Jugular Venous Catheterization‐Enhanced CT Angiography for In Vivo 3D Visualization of Cardiopulmonary Vasculature in Sprague‐Dawley Rats

ABSTRACT

Current in vivo imaging techniques for cardiopulmonary vascular evaluation in Sprague‐Dawley (SD) rats face limitations, including structural disruption, inadequate contrast filling, and invasiveness. This study developed a reliable, minimally invasive computed tomography angiography (CTA) technique via jugular vein catheterization for enhanced cardiopulmonary vascular imaging in live SD rats. Jugular vein catheterization was performed in 22 anesthetized healthy male SD rats (320–480 g), followed by dual‐source CT angiography with iopamidol contrast. Three‐dimensional vascular reconstruction was performed, and pulmonary artery width alongside left and right ventricular diameters was measured. CT‐derived measurements were compared with ultrasound data using Bland‐Altman analysis. CTA achieved clear visualization of pulmonary arteries, cardiac chambers, and aortic structures, demonstrating complete contrast filling and anatomical detail. Three‐dimensional reconstructions precisely delineated mediastinal and vascular relationships. Pulmonary artery widths measured by CT and ultrasound showed strong agreement (p > 0.05), validating reliability. Jugular catheterization enabled stable contrast delivery with minimal trauma. Jugular vein catheterization combined with CT angiography provides a safe, accurate, and minimally invasive method for in vivo cardiopulmonary vascular imaging in SD rats. This technique offers high anatomical resolution, compatibility with hemodynamic assessments, and reduced experimental trauma, establishing this approach as a valuable tool for cardiopulmonary disease model research. CT‐derived measurements were compared with ultrasound data using Bland‐Altman analysis. CTA achieved clear visualization of cardiac chambers and pulmonary arteries, demonstrating complete contrast filling and anatomical detail. Three‐dimensional reconstructions precisely delineated mediastinal and vascular relationships. Pulmonary artery widths measured by CT and ultrasound showed strong agreement (p > 0.05), validating reliability. Jugular catheterization enabled stable contrast delivery with minimal trauma. Jugular vein catheterization combined with CT angiography provides a safe, accurate, and minimally invasive method for in vivo cardiopulmonary vascular imaging in SD rats. This technique offers high anatomical resolution, compatibility with hemodynamic assessments, and reduced experimental trauma, establishing this approach as a valuable tool for cardiopulmonary disease model research.

1. Background

Sprague‐Dawley (SD) rats are frequently used as experimental animals in life science research, particularly for creating models of cardiopulmonary vascular diseases. Computed tomography (CT) has emerged as a vital imaging technique in examining the cardiopulmonary vascular system, especially through CT angiography [1]. This method is essential for assessing cardiopulmonary vascular diseases and is also useful for differentiating various pulmonary vascular conditions [2, 3]. As life sciences and information technology continue to advance, the application of CT in experiments with rats is becoming more common.

In recent years, various studies have utilized fillers or contrast agents injected into the cardiopulmonary blood vessels of isolated rats for imaging to examine vascular structures and alterations [4, 5, 6]. However, this approach can lead to unpredictable effects due to the disruption of the thoracic cavity's normal structure, which restricts its use in in vivo experiments. To overcome these challenges, some researchers have turned to tail vein injections of contrast agents in an effort to achieve cardiopulmonary vascular imaging in live rats [7, 8]. Nonetheless, it has been noted that the contrast agent fails to adequately fill the vessels, resulting in images that lack clarity. This poses a significant obstacle for disease observation and diagnosis.

Moreover, various studies have effectively visualized hepatic and abdominal vessels by administering contrast agents into major vessels (such as the femoral vein [9, 10], jugular vein [11], and carotid artery [12]) using high‐pressure syringes for angiographic purposes. The imaging outcomes from these studies are clear and show positive results; however, there have been no reports on their use in the cardiopulmonary vasculature. Jugular vein catheterization is often used in rat models for nutritional support and medication delivery [13, 14], especially in pulmonary hypertension models, while right heart catheterization is commonly utilized to indirectly evaluate pulmonary artery pressure [15, 16]. As a result, there is a growing focus on developing techniques for more precise, safe, and minimally invasive imaging of the cardiopulmonary vascular system in living SD rats.

This study integrated jugular vein catheterization and CT angiography techniques to assess the cardiopulmonary vascular system in SD rats. The goal was to establish a more dependable reference method for CT examinations of related disease models and to investigate new experimental strategies, ultimately offering enhanced technical support for the diagnosis and study of cardiopulmonary vascular diseases in the future.

2. Materials and Methods

2.1. Animal

All animal procedures conducted in this study adhered to the regulations set forth by the Animal Care and Use Committee of Gansu University of Traditional Chinese Medicine, which granted approval for the experiment. The Sprague‐Dawley (SD) rats utilized were sourced from SPF (Beijing) Biotechnology Co. Ltd., and were kept individually in a controlled setting with a temperature of 22 ± 2°C and humidity levels between 40% and 60%. They were maintained on a light‐dark cycle and had unrestricted access to food and water. Healthy male SD rats weighing between 320 and 480 g were chosen to ensure the consistency and reliability of the experimental process.

2.2. Marker Detection

Measurements were taken on 10 SD rats, which included body weight, body length, sternal length, and the distance from the suprasternal fossa to the right atrium. Following euthanasia (administered via intraperitoneal injection of 100 mg/kg chloral hydrate), the distance from the jugular suprasternal fossa to the right atrium was recorded, and the ratio of this distance to the sternal length was determined to be roughly 2/5, serving as a reference for future catheterization procedures (Supplementary Table 1).

2.3. Rat Jugular Vein Catheterization

SD rats were anesthetized using an intraperitoneal injection of 2% sodium pentobarbital (0.3 mL/100 g), then placed on the operating table and secured. The surgical incision was centered in the supraclavicular fossa, approximately 0.5–1 cm on the right side, and the skin was disinfected. An incision was made vertically along this point, about 2–3 cm in length, and the subcutaneous tissue was carefully separated layer by layer to expose the jugular vein, which was about 1–1.5 cm in length. Two absorbable sutures were passed through the jugular vein, with a vascular clip applied to the proximal end and ligation performed on the distal end. A small “V” shaped incision was made in the jugular vein near the ligation site using scissors, and a pre‐flushed jugular venous catheter was slowly inserted to a depth of 2/5 of the sternum length. Once blood flow was observed in the catheter, it and the proximal blood vessels were secured with sutures. To prevent the rat from dislodging the catheter, the distal end was threaded subcutaneously from the right thoracic area to the dorsal back, and the skin on the neck and back was sutured layer by layer after sealing the heparin cap, ensuring the catheter and cap were securely fixed (Figure 1A‐E).

Figure 1.

Jugular venous catheter placement process. A. Exposure of the jugular vein(red arrow) after disinfection. B. Vascular clamp clamps the proximal end, makes it congested and bulging, and ligates the distal end. C. Placement of a jugular venous catheter. D. Thread the intravenous catheter under the skin on the right side of the back. E. Fixation of the catheter on the rat's back.

2.4. CT Angiography

Following the successful placement of the jugular venous catheter, the rat was anesthetized once more using sodium pentobarbital at a dosage of 50 mg/kg, administered intraperitoneally, and positioned prone on the scanning table. The chest of the rat was initially scanned using dual‐source CT (Siemens SOMATOM Force, Germany) from the supraclavicular fossa to the lower border of the ribs. Next, the heparin cap on the jugular venous catheter was removed, and a pressurized syringe was used to inject a contrast medium (iopamol mixed with normal saline in a 1:1 ratio, totaling 2 mL at a rate of 0.1 mL/s). After the contrast agent was injected, a series of perfusion scans were conducted to capture CT images of the cardiopulmonary vasculature over 20 phases, lasting approximately 30 s.

2.5. Animal Ultrasound

For ultrasound detection of the pulmonary arteries, rats are anesthetized using isoflurane at a concentration of 3% and positioned on an animal ultrasound table. The skin on the rat's neck and chest is prepared, and a suitable amount of couplant is applied to both the rat's chest wall and the ultrasound probe. The B‐ultrasound probe is then positioned at the main trunk and bifurcation of the pulmonary artery, the ultrasound image is captured, and the width of the pulmonary artery is measured and documented.

2.6. Statistical Analysis

Statistical analysis of landmark measurement data was conducted using IBM SPSS Statistics 25 software, where the mean and standard error (SEM) for each data group were computed. Bland‐Altman analysis was carried out with GraphPad Prism 9 software. Additionally, CT images in DICOM format were reconstructed into 3D using the Radiant database (R64) for further analysis and comparison.

3. Results

3.1. Chest CT Angiography After Catheterization of the Jugular Vein in Rats Showed Clear Imaging of the Heart and Pulmonary Artery

In a thoracic CT angiography scan conducted on 22 SD rats following jugular vein catheterization, the outlines of the right atrium, right ventricle, left atrium, and left ventricle were distinctly visible due to the injection of contrast medium (iopamol). The differences in size between the left and right heart chambers, as well as the overall shape of the heart, were easily identifiable (Figure 2A‐D). The imaging of the pulmonary artery revealed a clear outline of the main pulmonary artery trunk and its left and right branches, with the contrast medium filling appearing intact (Figure 3A‐C). Additionally, the morphology of the pulmonary artery was observable in the sagittal image (Figure 3D), providing a solid anatomical foundation for future functional assessments.

Figure 2.

Cardiac imaging with CT angiography in the chest of a rat. A. Bilateral cardiac chambers without perfusion. B. Right ventricular perfusion imaging. C. Left ventricular perfusion imaging. D. Both the left and right ventricular perfusion imaging. LV, left ventricular; RV, right ventricular.

Figure 3.

Pulmonary artery image of rat chest CT angiography. A. Pulmonary artery without perfusion. B. Pulmonary artery trunk perfusion imaging with beginning imaging of left and right pulmonary artery branches. C. Perfusion imaging of left and right pulmonary artery branches. D. The pulmonary artery can be clearly seen on sagittal CT (white arrow). PA, pulmonary artery; TA, thoracic aorta.

3.2. Three‐Dimensional Visualization of CT Angiography in the Chest of Rats

By digitally reconstructing the enhanced CT scan results of a rat's chest, the 3D CT reconstruction image effectively displayed the anatomical features of the pulmonary artery, aorta, and the cardiopulmonary vascular system (Figure 4A‐F). CT 3D imaging can distinctly identify the relationships among mediastinal tissues, organs, and blood vessels, aiding in the assessment of blockages, ligations, and compressions of mediastinal tissues. This provides a visual foundation for evaluating both anatomical and pathological changes in the vascular system.

Figure 4.

Three‐dimensional reconstruction of rat chest CT. A. CT angiography 3D reconstruction (frontal view). B‐F. Three‐dimensional reconstruction of cardiopulmonary angiography showed the pulmonary artery (white dovetail arrow) and aorta (white arrow). B. CT cardiopulmonary angiography (frontal view). C. CT cardiopulmonary angiography (left anterior lateral view). D. CT cardiopulmonary angiography (frontal view, image after deboning). E. CT cardiopulmonary angiography (left posterior lateral view, image after deboning). F. CT cardiopulmonary angiography (posterior view, image after deboning).

3.3. Rat CT Pulmonary Artery Imaging Has Strong Reliability

Following CT imaging of the pulmonary arteries, left ventricular diameter, and right ventricular diameter in SD rats, the width of these structures was assessed and compared with measurements obtained by echocardiography. Pulmonary artery (PA) width measurements were successfully obtained using both transthoracic UCG and CT perfusion imaging in all 22 rats included in the comparative analysis. Representative images depicting the PA width measurement procedures for each modality are presented in UCG and CT perfusion imaging (Figure 5A and B). Statistical comparison of the PA width values derived from the two imaging techniques revealed no significant difference. A paired Student's t‐test performed on the measurements yielded a p‐value greater than 0.05 (Figure 5C), indicating that the mean PA widths measured by UCG and CT perfusion imaging were not statistically distinct. To further assess the agreement between the two measurement methods beyond simple mean comparison, Bland‐Altman analysis was conducted (Figure 5D). At the same time, the consistency between CT and ultrasound measurements of left ventricular and right ventricular structures was also high (Figure S1, S2). This analysis plotted the difference between paired UCG and CT measurements against their average value. The Bland‐Altman plot demonstrated good agreement between UCG and CT perfusion imaging for assessing Pulmonary artery width in this rat model. The majority of data points fell within the limits of agreement, suggesting consistent measurement performance across the range of observed PA widths without evidence of significant systematic bias between the two techniques.

Figure 5.

CT angiography validation. A. UCG measurement of pulmonary artery width in rats; B. CT perfusion imaging measurement of pulmonary artery width. C. shows no statistical significance by performing a t‐test on UCG versus CT perfusion imaging pulmonary artery width. D. Bland‐Altman analysis was performed on UCG and CT pulmonary artery width measurements (n = 22).

4. Discussion

In this study, we explored the effectiveness of CT angiography for examining the cardiopulmonary vascular system in living SD rats. The findings indicated that this method can clearly visualize the heart and pulmonary arteries while preserving the integrity of the thoracic cavity, and the measurement data obtained are both reliable and safe. As a minimally invasive imaging technique, CT angiography allows for high‐quality visualization of the cardiopulmonary system through jugular catheterization, minimizing trauma to the animals and providing robust technical support for studies on cardiopulmonary vascular diseases.

The use of in vivo imaging in this study offers significant advantages over ex vivo approaches by preserving the physiological integrity of the vascular system. This methodology maintains native hemodynamic conditions, including physiological pressure, shear stress, and vascular tone, thereby enabling functional assessment of vessels within an intact circulatory environment. Furthermore, in vivo imaging uniquely captures systemic disease influences on vascular pathophysiology, which are challenging to replicate in isolated ex vivo preparations due to the absence of integrated neurohormonal and immune responses [17]. Beyond physiological relevance, in vivo imaging provides critical dynamic and structural insights that ex vivo models cannot achieve. In contrast, ex vivo analyses yield only static endpoint data and inherently disrupt vascular anatomy, precluding observation of vessel‐tissue interactions [18].

CT angiography effectively reveals the main pulmonary artery and its left and right branches, as well as the outlines of the heart chambers. The measurements of pulmonary artery width and left, right ventricular diameter obtained through CT imaging align closely with those from echocardiography, indicating that CT angiography is both feasible and accurate for evaluating the cardiopulmonary vascular system in rats. This is particularly relevant for research on pulmonary artery width, ventricular size, and hemodynamic changes, as it yields precise imaging data.

Moreover, the technique of placing a jugular vein catheter offers a safe and stable access point for conducting CT angiography in rats, simplifying the procedure and significantly lowering the risk of trauma to the animals. This method is not only effective for angiography but also for measuring right ventricular pressure in cardiopulmonary disease models, thereby broadening its applicability in experimental research.

However, it is important to note that using jugular vein cannulation in rat models carries certain risks, including catheter blockage, leakage, bleeding, and infection, which could result in experimental failure [19]. Therefore, implementing strict disinfection protocols, securing the catheter, and conducting postoperative evaluations are crucial to ensure the success of the experiment and the reliability of the data. This study also has several limitations: 1. It has not been further validated in models of pulmonary vascular and cardiovascular diseases (such as pulmonary hypertension, heart failure, etc.); 2. The survival period of rats after CT cardiovascular and pulmonary imaging has not been further observed. These will be issues that need to be addressed in our next study.

In summary, this study highlights the high feasibility and safety of CT angiography for assessing the cardiopulmonary vascular system in living SD rats. This technique serves as an effective imaging tool for developing models of cardiopulmonary vascular‐related diseases and will aid in advancing experimental research on cardiopulmonary conditions in the future.

Author Contributions

Tongtong Gao: methodology, writing – original draft, writing – review and editing. Huan Liu: methodology, writing – original draft, writing – review and editing. Jianwei He: writing – review and editing. Like Ma: data curation, writing – review and editing. Mengyu Wu: writing – review and editing. Yunshan Cao: methodology, supervision, writing – review and editing. Xiaozhou Long: funding acquisition, methodology, supervision, and writing – review and editing.

Ethics Statement

The studies involving humans were approved by the Medical Ethics Committee of Gansu University of Chinese Medicine.

Conflicts of Interest

The authors declare no conflicts of interest.

Supporting information

Figure S1: CT angiography validation. UCG measurement of right ventricular diameter in rats; B. CT perfusion imaging measurement of right ventricular diameter. C. shows no statistical significance by performing a t‐test on UCG versus CT perfusion imaging right ventricular diameter. D. Bland‐Altman analysis was performed on UCG and CT right ventricular diameter measurements(n = 22).

Figure S2: CT angiography validation. A. UCG measurement of left ventricular diameter in rats; B. CT perfusion imaging measurement of left ventricular diameter. C. shows no statistical significance by performing a t‐test on UCG versus CT perfusion imaging left ventricular diameter. D. Bland‐Altman analysis was performed on UCG and CT left ventricular diameter measurements(n = 22).

Supplementary Table 1: Measurement of each landmark point in rats.

Data Availability Statement

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.