Data analysis

Optical traces were conditioned and analyzed in Neuroplex (RedShirtImaging, LLC). Bleach correction was done by subtracting an exponential fit from the optical trace. Each trace was a product of temporal averaging (n = 4 sweeps), spatial averaging (21 – 37 pixels), low-pass Gaussian filter with 77 Hz cutoff, and high-pass Tau filter (10), unless stated otherwise. Optical signal amplitude was measured in Neuroplex as fractional change in light intensity (ΔF/F in %); typical for calcium and voltage imaging experiments [22]. Resting fluorescence intensity (RFI) was measured at standard illumination – fixed LED intensity, by averaging the resting light intensities of 37 neighboring pixels in cortical layer 2/3. Signal to noise ratio (SNR) was measured as the optical signal amplitude (filtered at 77 Hz) divided by peak-to-peak noise measured at unfiltered baseline preceding a biological signal. Temporal Summation (paired pulse facilitation) was calculated as a ratio of the amplitudes of the third peak divided by the first, in the same optical trace. Distance Attenuation was also calculated as a ratio of corresponding peak amplitudes at two recording locations (ROIs). Quantifications were organized in MS Excel. Results are presented as mean ± standard error of the mean (SEM), unless stated otherwise. Statistical significance was determined with unpaired, two-tailed Student’s t-tests. Results were considered statistically significant if the P ≤ 0.05.