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The rodent hypoxia-ischemia (HI) model, referred to as the Vannucci model, is the most commonly used model for studying perinatal hypoxic-ischemic brain injury (Zhu et al., 2009; Vannucci and Hagberg, 2004). In the Vannucci model, brain injury is acquired by combining a permanent unilateral common carotid artery ligation with subsequent exposure to hypoxia (Rice et al., 1981). The Vannucci model was originally developed in rat pups at postnatal day (PND)7 (Rice et al., 1981), an age at which the development of the rat brain corresponds to that of the human infant at gestational weeks 32-34 (Zhu et al., 2009; Vannucci et al., 2009). The Vannucci HI model has since been adapted to mouse models of perinatal brain injury, and this has allowed the technique to be used with a broader array of genetically modified animals. Mice at PND9 have been the most commonly used, and these correspond to the human near-term infant (Zhu et al., 2009). In the present protocol, the Vannucci model has been adapted to serve as a model of preterm brain injury in C57bl/6J mice at PND5, an age where the development of the mouse brain corresponds to the brain of a human preterm infant (Zhu et al., 2009; Albertsson et al., 2014). The injury acquired with this protocol is characterized by local white matter injury combined with small areas of focal cortical injury and hippocampal atrophy (Albertsson et al., 2014).