(A) Schematics for iClass training. Mice explored in an open field containing a rectangle in the center, which is marked red and 1/10 of the total area, for 5 consecutive days. In the first day (Pre) and last day (Test) mice received no light stimulation. During the 3 consecutive training days (L1, L2, L3), light pulses (483 nm) at 20 Hz were delivered to activate the VTA-mOT DAergic projection when mice were in the rectangle (bottom), and terminated until they left the rectangle (Top). (B–D) ChR2-mCherry-expressing mice (n = 7) increased entry times (B) and retention (C, D) for central area when compared with control mice (n = 7). The dashed line represents the percentage of the marked rectangle to the total. *p<0.05, **p<0.01, ***p<0.001. The preference was turned up at the first training day, increased with the training sections, and maintained to the test day (B, C). Learning curves showed that mice took 3–6 min to overcome their instinctive avoidance for the central area in the first training day (D). The center preference increased with training time and days. (E, F) Representative iClass tracks illustrated light-evoked center preference in ChR2-expressing mice (bottom, (F) but not in the control mice (top, (E).