Day 49 (direct contact) or 56 (indirect contact) neurons were incubated for 10 minutes at 37°C, 5% CO2 in Fluo-5-AM ester 1 μM in neurobasal medium without growth factors and imaged under constant perfusion at a 0.4ml/minute rate with Tyrode’s solution (concentration in mM: 119 NaCl. 2.5 KCl, 2 CaCl2, 2 MgCl2, 25 HEPES, 30 Glucose) in an AxioObserver.Z1 Zeiss epifluorescence microscope using a 40X-oil objective (Numerical aperture, 1.30), 488nm excitation laser, 488±10 BP excitation filter, 525±50 emission filter and 500FT beam splitter. Images were acquired at 2Hz rate with a Hamamatsu EM-CCD camera and Axiovision 4.8 software. For drug addition, perfusion was stopped and half the volume of Tyrode’s was replaced with Tyrode’s containing the corresponding drug twice concentrated as the final working dilution. Lastly, drugs were washed away for 20 minutes at the aforementioned perfusion rate. For electrical field stimulation, the stimulation paradigm was set using a Master-8 (A.M.P.I.). 30mA pulses were generated with a stimulus isolator (World Precision Instruments) connected to two platinum electrodes placed above the region being imaged. For analysis, regions of interest (ROIs) were
