Labelling retinal ganglion cells for imaging

OGB-1-labelled GCL cells were targeted with electrodes (5–15 MΩ) subsequent to two-photon recordings. Single cells in the GCL were dye-filled with SR101 (Invitrogen) using the buzz function (100-ms pulse) of the MultiClamp 700B software (Molecular Devices). Pipettes were carefully retracted as soon as the cell began to fill. Approximately 20 min were allowed for the dye to diffuse throughout the cell before imaging started. After recording, an image stack was acquired to document the cell’s morphology, which was then traced semi-automatically using the Simple Neurite Tracer plugin implemented in Fiji. In cases of any warping of the IPL we used the original image stack to correct the traced cells using custom-written scripts in IGOR Pro (for details, see ref. ^{37 (link)}).