in neuronal injury in mice treated with post-arrest hypothermia. ^*p<0.05 compared to normothermia.

Fig. 4

Mild therapeutic hypothermia preserves synaptic plasticity in a sex-specific manner. (A) Time course of fEPSP slope (mean±SEM) from juvenile male mice 7 days following CA/CPR recovered at normothermia (37 °C, red), 32 °C (light blue) and 30 °C (dark blue). Arrow indicates timing of theta-burst stimulation (40 pulses, 100 Hz). (B) Time course of fEPSP slope (mean±SEM) from juvenile female mice 7 days following CA/CPR recovered at normothermia (37 °C, red) and 32 °C (light blue). Arrow indicates timing of theta-burst stimulation (40 pulses, 100 Hz). (C) Quantification of change in fEPSP slope after 60 min following TBS normalized to baseline, set to 100%. ^*p<0.05 compared with control mice. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)