Figure 3. Outcomes of cognitive tests: polypharmacy treatment impaired non-spatial memory. (A) Number of entries and percentage of spontaneous alternations performed by control and polypharmacy mice in the Y Maze test. (B) NOR test, day 3: the dot plots express the discrimination index (a score above 0 indicates that the mice explored the novel object more than the familiar one). *p<0.001, t-Student test. Histograms on the right show the average of time spent in exploring the two objects by control and polypharmacy animals, ***p<0.001, two-way ANOVA repeated measurements. Note that control mice spent about double the time exploring the new object compared to the familiar one; on the contrary, treated animals did not differentiate between the two objects, as indicated by the discrimination index. (C) The heatmaps visually represent day 3 of NOR test and specifically the area explored around the objects by the animals, showing that only in control group there is a clear preference for the novel object compared to the familiar (in red color the most visited areas). Fam and Nov = familiar and novel object respectively. (D) Contextual and cue FC test: the graph on the left shows the percentage of freezing time measured on day 1 (habituation phase) vs day 2 (context test); the graph on the right expresses the freezing percentage measured before vs during the cue stimulus (sound). *p<0.05, **p<0.01, ***p<0.001, two-way ANOVA repeated measurements test. Ctrl= control, Poly= polypharmacy, n= 10 animals per group. All data are presented as mean ± SEM.