A custom-made MATLAB script (based on ref. 16) was used to analyze live cell calcium traces and derive various parameters reflecting characteristics of neuronal activity. In brief, regions of interest (ROIs) were drawn based on time projection images of the recordings. For each ROI traces of fluorescence intensity over time were created and used as substrate for subsequent analyses. Fluorescence traces were normalized to the initial fluorescence intensity (F/F0) and average calcium bursting frequency and amplitude were calculated for active cells. Active cells were defined as cells showing at least one peak (i.e. calcium burst) in the fluorescence signal. Fluorescence signals of individual active neurons per well were compared to calculate an average correlation score, indicative of synchronicity of calcium signals (Pearson correlation; -1 up to 1).
