Procedure: Echocardiography

1. Mice are placed in an induction chamber and anesthetized.
2. Once sedated, each mouse is moved to a nose cone for hair removal using hair-removal cream (apply only to the area that will be utilized for imaging); the area is wiped clean with wet gauze.
3. Each mouse is moved to the imaging platform and its paws are taped to the ECG lead plates and a rectal probe is inserted. Body temperature is maintained at 36-37°C.
4. During imaging, anesthesia is reduced to maintain proper heart rate; if the mouse shows signs of wakefulness, more anesthesia is used.
5. Imaging gel is applied to the area to be imaged and the imaging probe is lowered until it makes contact with the gel, making sure that all areas of the probe are covered with gel.
6. The image of the heart is taken in the short-axis mode with papillary muscles being the point of reference. The papillary muscles should be parallel to the screen; some manipulation of the platform may be necessary in order to get a clear image.
7. Once the papillary muscles are parallel in B-Mode, press the M-mode button and place the yellow line in the middle of the left ventricle (LV). Change the display window to 1000 ms. If image is not clear, move the yellow line to the left or right; or leave the line in the center of the LV and move the platform left or right, forward or backwards.
8. Save the M-mode data by pressing the Cine store button when a good clear image is obtained; save at least 4 short loops of data.
9. To view the heart two-dimensionally in the parasternal long-axis choose the Presets: B-mode, parasternal long-axis view.
10. Place the transducer vertically to the animal's body on the left side of its sternum with the notch of the transducer pointing to the animal's head; optimal parasternal views are obtained by adjusting gain settings or visualization of endocardial and epicardial walls; a proper image in this orientation includes the left atrium and ventricle, a slight portion of the right ventricular wall and the output of the aorta, with the heart forming a gourd-like structure; the beginning ascending aorta and the apex of the heart are on the same horizontal line.
11. When a good clear image is obtained press the Cine store button to save data; each mouse should have at least 3 short B-Mode, parasternal long-axis view images captured.
12. Once imaging is complete, remove mouse from platform and allow to recover atop a heating pad.
13. Offline analysis: To define the diameter of the aortic arch, ensure that this measurement is done during systole of the heart. The diameter is to be measured perpendicular to the aortic wall. Use a ruler to measure the diameter; place it on the right of aortic valves and on the left of the beginning right atrium. Each mouse should have a least 3 ascending aorta diameters taken.