Current evidence suggests a central role for autophagy in many inflammatory brain disorders, including Alzheimer's disease (AD). Furthermore, it is also well accepted that some inhalation anesthetics, such as isoflurane, may cause AD-like neuropathogenesis and resultant postoperative cognitive dysfunction, especially in the elderly population. However, the impact of inhalation anesthetics on autophagic components in the brain remains to be documented. Hence, our objective was to investigate the effects of different durations of isoflurane exposure on hippocampus-dependent learning and hippocampal autophagy in aged rats. Aged Sprague-Dawley rats (20 months old) were randomly exposed to 1.5% isoflurane or 100% oxygen for 1 or 4 h. Animals were then trained in the Morris water maze (4 trials/day for 5 consecutive days). Hippocampal phagophore formation markers, beclin 1 and protein microtubule-associated protein 1 light chain-3B (LC3B), as well as p62, an indicator of autophagic flux, were quantified by western blotting. There was no significant difference in the escape latencies and time spent in the target quadrant, as well as hippocampal expression of beclin 1, LC3B-II, and p62 at 24 h post-anesthesia between the 1-h isoflurane-exposed rats and their controls (P >0.05). Four-hour exposure to isoflurane resulted in spatial learning and memory deficits, as evidenced by prolonged escape latencies on days 4 and 5 post-anesthesia and less time spent in the target quadrant than sham-exposed animals (P <0.05). These events were accompanied by a decline in hippocampal expression of LC3B-I, LC3B-II, and beclin 1 24 h after isoflurane (P <0.01 and P <0.05). Nevertheless, no significant change in p62 expression was found. Further kinetics study of autophagic changes induced by 4 h of isoflurane showed a transient upregulation of LC3B-I, LC3B-II, and beclin 1 at the end of exposure and a subsequent striking decrease within 12-24 h post-anesthesia (P <0.05). Hippocampal p62 peaked at 6 h but subsequently resolved. These results from our pilot in vivo study support a duration-dependent relationship between 1.5% isoflurane exposure, and spatial cognitive function as well as hippocampal phagophore formation.