Hypoxia-Ischemia and Therapeutic Hypothermia in the Neonatal Mouse Brain – A Longitudinal Study

Abstract

Therapeutic hypothermia is standard of care for infants with hypoxic ischemic encephalopathy. Murine models of hypoxic-ischemic injury exist; however, a well-established mouse model of therapeutic hypothermia following hypoxic-ischemic injury is lacking. The goal of this study was to develop a full-term-equivalent murine model of therapeutic hypothermia after hypoxia-ischemia and examine magnetic resonance imaging, behavior, and histology in a region and sex specific manner. Hypoxic-ischemic injury was induced at postnatal day 10 in C57BL6 mice using a modified Vannucci model. Mice were randomized to control, hypothermia (31˚C for 4h), or normothermia (36˚C) following hypoxic-ischemic injury and stratified by sex. T2-weighted magnetic resonance imaging was obtained at postnatal day 18 and 30 and regional and total cerebral and cerebellar volumes measured. Behavioral assessments were performed on postnatal day 14, 21, and 28. On postnatal day 18, normothermic mice had smaller cerebral volumes (p < 0.001 vs. controls and p = 0.009 vs. hypothermia), while at postnatal day 30 both injured groups had smaller volumes than controls. When stratified by sex, only normothermia treated male mice had smaller cerebral volumes (p = 0.001 vs. control; p = 0.008 vs. hypothermia) at postnatal day 18, which persisted at postnatal day 30 (p = 0.001 vs. control). Female mice had similar cerebral volumes between groups at both day 18 and 30. Cerebellar volumes of hypothermia treated male mice differed from control at day 18, but not at 30. Four hours of therapeutic hypothermia in this murine hypoxic-ischemic injury model provides sustained neuroprotection in the cerebrum of male mice. Due to variable degree of injury in female mice, response to therapeutic hypothermia is difficult to discern. Deficits in female behavior tests are not fully explained by imaging measures and likely represent injury not detectable by volume measurements alone.

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

“…Others have tested different hypothermia temperature targets and some have found that there was no further neuroprotection (Weinrauch et al, 1992; Colbourne and Corbett, 1995), though some have found additional protective effect at 31 °C versus 34 °C in a neonatal rat model (Yager et al, 1993). Further, hypothermia in a similar neonatal model in mice has been observed to provide neuroprotection (as measured by MRI) and improved behavioral testing in males out to 20 days after ischemic injury, whereas the effect on females was variable and transient (Burnsed et al, 2015). A major difference between our findings and previous studies is the age of the rodents at the time of ischemia, as well as the mechanism of injury that is incurred.…”

“…This may be particularly important in light of recent human studies showing lack of effect of hypothermia after out-of-hospital CA in children (Moler et al, 2015), suggesting a need for a better understanding through basic science. Further, it was also somewhat unexpected that there was a difference in the response to hypothermia in male and female juvenile mice, though questions regarding the depth of hypothermia have recently emerged from human trials (Nielsen et al, 2013) and previous reports have suggested sexually dimorphic responses in some behavior tasks in neonatal hypoxia–ischemia models (Burnsed et al, 2015; Smith et al, 2015). Lee and colleagues found that exposing neonatal male rats to 30 °C or 33 °C for varying lengths of time did not further reduce the loss of residual brain volume or motor behavior tasks one month after a model of neonatal hypoxic injury (Lee et al, 2010).…”

Brain-immune interactions in perinatal hypoxic-ischemic brain injury

“…Compared to 129Sv (HI resistance) and C57BL/6 mice (high mortality), CD1 mice display maximum neonatal HI brain injury with very low mortality rate (Sheldon et al , 1998). Adapting this model to the neonatal mice has also revealed that the duration of hypoxic exposure producing a moderate to severe injury in mice is significantly shorter (20–60 min) (Burnsed et al , 2015; Reinboth et al , 2016; Ten et al , 2004). …”

Sestrin2 induced by hypoxia inducible factor1 alpha protects the blood-brain barrier via inhibiting VEGF after severe hypoxic-ischemic injury in neonatal rats

“…Hypoxic ischemic (HI) encephalopathy remains the major cause of perinatal brain injury, resulting in short and long term disabilities (Burnsed et al, 2015; Chicha et al, 2014). It affects 60% of preterm infants and 1–8 cases per 1000 births (Vannucci, 2000).…”