Although the world is three-dimensional, spatial cognition has been extensively studied mostly in twodimensional environments. Several studies showed a difference in the way freely moving species learn spatial information in the two dimensions and integrate it into an internal representation of space as compared to surface-bound species, which prioritize the horizontal dimension and encode it with higher resolution. Cephalopods have remarkable spatial abilities and can move freely in a volume, yet some species, such as cuttlefish, are bottom-dwelling animals, making them an interesting model for the study of three dimensional navigation. We tested the relative preference of vertical vs. horizontal information in trained Sepia officinalis cuttlefish. Animals were trained to approach one of two visual cues arranged in a 45° configuration, either up and right or down and left. After reaching a learning criterion, the animals were sequentially presented 3 different test configurations of the two visual cues, with training trials in between. In the first two tests the animal was presented either with a horizontal or a vertical configuration of the two visual cues. These tests assessed whether the animal could extract the vertical or the horizontal coordinates from the previously learned spatial location of the rewarded visual cue. Most animals made a correct choice in both tests. The last test presented a conflict situation, in which the two visual cues were arranged in a 45° configuration opposite to the training one. Animals significantly chose the visual cue which was consistent with the previously learned vertical information. Hence, cuttlefish can separately learn and extract the vertical and horizontal coordinates of a three-dimensional location, and the vertical component of space seems to have a greater importance than the horizontal component.