Focused ultrasound (FUS) with the presence of microbubbles induces blood brain barrier (BBB) opening in targeted areas and facilitates drug delivery. However, recent studies have indicated that FUS-BBB opening with excessive exposure levels may be associated with inflammatory response and cellular/tissue damage. Multiple weekly FUS exposures have been shown to be safe for human subjects. However the effect of more frequent FUS exposures is still unknown. This study examines whether frequent focused ultrasound blood brain barrier opening is associated with aggravated behavioral, histopathologic change or brain tissue damage. Two protocols of focused ultrasound blood brain barrier opening were devised using different microbubble doses (0.15 µl/kg and 0.4 µl/kg). Focused ultrasound exposure at a threshold level of BBB-opening, below-threshold level, or above level for intracerebral hemorrhage were delivered every 2 days. Animal behavioral and physiological changes were examined and recorded. Brain tissue was examined for hemorrhage and apoptosis. Results indicate that frequent exposure of excessive focused ultrasound (1.4 mechanical index) produced minor and short-term behavioral changes despite significant tissue damage, while frequent BBB opening with threshold or below-threshold FUS exposure (0.33–0.8 mechanical index) did not cause behavioral or histological change. Immunofluorescent examination of rat brain tissue indicated that excessive doses of microbubble administration induce an apparent cellular apoptotic response, which may be exacerbated by intracerebral hemorrhage. Experimental results suggest that frequent focused ultrasound blood brain barrier opening with sufficient ultrasound exposure level and a microbubble dose can be safe and pose minimal risk to brain tissue.

Despite extensive efforts in recent years, the blood-brain barrier (BBB) remains a significant obstacle for drug delivery. This study proposes using a clinical extracorporeal shockwave instrument to open the BBB, combined with a laser assisted bi-axial locating platform to achieve non-invasive, controllable-focus and reversible BBB opening in the brains of rats. Under shockwave treatment with an intensity level of 5 (P–9.79 MPa, energy flux density (EFD) 0.21 mJ/mm2) and a pulse repetition frequency of 5 Hz, the BBB could be opened after 50 shocks without the use of an ultrasound contrast agent. With the proposed method, the BBB opening can be precisely controlled in terms of depth, size and location. Moreover, a shockwave based gene transfection was demonstrated using a luciferase gene.