To analyze the retinal ganglion cell layer (RGL) by optical coherence tomography (OCT) in toxic and nutritional optic neuropathy and to correlate its thickness and volume with functional damage. We conducted an observational cross-sectional study in healthy subjects and in patients with toxic optic neuropathy observed in the Neuro-Ophthalmology Department of Central Lisbon Hospital Center. Complete ophthalmologic examination, OCT (Heidelberg Spectralis), and automated static perimetry were performed. Thickness and macular volume of RGL layer and inner plexiform layer were measured after manual segmentation. The study included 16 eyes of 12 healthy subjects and 16 eyes of 8 patients with toxic and nutritional optic neuropathy. Age and gender did not differ between the 2 groups. Ethambutol was the cause of toxic optic neuropathy in 4 patients and nutritional factors (tobacco-alcohol) in 4 patients. A statistically significant decrease in thickness and volume of RGL, in all quadrants at 2 and 3 mm, was detected in individuals with optic neuropathy compared with controls (P < 0.01). A positive correlation between RGL thickness and mean deviation (MD) and between RGL volume and MD was detected (P < 0.05). There was a negative correlation between MD and time of disease (r = 0.846 P = 0.001) and a positive correlation between MD and visual acuity in logMAR (r = 0.739 P = 0.006). A majority of the structural parameters also correlated negatively with time of disease (P < 0.05). Decreased RGL thickness and volume detected in this study support a mechanism of RGL toxicity. RGL analysis may contribute to the diagnosis and management of toxic and nutritional optic neuropathies.