Ultrasound imaging utilizes the interaction of sound waves with living tissue to produce an image of the tissue or, in Doppler-based modes, determine the velocity of a moving tissue, primarily blood. These dynamic, real time images can be analyzed to obtain quantitative structural and functional information from the target organ. This versatile, noninvasive diagnostic tool is widely used and accepted in human and veterinary medicine. Until recently its application as a research tool was limited primarily to larger, nonrodent species. Due to advances in ultrasound imaging technology, commercially available ultrasound systems now have the spatial and temporal resolution to obtain accurate images of rat and mouse hearts, kidneys, and other target tissues including tumor masses. As a result, ultrasound imaging is being used more frequently as a research tool to image rats and mice, and particularly to evaluate cardiac structure and function. The developing technology of ultrasound biomicroscopy has even greater spatial resolution and has been used to evaluate developing mouse embryos and guide site-specific injections into mouse embryos. Additional ultrasound imaging technologies, including contrast-enhanced imaging and intravascular ultrasound transducers adapted for transesophageal use, have been utilized in rats and mice. This paper provides an overview of basic ultrasound principles, equipment, and research applications. The use of noninvasive ultrasound imaging in research represents both a significant refinement as a potential replacement for more invasive techniques and a significant advancement in research techniques to study rats and mice.

Ultrasound imaging: principles and applications in rodent research.

Background—No noninvasive technique is currently capable of “real-time” assessment and monitoring of cerebral blood flow (CBF). We hypothesized that cerebral perfusion could be accurately measured and monitored in “real time” with contrast-enhanced ultrasound (CEU). Methods and Results—Cerebral perfusion was assessed in 9 dogs through a craniotomy with CEU at baseline and during hypercapnia and hypocapnia while normoxia was maintained. Cerebral microvascular blood volume (A), microbubble velocity (), and blood flow (A) were calculated from time-versus–acoustic intensity relations. Compared with baseline, hypercapnia and hypocapnia significantly increased and decreased CBF, respectively, as measured by CEU. These changes in blood flow were mediated by changes in both A and. A good correlation was found between A derived from CEU and CBF measured by radiolabeled microspheres (y0.67x0.04, r0.91, P0.001). Conclusions—Changes in both cerebral microvascular blood volume and red blood cell velocity can be accurately assessed with CEU. Thus, CEU has the potential for bedside measurement and monitoring of cerebral perfusion in real time in patients with craniotomies or burr holes. (Circulation. 2001;104:2582-2587.) n current clinical practice, noninvasive assessment of cerebral blood flow (CBF) can be achieved by use of various techniques, such as CT, MRI, and single photon emission CT (SPECT). These techniques, however, are ill suited for measurement of CBF at the bedside or in the operating room. Furthermore, they cannot be used repeatedly, in rapid succession, to measure changes in CBF that may occur in the natural course of events or in response to treatment. In addition, these accurate, albeit expensive, modalities cannot be used for monitoring regional CBF in patients with acute ischemic stroke or intracranial hemorrhage, or for the serial assessment of global CBF in patients with increased intracranial pressure. We have previously shown that blood flow to various tissues, including the myocardium,1 skeletal muscle,2 and kidney,3 can be measured by contrast-enhanced ultrasound (CEU). This technique, which uses small gas-filled microbubbles, can provide an assessment of microvascular blood volume (MBV) as well as red blood cell velocity (RBCv). These parameters can be obtained during a continuous intravenous infusion of microbubbles. After steady state is achieved, the microbubbles within tissue are destroyed with high-energy ultrasound, and the rate at which they replenish the ultrasound beam is measured, which reflects RBCv. The concentration of microbubbles when the beam is fully replenished is proportional to MBV. We therefore hypothesized that CEU can be used to assess CBF noninvasively. We performed experiments in dogs in which CBF was measured with both CEU and radiolabeled microspheres during rest, hypercapnia, and hypocapnia. The aim was to demonstrate proof of principle so that future technical developments could assist in transferring this method to the clinical setting. Methods

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Quantification of Cerebral Perfusion With “Real-Time” Contrast-Enhanced Ultrasound

OBJECTIVE To evaluate the use of transient response second harmonic imaging (HI) by means of ultrasound to assess abnormalities of cerebral echo contrast agent enhancement in patients with acute stroke. METHODS The study comprised 25 patients with acute onset of hemispheric stroke ( RESULTS Adequate cerebral contrast enhancement could be seen in 21 subjects. In seven, a large hemispheric deficit of contrast enhancement affecting the entire middle cerebral artery (MCA) territory was detectable; the lentiform nucleus was affected in three subjects. Assessment of cerebral contrast abnormalities was possible in two patients with superficial MCA infarctions but in none of the patients with lacunar ischaemias. None of the control persons had focal deficits of cerebral echo contrast enhancement. In all patients with complete MCA infarction and striatocapsular infarction, presumed ischaemic areas in HI examinations correlated with final CT findings. Overall sensitivity and specifity of HI examinations for predicting size and localisation of the infarction were 75 and 100%, respectively. During follow up, reappearance of contrast enhancement was determined in three patients, in two patients circulatory arrest due to malignant brain oedema with missing contrast enhancement in the entire cerebral hemisphere could be seen. Extent of contrast enhancement deficits significantly correlated with the clinical status on admission and after 1 week (p CONCLUSIONS Second harmonic imaging is the first ultrasonic technique that enables visualisation of pathological cerebral echo contrast enhancement. Because this method identifies deficits of focal contrast enhancement in patients with acute stroke and allows estimation of the final infarct size and clinical prognosis, it may help to select and monitor patients for invasive therapies.

https://jnnp.bmj.com/content/jnnp/69/5/616.full.pdf

Ultrasonic evaluation of pathological brain perfusion in acute stroke using second harmonic imaging

Stroke is one of the major causes of death and disabilities in industrialized countries. Ultrasound imaging is a largely wide spread bedside technique that is easily accessible and valuable in case of emergency but suffers from the fact that the ultrasound wave has to cross the skull for brain imaging. However, ultrasound contrast agents and new contrast-specific imaging modalities have helped to improve the diagnostic quality of transcranial ultrasonography. This review article surveys and discusses the current state of microbubbles technology and the use of contrast-enhanced transcranial ultrasound for the assessment of brain perfusion. Future aspects and expectations in contrast agent functionality, such as targeting and drug or gene delivery, acceleration of thrombolysis, and imaging technology, are also discussed.

Ultrasound contrast agents for brain perfusion imaging and ischemic stroke therapy

Background and Purpose— Initial reports indicate that transcranial harmonic imaging after ultrasound contrast agent bolus injection (BHI) can detect cerebral perfusion deficits in acute ischemic stroke. We evaluated parametric images of the bolus washout kinetics. Methods— Twenty-three patients with acute internal carotid artery infarction were investigated with perfusion harmonic imaging after SonoVue bolus injection ≤40 hour after the onset of symptoms. The findings were compared with those of cranial computed tomography (CCT) and clinical course 4 months after stroke. Results— Images of pixel-wise peak intensity (PPI) and time to peak intensity could be calculated for all patients. Spearman rank correlations of r=0.772 (P<0.001) and r=0.572 (P=0.008) between area of PPI signal decrease and area of infarction in the follow-up CCT as well as outcome after 4 months were obtained, respectively. Conclusions— In the early phase of acute ischemic stroke, BHI after SonoVue bolus injection is a useful ultrasoun...

Ultrasound Perfusion Imaging in Acute Middle Cerebral Artery Infarction Predicts Outcome

Background and Purpose—Through harmonic gray-scale imaging, it is possible to analyze brain tissue perfusion with different ultrasound methods. Methods—In 12 healthy volunteers, 2 doses (0.5 and 1.5 mL) of Optison, a perfluoropropane-containing contrast agent, were injected intravenously and produced a strong increase of brightness in the brain parenchyma. We used harmonic imaging for quantification of ultrasound intensity in the thalamus, ipsilateral temporoparietal white matter (TPWM), and ipsilateral lateral fissure at both sides. Time-intensity curves were calculated, and peak increase (PI) of intensity and the area under the time-intensity curve (AUC) from baseline were compared. Results—We found a significant dose dependence of the AUC in all regions at both sides. PI only showed a significant dose dependence in the TPWM but not in the ipsilateral thalamus and lateral fissure. No side differences for AUC and PI were detected in all regions and doses used. We found a significantly higher value of the...

Harmonic Imaging of the Human Brain Visualization of Brain Perfusion With Ultrasound

Background and Purpose —It is unclear which harmonic imaging mode (power Doppler or gray-scale imaging) is superior and which measuring method is the most robust for the description of brain perfusion.

Methods —We performed an animal study on 6 beagles through the intact skull using a SONOS 5500 device and Optison injected intravenously in 3 different doses (0.15, 0.3, and 0.6 mL). Intensity versus heart-cycle plots for the brain parenchyma and the basal cerebral arteries were generated to evaluate the peak increase (PI) from baseline and the area under the curve (AUC).

Results —With harmonic gray-scale imaging, a homogeneous increase in echo contrast of the brain parenchyma was observed. The effect was dose dependent, resulting in a significant increase in PI as well as an insignificant increase of the AUC with 0.3 mL versus 0.15 mL contrast agent ( P =0.03 and P =0.65, respectively; n=5). With harmonic power Doppler, injection of the 3 different doses resulted in a nonsignificant increase in PI and AUC P =0.17, n=6 for both). After normalization of the brain signal to the peak arterial signal in individual dogs, a significant increase could be demonstrated ( P =0.03 and P =0.01, respectively; n=6). The signal pattern of harmonic power Doppler was inhomogeneous, with stronger signal increases in the anterior part of the brain.

Conclusions —Gray-scale imaging leads to a more homogeneous increase in echo contrast of the brain tissue and may be more suitable for displaying brain perfusion. The PI of the signal intensity seems the most robust parameter for the description of cerebral perfusion with both imaging modes under investigation.

Visualization of Brain Perfusion With Harmonic Gray Scale and Power Doppler Technology An Animal Pilot Study

Harmonic power-based duplex sonography is a new ultrasound method that improves the signal-to-noise ratio of extracranial vascular imaging. The authors evaluated this new method for transtemporal imaging of the basal cerebral arteries. Fundamental power-based duplex sonography (p-TCCS) and harmonic power-based duplex sonography (HI-p-TCCS) in combination with a novel perfluoropropane-containing ultrasound contrast agent (Optison) were investigated for the evaluation of the basal cerebral arteries in 12 healthy volunteers. The number of identified vascular segments and the blood flow velocities in the middle and posterior cerebral arteries were determined for p-TCCS and for two doses of Optison (0.5 and 1.5 mL) using HI-p-TCCS. Furthermore, the authors determined the time course of signal enhancement after Optison bolus injections. The results were compared using Friedman two-way ANOVA test. Significantly more arterial segments were visualized using HI-p-TCCS with enhancement of either 0.5 mL or 1.5 mL Optison (p < 0.01, each) than using p-TCCS. The spatial resolution was markedly increased with HI-p-TCCS, resulting in a striking difference in the detection of distal arterial segments and cortical and parenchymal branches. Except for the diastolic blood flow velocities (BFVs) in the M1 segment, the BFVs did not differ significantly between p-TCCS and HI-p-TCCS. Comparing HI-p-TCCS with 0.5 mL and 1.5 mL Optison, the authors found a small but significant reduction of the latency period (18.2 vs. 15.9 seconds, respectively; p < 0.01), a significant increase of the blooming phase (62.7 vs. 99.8 seconds, respectively; p < 0.0006) and a significant prolongation of the diagnostically useful signal enhancement (233.7 vs. 427.6 seconds, respectively; p < 0.004).

Tobias Katzer

Papers

Harmonic Imaging of the Human Brain Visualization of Brain Perfusion With Ultrasound

Visualization of Brain Perfusion With Harmonic Gray Scale and Power Doppler Technology An Animal Pilot Study

Transcranial harmonic power duplex sonography for the evaluation of cerebral arteries.