Background: The spinal cord and brain measurements are rarely investigated in neuromyelitis optica (NMO) patients with and without antibodies to aquaporin-4 (AQP4), directly compared to multiple sclerosis (MS) patients. Objectives: To investigate magnetic resonance imaging (MRI) features of both brain and spinal cord in NMO patients with and without antibodies to AQP4, compared with MS patients and healthy controls (HC). Methods: We recruited 55 NMO including 30 AQP4 (+) and 25 AQP4 (-), 25 MS and 25 HC. Brain and spinal cord MRIs were obtained for each participant. Brain lesions (BL), whole brain and deep grey matter volumes (DGMV), white matter diffusion metrics and spinal cord lesions were measured and compared among groups. Results: The incidence of BL was lower in the AQP4 (+) group than in the AQP4 (-) and MS groups (p < 0.05). In the AQP4 (+) group, there was a lower incidence of infratentorial lesions (ITL) and higher spinal cord lesions length than in the MS group (p < 0.05). The thalamic and hippocampal volumes were smaller in the AQP4 (-) group and MS group than in the HC group (p < 0.05). Conclusions: The NMO patients with AQP4 (-) showed higher prevalence of BL, ITL, and similar spinal cord lesion length, compared to AQP4 (+), and demonstrated deep grey matter atrophy, suggesting an intermediate phenotype between that of typical MS and NMO.