Transient deficits have been reported after unilateral entorhinal cortex (EC) lesion. To determine whether there is a more persistent deficit, adult male Sprague-Dawley rats with electrolytic or sham lesions of the left entorhinal cortex were examined on acquisition of a modified working memory task in the Morris water maze. This delayed matching-to-sample task, with a 1-h intertrial interval, reveals a significant deficit in total distance to platform in both presentation (Trial 1) and matching (Trial 2) in the rats with entorhinal lesions. We have also found that this test can be used to assess significant deficits in perseveration (repeated nonproductive movement) in rats with entorhinal lesions. The deficits can be seen up to 16 days postinjury. Administration of ganglioside GM1 resulted in a moderate improvement in performance in both water maze measures analyzed. All groups (sham operated, lesion with saline treatment, and lesion with ganglioside GM1 treatment) were given three other tests, which were used to evaluate possible contributing factors to deficient water maze performance. A one-trial test for exploration of novel objects revealed no significant, simple working memory deficit in any group. Plus maze testing, to assess possible differences in levels of anxiety or increased activity as a component of water maze performance, also revealed no differences in the three groups. All groups were also similar in motor activity, shown by monitoring of activity levels. The worsened water maze performance observed in rats with EC lesion may be related to deficits in working memory ability within the framework of acquisition of a more complex spatial learning task.