New microscope captures 3D blood flow and oxygenation at single-cell resolution

The brain relies on real-time delivery of oxygen and nutrients through its microvasculature, which threads through neural tissue like electrical wires. While modern imaging technologies allow researchers to follow the activity of individual neurons in the brain, they are not yet advanced enough to dissect the microvascular function at a comparable spatial scale. This gap hinders our understanding of cerebral small vessel disease and its contributions to cognitive impairment and dementia.

To address this challenge, a team of researchers at Washington University in St. Louis and Northwestern University, led by Song Hu, professor of biomedical engineering in the McKelvey School of Engineering, has developed super-resolution functional photoacoustic microscopy (SR-fPAM). By tracking the movement and oxygenation-dependent color change of red blood cells, SR-fPAM allows researchers to image blood flow and oxygenation at single-cell resolution in the mouse brain, which bridges a critical gap in functional microvascular imaging and could provide new insight into microvascular health and disease, such as stroke, vascular dementia and Alzheimer's disease.

Red blood cells, which are abundant in blood vessels, naturally absorb light due to hemoglobin, the molecule responsible for oxygen transport. When illuminated with short laser pulses, hemoglobin generates ultrasound waves, a phenomenon known as the photoacoustic effect. While conventional photoacoustic microscopy can image blood vessels without labeling them, it does not provide single-cell resolution in 3D.

Hu's team addressed this limitation by developing a high-speed photoacoustic microscope capable of repeatedly imaging the same brain region at millisecond intervals, allowing them to trace red blood cell movement in single files through capillaries and in groups through larger vessels. By tracking these cells across sequential frames and accumulating their trajectories computationally, the researchers were able to reconstruct 3D microvascular structures at single-cell resolution.