A defining characteristic of optic neuropathies, such as glaucoma, is progressive loss of retinal ganglion cells (RGCs). Current clinical tests only provide weak surrogates of RGC loss, but the possibility of optically visualizing RGCs and quantifying their rate of loss could represent a radical advance in the management of optic neuropathies. In this study we injected two different adeno-associated viral (AAV) vector serotypes in the vitreous to enable green fluorescent protein (GFP) labelling of RGCs in wild-type mice for in vivo and non-invasive imaging. GFP-labelled cells were detected by confocal scanning laser ophthalmoscopy 1-week post-injection and plateaued in density at 4 weeks. Immunohistochemical analysis 5-weeks post-injection revealed labelling specificity to RGCs to be significantly higher with the AAV2-DCX-GFP vector compared to the AAV2-CAG-GFP vector. There were no adverse functional or structural effects of the labelling method as determined with electroretinography and optical coherence tomography, respectively. The RGC-specific positive and negative scotopic threshold responses had similar amplitudes between control and experimental eyes, while inner retinal thickness was also unchanged after injection. As a positive control experiment, optic nerve transection resulted in a progressive loss of labelled RGCs. AAV vectors provide strong and long-lasting GFP labelling of RGCs without detectable adverse effects.