(A), Running path of a mouse (left) and calcium traces of example cells (right) in a representative single trial. Mice ran toward two reward locations: one previously learned (R1) and one new (R2). Calcium traces for two cells that reactivated at the reward location and two that did not. Arrows mark reactivations. Vertical line marks time of arrival at R2. (B), Percentage of cells that reactivated at one or both of the reward locations or other locations. Points mark percentage in single trials. Box-and-whisker plots: median, 25 and 75th percentile, and extreme values. n=444 trials. (C), Traces centered on activity peaks for two example cells: one that reactivated at the new reward location in trial 1 (top) and one that did not reactivate in any of the trials (bottom). (D), Histogram of activity peaks normalized to maximum AUC value compared between cells that reactivated for the first time in the preceding trial and cells that did not reactivate in any of the trials. Triangles mark mean values. Permutation tests for repeated measures ANOVA, model for reactivations at the reward locations: significant effect of trial (F(6, 96)=4.51, p<0.001) and reactivation (F(1,16) = 10.15, p=0.006), model for reactivations at non-rewarded locations: significant effect of trial (F(6, 96)=7.85, p=0.001) and non-significant of reactivation (F(1, 16)=0.56, p=0.47). n=6785 cell trials from 1405 cells. (E), The duration from activity during locomotion to the time of first reactivation did not correlate with activity peaks in the following trial, suggesting a long-lasting eligibility trace.(F), Mean calcium peak in each trial for cells that reactivated in trial 1 or trial 4 compared to cells that did not reactivate in any trial. Ribbons extend +/- 1 SEM. Cells that reactivated in trial 1 had significantly higher normalized calcium peaks in all trials. Cells that reactivated for the first time in trial 4 had significantly higher normalized calcium peaks in trials 4, 5, 7 and 8 but not in trial 6 and trials before the reactivation. Permutation t-tests with Benjamini-Hochberg correction for multiple comparisons and between-animal random effects. n=6721 cell trials from 681 reactivated in trial 1 cells, n=769 from 96 reactivated in trial 4 cells, and n=5980 from non-reactivated 607 cells. **p<0.01.