Exceeding two probe trials in the water maze test may lead to behavioral extinction in mice

Water maze tests are widely used in studying memory, dementia, and related drug efficacy. However, there have been no consensus on the number or duration of probe trials-different labs just use their own protocols, which makes it difficult to compare findings across studies. To address this, we conducted two types of water maze tests (visual-cued and acoustic-cued) in mice to determine the appropriate number of probe trials. In each test three probe trials were performed, with each trial lasting for 60 s. The results of both test types showed significant regression of assessment parameters from the second to the third probe trial, including an increased first crossing time and decreased target crossings, target quadrant time, and target quadrant path length. In contrast, no significant differences were observed between the first and second trials for any parameter. To reinforce the above findings, we partly repeated the experiments. The partial replication of the experiments yielded essentially the same results. Therefore, we believe that the number of 60-second probe trials of water maze tests in mice should not exceed two since more than two probe trials may lead to behavioral extinction.