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AbstractRats show spontaneous preference for exploring novel rather than familiar objects. Thus, exploratory activity can be used to evaluate recognition memory. To date, the spontaneous novelty-preference test of object recognition has been used to study long-term recognition memory formation in adult rats (Mumby et al., 2002 Behav Brain Res 132:215-26), but not in aged rats. In the present study we used this behavioral test with 25-27 month-old Wistar rats (n=9), but we found that it was not able to elicit recognition memory. Thus, we developed a new experimental protocol and we tested a second group of rats of the same age (n=18). Each animal received 5 training sessions (1 per day on days 1 and 2, and 1 on day 3) lasting 5 min in a small box (48x26,5x21 cm) containing two identical plastic cubes (8 cm high). We found that the time spent in exploring this pair of objects significantly (P&lt;0.01) decreased on the third session. Twenty-four h after the training 8 rats were tested in the same box, in which one of the two cubes (familiar object) was replaced by a plastic pyramid (8 cm high, novel object). The animals displayed a significantly (P&lt;0.05) longer exploration time of the novel object in comparison with the familiar one. When we applied the new protocol to adult animals (4-6 month-old, n=19), we found that the time spent in exploring the pair of familiar objects significantly (P&lt;0.01) decreased on the second training session, and the time spent in exploring the novel object in comparison with the familiar one was significantly (P&lt;0.05, n=10) longer after 24 h. These findings indicate that the new protocol is able to induce long-term recognition memory formation in aged animals, and allows the evaluation of age-related differences in learning ability.