Optimization of behavioral testing in a long-term rat model of hypoxic ischemic brain injury

•In this study we determined the optimal behavioral tests for long-term functional assessment in a rodent model of neonatal brain injury•We demonstrate that Z-scores are a useful tool for standardizing behavioral outcomes and can be used to create a functional composite score.•Rats exposed to a hypoxic ischemic brain injury showed deficits in the negative geotaxis test, cylinder test and novel object recognition test.•This study can inform future studies on behavioral tests most appropriate for long-term assessment in rodent models of neonatal brain injury. Hypoxic ischemic (HI) brain injury is a significant cause of childhood neurological deficits. Preclinical rodent models are often used to study these deficits; however, no preclinical study has determined which behavioral tests are most appropriate for long-term follow up after neonatal HI. HI brain injury was induced in postnatal day (PND) 10 rat pups using the Rice-Vannucci method of unilateral carotid artery ligation. Rats underwent long-term behavioral testing to assess motor and cognitive outcomes between PND11-50. Behavioral scores were transformed into Z-scores and combined to create composite behavioral scores. HI rats showed a significant deficit in three out of eight behavioral tests: negative geotaxis analysis, the cylinder test and the novel object recognition test. These individual test outcomes were transformed into Z-scores and combined to create a composite Z-score. This composite z-score showed that HI rats had a significantly increased behavioral burden over the course of the experiment. In this study we have identified tests that highlight specific cognitive and motor deficits in a rat model of neonatal HI. Due to the high variability in this model of neonatal HI brain injury, significant impairment is not always observed in individual behavioral tests, but by combining outcomes from these individual tests, long-term behavioral burden can be measured.