High-resolution fluorescence-guided transcranial ultrasound mapping in the live mouse brain.
Authors: Estrada H, Robin J, Özbek A, Chen Z, Marowsky A, Zhou Q, Beck D, le Roy B, Arand M, Shoham S, Razansky D
Understanding the physiological impact of transcranial ultrasound in rodent brains may offer an important preclinical model for human scale magnetic resonance–guided focused ultrasound methods. However, precision tools for high-resolution transcranial ultrasound targeting and real-time in vivo tracking of its effects at the mouse brain scale are currently lacking. We report a versatile bidirectional hybrid fluorescence-ultrasound (FLUS) system incorporating a 0.35-mm precision spherical-phased array ultrasound emission with a fiberscope-based wide-field fluorescence imaging. We show how the marriage between cortex-wide functional imaging and targeted ultrasound delivery can be used to transcranially map previously undocumented localized fluorescence events caused by reversible thermal processes and perform high-speed large-scale recording of neural activity induced by focused ultrasound. FLUS thus naturally harnesses the extensive toolbox of fluorescent tags and ultrasound’s localized bioeffects toward visualizing and causally perturbing a plethora of normal and pathophysiological processes in the living murine brain.
Introduction
Purpose
Transcranial ultrasound stimulation
Study Objective
Develop and demonstrate a bidirectional hybrid fluorescence-ultrasound (FLUS) system for precise transcranial focused ultrasound targeting and real-time fluorescence-guided monitoring of its effects in the mouse brain.
Animal model / Human subject
Mouse (Mus musculus); strain: C57BL/6J-Tg (Thy1-GCaMP6f); age: 6-9 weeks; sex: male and female
Disease model
Healthy
MRI or image guidance method
Fluorescence-guided (fiberscope-based wide-field fluorescence imaging)
Targeted brain region(s)
Cerebral cortex
Outcomes and Safety
Summary of Outcomes
Transcranial focused ultrasound produced highly localized, reversible fluorescence dips attributable to focal heating (no histological damage at 3 MPa/150 ms), while more intense or longer sonications caused pathological effects: 3.9 MPa/100 ms induced blood–brain barrier disruption and decreased functional connectivity, and stronger/longer stimulations (e.g., 4.8 MPa/150 ms and 300 ms exposures) reliably triggered cortical spreading depression. Parameters found to produce these effects: 3 MPa, 150 ms → thermal fluorescence dips (safe); 3.9 MPa, 100 ms → BBB disruption and network changes; 3–3.9 MPa, 100–300 ms (intermediate) → occasional CSD; 4.8 MPa, 150 ms (and 300 ms long pulses) → consistent CSD.
Safety-related matter
No histological signs of brain damage or changes in functional connectivity were found after pulsed sonications at 3 MPa (sequence 1). However, more intense or longer sonications (e.g., 3.9 MPa or 300 ms) produced adverse effects including cortical spreading depression, negative network-level functional connectivity changes, blood–brain barrier disruption (IgG extravasation), and perivascular microglial/astrocytic gliosis indicative of local inflammation.
Brain Region
Ultrasound Parameters
Ultrasound instrument
FLUS (bidirectional hybrid fluorescence–ultrasound system); 0.35-mm precision spherical phased-array ultrasound emitter; manufacturer: None
FUS Frequency
3 MHz
FUS Intensity
284-724 W/cm2
FUS Pressure
3-4.8 Mpa
FUS Mode
pulsed
Pulse duration
150, 100-300 ms
Duration of a single FUS session
150 ms
Focal Characteristics
focal depth: None, focal length: None, aperture size: None
Treatment frequency
multiple sessions
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