Dual-modal metabolic analysis reveals hypothermia-reversible uncoupling of oxidative phosphorylation in neonatal brain hypoxia-ischemia.

Hypoxia-ischemia (HI), which disrupts the oxygen supply-demand balance in the brain by impairing blood oxygen supply and the cerebral metabolic rate of oxygen (CMRO(2)), is a leading cause of neonatal brain injury. However, it is unclear how post-HI hypothermia helps to restore the balance, as cooling reduces CMRO(2). Also, how transient HI leads to secondary energy failure (SEF) in neonatal brains remains elusive. Using photoacoustic microscopy, we examined the effects of HI on CMRO(2) in awake 10-day-old mice, supplemented by bioenergetic analysis of purified cortical mitochondria. Our results show that while HI suppresses ipsilateral CMRO(2), it sparks a prolonged CMRO(2)-surge post-HI, associated with increased mitochondrial oxygen consumption, superoxide emission, and reduced mitochondrial membrane potential necessary for ATP synthesis-indicating oxidative phosphorylation (OXPHOS) uncoupling. Post-HI hypothermia prevents the CMRO(2)-surge by constraining oxygen extraction fraction, reduces mitochondrial oxidative stress, and maintains ATP and N-acetylaspartate levels, resulting in attenuated infarction at 24 hr post-HI. Our findings suggest that OXPHOS-uncoupling induced by the post-HI CMRO(2)-surge underlies SEF and blocking the surge is a key mechanism of hypothermia protection. Also, our study highlights the potential of optical CMRO(2) measurements for detecting neonatal HI brain injury and guiding the titration of therapeutic hypothermia at the bedside.