Microvascular and astrocytic responses to repeated magnetic resonance-guided focused ultrasound

Focused ultrasound (FUS), in combination with microbubbles, enables the transient and localized opening of the blood-brain barrier (BBB) for targeted drug delivery. While this technique has shown promise in preclinical models, the safety and neurovascular consequences of repeated BBB permeabilization remain incompletely understood. In this study, we performed a longitudinal assessment of vascular, microstructural, and glial responses following repeated FUS-mediated BBB opening in healthy rats using magnetic resonance (MR)-guided targeting and MR vascular fingerprinting. FUS sessions were conducted on a weekly or twice-weekly basis over a period of four weeks, employing two distinct acoustic pressures, 390 kPa and 440 kPa, respectively. The BBB opening was reproducible across sessions, with stable contrast enhancement observed with DCE-MRI and a homogeneous cumulative open BBB volume. No significant hemorrhages or edema were detected; however, transient cerebral blood flow reductions of approximately 15-20 % were observed following each session. After eight FUS sessions at 440 kPa, subtle vascular remodeling was observed, including increased vessel radius and reduced tissue oxygen saturation. This finding was confirmed by both MRI and histological analysis. Astrocytic activation, as determined by GFAP immunostaining, was minimal after a single or weekly FUS exposure but became significant with biweekly treatments, indicating a cumulative neuroimmune response. These results support the feasibility of weekly FUS-induced BBB opening at moderate acoustic pressures. However, they also underscore the importance of protocol optimization to prevent glial and vascular stress in long-term therapeutic applications.