Using ultrasonography to define fetal-maternal relationships: moving from humans to mice

Abstract

Ultrasound scanning is a noninvasive, accurate, and cost-effective method to create images of the female reproductive tract clinically and in research. Ultrasonography is particularly valuable for studying the dynamic relationships among mother, placenta, and fetus during pregnancy because this modality does not disturb the ongoing course of gestation. Importantly, the complex vascular changes in the mother induced by pregnancy and the vascular system generated to support placental function can be assessed quantitatively and functionally by ultrasonography. Many mouse models are available that address aspects of human placental function and dysfunction, but high-quality microultrasound technology suitable for use in pregnant mice has become widely available only recently. This technical advance now enables real-time recording of maternal-fetal interactions in pregnant rodents. The ability to perform microultrasonic analyses of parameters such as uterine arterial remodeling, hemodynamic changes, placental development, and fetal growth in mice now permits research that uses the same imaging platform as that for human patients. This capability will enhance the translation of information derived from rodent studies to the clinic.

Figure 1.

Positioning and ultrasound imaging of an anesthetized, dorsally recumbent pregnant mouse (A) as an arial view and (B) in transverse cross-section. Studies can be performed on mice at different gestational stages. All hair was removed from the ventral abdomen after the mouse was anesthetized with approximately 2.0% (1.5% to 2.5%) isoflurane by means of an oxygen mask. The mouse then was placed on the platform and held in position with surgical tape. A thick layer of warm water-based coupling gel was applied over the skin of the area to be imaged. A 40-MHz transducer probe was applied to the skin to collect images. The transabdomenal area (boxed region in A) is suitable for detection of the pregnant uterus. To acquire optimal images, the scanning probe needs to carefully be adjusted to the natural orientation of each implantation site. All waveforms are saved for later offline analysis. Maternal heart and respiration rates were monitored by using an autom ated system. Body temperature was maintained as 36 to 37 °C by the warmed platform, which is supported by an integrated rail system. Figures are not to scale.

Figure 2.

Examples of ultrasound microscope B-mode images from a gd16 BALB/cAnNCrl mouse. (A) Fetal, placental, and maternal decidual areas. The angle of the Doppler beam was less than 30°. Doppler velocity waveforms from the (B) umbilical artery, (C) uterine artery, and (D) transformed spiral artery are shown. (E) Compared with the flow pattern associated with pregnancy (panel C), the uterine artery of virgin BALB/c mice yielded a higher-resistance Doppler waveform. Comparing the time interval between systolic (or diastolic) peaks of the uterine artery and umbilical artery reveals that in mice, the fetal heart beat is much slower than the maternal heart beat. The reverse is seen in human pregnancy. The difference between systolic and diastolic peaks (red lines in panels C and D) was greater in the source uterine artery than in its modified spiral artery tributary. These differences can be evaluated by multiple indices including pulsatility index, resistance index, ratio of systolic to diastolic flow, and so on. Pulse repetition frequency, 10 kHz; wall filter, 100 Hz; display window, 2000 ms; sound speed, 1540 m/s; Doppler gain, 5.00 DB. The red line in image A is a wire frame overlay for pulsed-wave Doppler ultrasonography.

Figure 3.

Volumetric measurement of the same gd14 BALB/cAnNCrl placenta by 2 techniques. After the parameters of the target scan area were set (step size, 0.1 mm; display at 20 to 30 dB), a 3D-mode image was acquired by using a 40-MHz probe. The 3D-mode volume tool was used in cube view (A and C) to measure the object volume of the placenta. For automated calculation of rotational segmentation (A, B), a rotational axis (white bar) was set manually, and a 3D image representative of the volume around the axis was created (B). This image usually excluded the edges of the placental disc and did not account for irregularities in shape. For the manual, parallel serial collection of volume estimate, a manual outline of the placenta was drawn (red), and a series of contours and calculations of volume within each contour was built. The resulting, integrated parallel segmentation 3D image is shown in (D). Typically, rotational segmentation should be used only when the volume has ideal geometric shape. Panels A and C show are irregular contours (*) on the placenta. The yellow central component of each image is the Doppler flow within the umbilical cord; the red edges are regions of slower flow.