Background: Retinal vascular abnormalities may be associated with glaucomatous damage. Adaptive optics (AO) is a new technology that enables the analysis of retinal vasculature at the cellular level in vivo. The purpose of this study was to evaluate retinal arteriolar parameters using the rtx1 adaptive optics fundus camera (AO-FC) in patients with primary open-angle glaucoma (POAG) at different stages and to investigate the relationship between these parameters and changes in spectral-domain optical coherence tomography (SD-OCT) and perimetry. Methods: Parameters of the retinal supratemporal and infratemporal arterioles (wall thickness (WT), lumen diameter (LD), total diameter (TD), wall-to-lumen ratio (WLR), and cross-sectional area of the vascular wall (WCSA)) were analysed with the rtx1 in 111 POAG eyes, which were divided into three groups according to the severity of the disease, and 70 healthy eyes. The associations between RTX1 values and the cup-to-disk ratio, SD-OCT parameters, and visual field parameters were assessed. Results: Compared with the control group, the POAG groups showed significantly smaller TD and LD values (p < 0.05) and significantly higher WLR and WT values (p < 0.05) for the supratemporal and infratemporal arterioles. TD was significantly positively correlated with the retinal nerve fibre layer (RNFL) and ganglion cell complex (GCC) (p < 0.05). LD was significantly positively correlated with the RNFL, GCC, and rim area (p < 0.05). The WLR was significantly negatively correlated with the RNFL, GCC, rim area, and MD (p < 0.05), while it was significantly positively correlated with the cup-to-disc ratio and PSD (p < 0.05). Conclusions: The results suggest that vascular dysfunction is present in POAG, even at a very early stage of glaucoma, and increases with the severity of the disease.