Longitudinal three-photon imaging for tracking amyloid plaques and vascular degeneration in a mouse model of Alzheimer’s disease

Significance: Vascular abnormalities may contribute to amyloid-beta accumulation and neurotoxicity in Alzheimer's disease (AD). Monitoring vascular degeneration as AD progresses is essential. Three-photon fluorescence microscopy enables high-resolution deep tissue imaging with minimal invasiveness and photodamage. Aim: mouse model. Approach: A cranial window allowed repeated 3P imaging at 4-week intervals beginning at 5 weeks after surgery. Vessels labeled with Texas-Red were segmented using DeepVess, whereas plaques labeled with methoxy-XO4 were segmented using custom scripts. Quantitative analyses assessed vascular parameters (diameter, tortuosity, length, inter-vessel distance, total volume) and plaque metrics (radius, total volume). Results: We imaged the same field over 4 weeks, quantifying an overall decrease in vasculature and an increase in amyloid plaques between two sessions. Significant changes in vessel diameter, inter-vessel distance, and alterations in vessel length and plaque radius were observed. Changes in vessel tortuosity were not significant. Conclusions: We demonstrate the potential of three-photon imaging to track vascular and amyloid-related changes in deep cortical structures. It offers a tool for studying the interplay between vascular and amyloid pathologies in AD, supporting future research into disease mechanisms and therapeutic strategies.