We investigated the earliest time at which irreversible damage takes place after hypoxia-ischemia in the Levine preparation of rats. In 60 rats anesthetized with chloral hydrate and maintained at one of three body temperatures, we unilaterally ligated the left common carotid artery and placed electrodes in the striatum to measure impedance (reflecting the extracellular space) during hypoxia, recovery, and/or cardiac arrest. We measured blood gases and pH at regular intervals during hypoxia in 47 rats and assessed blood-brain barrier function with Evans blue and tissue damage using Na+:K+ ratios. Shortly after hypoxia, impedance normalized in 24 rats without brain damage (normal Na+:K+ ratios, 4 hours of recovery). Sustained elevation of striatal impedance during recovery in six rats was related to an elevated Na+:K+ ratio and a disrupted blood-brain barrier. Damage was not obviously related to blood gases, pH, or the net reduction of the extracellular space during hypoxia. Hypothermia in 17 rats prevented impedance changes, and no striatal damage was found. Thus, irreversible brain damage very likely occurs during or very shortly after hypoxia. Persistent reduction of the extracellular space indicates tissue damage and can be used to monitor potential in vivo therapeutic measures.