A new method for preparing a rat intracerebral hemorrhage model by combining focused ultrasound and microbubbles

Abstract

Background: We aimed to prepare a non-invasive, reproducible, and controllable rat model of intracerebral hemorrhage with focused ultrasound (FUS).

Methods: A rat intracerebral hemorrhage (ICH) model was established by combining FUS and microbubbles (μBs), and edaravone was used to verify whether the free radical scavenger had a protective effect on the model. The brain tissue of each group was sectioned to observe the gross histology, blood-brain barrier (BBB) permeability, cerebral infarction volume, and histopathological changes.

Results: Compared with the FUS group, the BBB permeability was significantly increased in the FUS + μBs (F&B) group (p = 0.0021). The second coronal slice in the F&B group had an obvious hemorrhage lesion, and the FUS + μBs + edaravone (F&B&E) group had smaller hemorrhage areas; however, ICH did not occur in the FUS group. The cerebral infarction volume in the F&B group was significantly larger than that in the FUS group (p = 0.0030) and F&B&E group (p = 0.0208). HE staining results showed that nerve fibrinolysis, neuronal necrosis, microglia production, and erythrocytes were found in both the F&B group and the F&B&E group, but the areas of the nerve fibrinolysis and neuronal necrosis in the F&B group were larger than the F&B&E group.

Conclusions: A rat ICH model was successfully prepared using the μBs assisted FUS treatment, and edaravone had a therapeutic effect on this model. This model can be used to study the pathophysiological mechanism of ICH-related diseases and in preclinical research on related new drugs.

Keywords: ICH model; focused ultrasound (FUS); microbubbles (μBs); rat.

FIGURE 1

(A) The connections of the device. (B) A plane diagram of the target site for ultrasound intervention. (C) A schematic image representing the target coordinates of FUS. (D) Schematic diagram of the intervention procedure. (E) Experimental timeline.

FIGURE 2

(A) Representative images of brain slices detected by the IVIS® Lumina III system in three groups. The relative fluorescence intensity is color graded as low (blue), medium (green), and high (red). (B) Quantification the fluorescence intensity of brain slices in three groups. (C) Representative images of brain slices before and after TTC staining in three groups. The red pixel in the red box is the cerebral hemorrhage lesion. (D) Cerebral infarction volume in two groups.

FIGURE 3

(A) Representative images of brain slices before and after TTC staining in two groups. The red pixel in the red box is the cerebral hemorrhage lesion. (B) Cerebral infarction volume in two groups. (C) Representative pictures of HE staining in two groups. Nerve fiber dissolution (green arrow), neuron necrosis (purple arrow), microglia (blue arrow), erythrocyte (red arrow). The corresponding HE stain results at different magnifications: 8× (bar = 1000 μm), 40× (bar = 200 μm), 200× (bar = 50 μm).