To assess interactions between dendrites of neighboring SACs, we injected pairs of cells with fluorescent dyes. Retinas from mice expressing OFP in SACs (Thy1-OFP3) were mounted RGC side up and perfused with Ames medium bubbled with 95%O2/5%CO2 at 25°C. OFP+ SACs were visualized with epifluorescence, and impaled with high resistance electrodes (50 MΩ) filled with a K+ based intracellular recording solution supplemented with 50 μM Alexa Fluor 568 (for targeting) and 200 μM of Alexa Fluor 488 or 647 (for filling, Invitrogen). Square voltage pulses of ~3V were applied to SACs at 50 Hz using a BK Precision Model 3011B function generator. After filling one SAC, the electrode was replaced with a second containing the contrasting dye and the second cell was filled. Images of labeled SAC pairs in live retinas were acquired at 40X on a Zeiss LSM 510 confocal microscope.
