In Vivo Visualization of Vasculature in Adult Zebrafish by Using High-Frequency Ultrafast Ultrasound Imaging

Objective:
Zebrafish has been recently considered an ideal vertebrate for studying developmental biology, genetics, particularly for modeling tumorigenesis, angiogenesis, and regeneration
in vivo
. However, when a zebrafish matures completely, its body loses transparency, thus making conventional optical imaging techniques difficult for imaging internal anatomy and vasculature. Acoustic wave penetration outperforms optical methods, high-frequency (>30 MHz) ultrasound (HFUS) was consequently an alternative imaging modality for adult zebrafish imaging, particularly for echocardiography However, visualizing peripheral vessels in a zebrafish by using conventional HFUS is still difficult.
Methods:
In the present study, high-frequency micro-Doppler imaging (HF
μ
DI) based on ultrafast ultrasound imaging was proposed for zebrafish dorsal vascular mapping
in vivo
. HF
μ
DI uses a 40-MHz ultrasound transducer, which is an ultrafast ultrasound imaging technology with the highest frequency available currently. Blood flow signals were extracted using an eigen-based clutter filter with different settings. Experiments were performed on an 8-month-old wild-type AB-line adult zebrafish.
Results:
Blood vessels, including intersegmental vessels, parachordal vessel, dorsal longitudinal anastomotic vessel, and dorsal aorta, from the dorsal side of the zebrafish were clearly observed in two-dimensional (2-D) and 3-D HFμDI.
Conclusion:
The maximum image depth of HFμDI and the minimal diameter of vessel can be detected were 4 mm and 36 μm, respectively; they were determined without any use of microbubbles. The maximum flow velocity range was approximately 3–4 mm/s on the dorsal vessels of the adult zebrafish.
Significance:
Compared with conventional ultrasound Doppler imaging, HF
μ
DI exhibited superior small vessel imaging.