This study examined whether the cooperation of the hippocampus and anterior thalamus via the fimbria-fornix is involved in the spatial memory. We compared the effect of contralateral lesions (Contra) with ipsilateral lesions (Ipsi) of the fimbria-fornix and anterior thalamus on the performance of an object exploration task and the Morris water maze task. If the hippocampus and anterior thalamus take part in a same functional system via fornix, the performance of Contra group will be more disruptive than that of Ipsi group. In the object exploration task, Contra and Ipsi groups did not differ from Control group in the performance of object recognition test. However, the performance of Contra group was significantly impaired in the spatial recognition test, compared with two other groups. In the Morris water maze task, only Contra group showed deficits in spatial learning but not the Ipsi group. These results indicate that contralateral, but not ipsilateral lesion caused deficits in spatial memory, supporting the notion that the functional cooperation of hippocampus and anterior thalamus via fornix is vital for spatial memory.