Figure 4. HBOT alleviates the reduction in vessel diameter in 5XFAD mice and increases blood flux.In vivo two-photon microscopy imaging and measurements of diameter and velocity in blood vessels of the somatosensory cortex in 5XFAD mice using spatially optimized line scans. (A) Representative images of fluorescently stained vessels of the somatosensory cortex of an HBO-treated 5XFAD mouse (right panel) and a control 5XFAD mouse (left panel) before (upper panel) and after a month of treatment (lower panel). Line scan patterns are superimposed on the vessels. Lines of the scan path along the length were used to calculate RBC velocity (V), while lines across the diameter of the vessels were used to calculate diameter (D). The line scans generated from the path can be stacked sequentially as a function of time to produce a raw cascade image (right of each image). Vessel diameter was calculated as the full width at half-maximum of a time average of several scans across the width of a vessel. RBC velocity was calculated from the angle of the RBC streaks. (B–D) Quantification of vessel diameter (B), RBC velocity in the blood vessels (C) and RBC flow (D), normalized to each treatment group baseline value. Paired t-tests and student t-tests were performed. Values represent means ± SEM.* P < 0.05, *** P < 0.001.