New imaging system maps retinal oxygen in unprecedented detail

The retina consumes oxygen at one of the highest rates of any tissue in the body, and disruptions in its oxygen supply are linked to blinding diseases such as glaucoma, age-related macular degeneration, and diabetic retinopathy. Yet researchers have struggled to noninvasively measure oxygen levels at the fine scale of retinal capillaries, where early disease changes often occur.

Now, a team from Johns Hopkins University and the University of Pennsylvania has developed a multimodal system that combines two advanced optical techniques to image both retinal structure and oxygen metabolism in live mice. The approach, reported in Neurophotonics, merges visible light optical coherence tomography (VIS-OCT), which produces 3D images of tissue microanatomy, with phosphorescence lifetime imaging scanning laser ophthalmoscopy (PLIM-SLO), which provides precise measurements of oxygen partial pressure (pO₂) within blood vessels.

To carry out the oxygen measurements, the researchers injected mice with a specially designed probe (Oxyphor 2P) that alters its phosphorescence lifetime depending on oxygen levels. By scanning the eye with controlled pulses of light, the team could calculate pO₂ values down to the level of individual capillaries. Meanwhile, the VIS-OCT channel can capture high-resolution views of the retinal layers and blood-flow patterns. The two systems were synchronized to acquire data through the same optical path, ensuring that oxygen readings and structural images could be spatially aligned.

Tests in healthy mice showed that the system could track changes in retinal oxygenation in response to different inhaled oxygen levels. As expected, arterioles exhibited higher pO₂ than venules, with capillary values typically falling in between. The measurements also matched systemic blood oxygen saturation, producing oxygen dissociation curves consistent with known hemoglobin physiology. Importantly, the instrument could focus at different retinal depths to elucidate local capillary values.

By simultaneously recording anatomy and oxygen tension, the dual-channel system offers a powerful new tool for studying how retinal oxygen supply is altered in disease and during treatment. The authors note that the approach is non-destructive, allowing repeated measurements in the same animal over time and could be applied to mouse models of retinal disease to investigate how microvascular dysfunction contributes to vision loss. The simultaneous imaging capability is key to refining and validating fully label-free VIS-OCT oximetry by pairing the ground truth PLIM-SLO pO₂ with the registered VIS-OCT channel.

The researchers suggest that with further refinement—such as adaptive optics for sharper images—the platform may pave the way toward improved diagnostics and monitoring strategies in human eye disease.

For details, see the original Gold Open Access article by S. Nolen et al., “Multimodal retinal imaging by visible light optical coherence tomography and phosphorescence lifetime ophthalmoscopy in the mouse eye," Neurophotonics 12(3), 035015 (2025), doi: 10.1117/1.NPh.12.3.035015.