Brain slices (275 µm) were acutely prepared from retroviral injected animals and focally loaded with 5 µl of 1 mM Fluo4-AM (Invitrogen). Ca2+ imaging was performed as previously described (Shim et al., 2009). Different groups of cells were treated with muscimol (10 µM), followed by ionomycin (10 µM). Cells were excited at 488 nm, and Fluo-4 signal was collected at 505–550 nm. Images were acquired every 5 sec and analyzed using NIH Image J software. The Ca2+ signal change was determined by ΔF/F [ΔF/F = [(F1-B1)-(F0-B0)]/(F0-B0)], which was normalized to the mean fluorescence intensity measured at the baseline condition (set as 0%) and after the ionomycin treatment (set as 100%).
