Doppler velocity measurements from large and small arteries of mice

Abstract

With the growth of genetic engineering, mice have become increasingly common as models of human diseases, and this has stimulated the development of techniques to assess the murine cardiovascular system. Our group has developed nonimaging and dedicated Doppler techniques for measuring blood velocity in the large and small peripheral arteries of anesthetized mice. We translated technology originally designed for human vessels for use in smaller mouse vessels at higher heart rates by using higher ultrasonic frequencies, smaller transducers, and higher-speed signal processing. With these methods one can measure cardiac filling and ejection velocities, velocity pulse arrival times for determining pulse wave velocity, peripheral blood velocity and vessel wall motion waveforms, jet velocities for the calculation of the pressure drop across stenoses, and left main coronary velocity for the estimation of coronary flow reserve. These noninvasive methods are convenient and easy to apply, but care must be taken in interpreting measurements due to Doppler sample volume size and angle of incidence. Doppler methods have been used to characterize and evaluate numerous cardiovascular phenotypes in mice and have been particularly useful in evaluating the cardiac and vascular remodeling that occur following transverse aortic constriction. Although duplex ultrasonic echo-Doppler instruments are being applied to mice, dedicated Doppler systems are more suitable for some applications. The magnitudes and waveforms of blood velocities from both cardiac and peripheral sites are similar in mice and humans, such that much of what is learned using Doppler technology in mice may be translated back to humans.

Fig. 1.

Drawing and photos of a 20-MHz Doppler probe designed to measure blood flow velocity noninvasively in mice. The probe consists of a ∼1-mm square piezoelectric crystal, air-backed using Styrofoam, and mounted into a 2-mm-diameter stainless-steel tube with epoxy. A concave epoxy lens is molded to the front face to focus the sound beam at a depth of ∼4 mm. A 10-MHz probe uses a 3-mm tube and is focused at ∼6 mm.

Fig. 2.

Illustration of the setup used to make noninvasive ECG and Doppler measurements showing an anesthetized mouse taped to electrodes on a temperature-controlled circuit board, Doppler and ECG signal processors, and a display of ECG and Doppler signals from the left ventricular inflow and outflow tracts. The 23 × 30-cm-printed circuit board contains 4 stainless-steel ECG electrodes, an array of 50 surface-mount resistors, a temperature sensor, a large ground plane, electrical connections to an ECG amplifier, and a temperature controller. Vel, velocity; Δf, change in frequency. Labeled on the Doppler tracing are the opening (o) and closing (c) of the mitral (m) and aortic (a) valves, peak ejection velocity (P) and acceleration (Accel), and peak-early (E) and late-filling (A) velocities. From these signals, one can obtain accurate timing of cardiac events such as preejection time, filling and ejection times, and isovolumic contraction and relaxation times as indexes of systolic and diastolic ventricular function [from Hartley et al. (25)].

Fig. 3.

Doppler velocity signals from several peripheral arterial sites in an anesthetized mouse using a 2-mm-diameter 20-MHz probe. All signals were obtained with the mouse supine except for the renal signals, which were obtained with the mouse prone and the probe placed lateral to the spine. The coronary signals were obtained later from a second mouse. The vertical lines correspond to the R wave of the ECG and are used to measure velocity pulse arrival times for calculation of pulse wave velocity (PWV) [from Hartley, et al. (25)].

Fig. 4.

Velocity signals from the high-velocity jet in a mouse with a transverse aortic constriction (TAC) along with pressure signals from the right (RCPr) and left (LCPr) carotid arteries. The mouse shown had an arrhythmia such that the jet velocity (V) and the pressure drop (ΔP) varied with each cardiac cycle. The graph to the right shows data from the 5 pulses shown (including the very small one near the center) plus data from 4 other mice where the velocity was varied over a large range. SV, sample volume. As shown, the maximum jet velocity (in m/s) measured noninvasively can be used with the simplified Bernoulli equation (4V²) to estimate the maximum pressure drop (in mmHg) across the aortic constriction [from Li et al. (35)].

Fig. 5.

Baseline and hyperemic left main coronary velocity signals taken noninvasively from a mouse before, 1 day, and 21 days after TAC. The relative contributions of systolic (S) and diastolic (D) flow vary between baseline and hyperemia and with time after constriction [from Hartley et al. (23)].

Fig. 6.

Hyperemic/baseline (H/B) coronary velocity and systolic/diastolic (S/D) area ratios at 5 time points before and after TAC. H/B is an index of coronary flow reserve and is progressively reduced and nearly abolished (1.1) after 21 days of aortic constriction. Similarly, the amount of coronary flow that occurs during systole increases progressively and significantly after aortic constriction.

Fig. 7.

Velocity (in cm/s) and diameter change (in μm) signals taken noninvasively from the right and left carotid arteries of a mouse before and 1 h after TAC. After TAC, the pulsations in both velocity and diameter are increased in the right carotid artery proximal to the band and decreased in the left carotid artery distal to the band. The photo taken at death 2 wk later shows dilation of the right carotid artery because of increased pressure and/or wall shear stress. V_P, peak velocity.

Fig. 8.

Doppler velocity and diameter change signals from the carotid artery of a mouse showing some of the sophisticated analyses that can be done. Wave intensity (WI) is calculated from the product of the derivatives of pressure and flow (or diameter and velocity) and indicates the net magnitude and direction of energy transfer along the vessel. Forward and backward-traveling waves are calculated from the measured pressure and flow (or diameter and velocity) waves and can be used to estimate the magnitude and timing of wave reflections.

Fig. 9.

Velocity, diameter change, and forward and backward waves from the right carotid artery of a mouse before and after TAC. Note that the scales are different before and after constriction and that the second peak seen in the forward wave is missing after constriction.