Fig. 4: Hologram-enabled simultaneous biplane multi-target BBB opening in mice.

a Experimental timeline of AH-SiMBO. b Schematic diagram showing the single-element plane-wave transducer coupled with a biplane multi-target hologram for FUS BBB opening. c 3D and 2D images showing the three target regions: bilateral hippocampus and MS, visualized on two separate brain slices. d Axial T1-weighted MR images showing the three BBB opening regions, marked in colored ellipses. e Representative brain photographs showing the EB delivery in treated and control mice. f Representative 30 μm thick brain slices displaying leakage of the EB. g, h MFI given in arbitrary units (a.u.) for EB leakage was quantified using ImageJ for each target region (n = 5 mice in the FUS group; n = 3 mice in the control group). Lateral (i) and axial (j) offset between the three actual target positions (T1, T2, T3) and the corresponding designed target positions. Center line, mean; box limits, upper and lower quartiles; whiskers, 1x standard deviation; points, data points. The PNPs were 0.28, 0.30, and 0.33 MPa for targets 1, 2, and 3, respectively. BBB blood-brain barrier, FUS focused ultrasound, MS medial septum, EB Evans Blue, MFI mean fluorescence intensity. Two-way ANOVA with post-hoc Bonferroni test in (g, h). Data are presented as mean ± SD, ns not significant, ***P < 0.001, ****P < 0.0001.