High-Resolution Cardiac Positron Emission Tomography/Computed Tomography for Small Animals

The presented cardiac PET/CT protocol is useful for obtaining functional and morphological information in a variety of small animal models of cardiac disease. Advantages of PET/CT imaging as compared to other preclinical imaging modalities include, but are not limited to, high sensitivity and very high spatial temporal resolution, as well as robustness as it does not require manual positioning of probes by the operators. One of the main disease model that can be studied with this protocol is myocardial infarction.

However, other cardiometabolic diseases and also response to therapy can be investigated. For instance, our group is interested in the role of obesity and diabetes on cardiac metabolism and function. The steps included in this protocol can be easily followed by users without experience.

However, some steps such as caudal vein cannulations and blood sampling may require some training and experience to be done reproducibly. Demonstrating the procedure will be Federica La Rosa, a research fellow from our laboratory, Federico Granziera, a PhD student from Sant'Anna School of Advanced Studies, and Domiziana Terlizzi, veterinary doctor of Fondazione Toscana Gabriele Monasterio. To begin, place the mouse in the supine position head first on the scanner bed of the PET/CT tomograph putting its nose in the nose mask for anesthesia and gently blocking the head of the mask with adhesive tape.

Fix the upper and lower limbs on the scanner bed to prevent involuntary movements during imaging procedures which may lead to motion artifacts. Monitor the body temperature using a rectal probe and respiration rate using a respiration pillow. For mice, draw 100 to 150 microliters of 10 megabecquerels fluorine-18 FDG using an insulin syringe.

If the original concentration of the tracer in the vial is high, dilute the tracer with saline to a concentration of 50 to 100 megabecquerel per milliliter. Use the PET dose calibrator to measure the actual activity in the syringe. Annotate the pre-injection activity and time of measurement as these values will be used later using specific input modules of the PET scanner graphical user interface.

If using iomeprol, set the injection rate to 10 milliliters per hour and volume to 0.5 milliliters. Connect the syringe to the syringe pump and set the pump for the actual syringe size and diameter, then connect the syringe to the CA tubing and needle and prefill the tubing with the CA.Set the injection rate to 10 milliliters per hour, limiting the injection volume to 0.5 milliliters. For iomeprol injection, use a syringe pump allowing slow injection at a constant rate, with an already set injection rate of 10 milliliters per hour.

Limiting the injection volume to 0.5 milliliters, stop the injection after three minutes. After ensuring the tubing and needle are prefilled with CA, connect the needle attached to the CA tubing to the tail vein's cannula. Start the injection.

Close the scanner's lid and prepare for the Cine-CT scan. Press the Continue button on the tomograph's user interface after 60 seconds from the beginning of the injection so that the Cine-CT acquisition is started. The injection of CA will stop roughly at the same time as the Cine-CT scan completion.

Open the DICOM images of the dynamic PET scan. Select the heart plugin module. Zoom into the mouse or rat heart image and select the last timeframe or the sum of the last three to five timeframes for which most of the blood pool activity has already been washed out.

Follow the onscreen instruction to reorient the image along the principal axis of the animal heart. Do this interactively by moving the displayed markers for the heart base and apex. Next, select the Segmentation tool.

If the result of the automatic segmentation is not acceptable, refine the shape of the segmented myocardium or left ventricle cavity by enabling the manual mode ROI search disabled. In the modeling tool, select the appropriate kinetic model or dynamic PET analysis. In this case, select graphical and then Patlak to enable the Patlak plot analysis to compute the metabolic rate of glucose uptake for each cardiac sector.

Next, in the polar map tool, select the correct number of displayed heart segments. In this case, select 17 segments. Now, press the Fit button to perform the fitting procedure of the Patlak analysis.

At the end of the fitting procedure, observe the displayed polar map of the KI values. Load the DICOM images of the Cine-CT scan in the software. Then open the dynamic dataset with the built-in 4D viewer.

And using the 3D multiplayer reformation or MPR tool, reorient the image data along the short axis. Export the reoriented data to DICOM, ensuring that the entire 4D data are exported with preserved slice thickness and image bit depth of 16 bits per voxel. Open the exported 4D MPR images using the 4D viewer, then select a timeframe corresponding to end-diastole and browse through all the timeframes with the time slider on the main toolbar to ensure that the correct cardiac phase is selected.

On this timeframe, pick the closed polygon annotation tool and manually delineate the endocardial wall of the left ventricle. Do the same for 10 to 20 slices from the base to the apex ensuring that all the ROIs have the same name. Next on the ROI menu, select ROI volume, then generate missing ROIs to generate the ROIs on all the short axis slices by interpolation of the manually drawn ROIs.

Then select ROI volume, followed by compute volume to calculate the volume of the ROI group with the same ROI name. Next, browse through the timeframes, select a phase corresponding to end-systole, and calculate the volume of the ROI group with the same ROI name as demonstrated. Finally, calculate the stroke volume and ejection fraction using the equation described in the manuscript.

The results of the automatic myocardial and left ventricle cavity segmentation of a control CD1 mouse are shown here. Even in healthy subjects, lower reconstructed values around the apex are commonly observed in PET. The Patlak graphical analysis demonstrated an example of the regional KI, the scatter plot and linear regression analysis, and the values of the slope and intercept of the linear fit performed on each segment along with the corresponding coefficient of determination.

In the healthy rat, different shapes and sizes of the left ventricle are shown for the end-diastolic and end-systolic phases. With the 3D reconstruction of the segmented left ventricle volume, the calculation of volumes resulted in end-diastolic volume of 0.361 milliliters and an end-systolic volume of 0.038 milliliters. A volume rendering of the same rat heart for the end-diastole and end-systole allows for depicting the iodine enhanced chambers and vessels so their value is more qualitative than quantitative.

Maintain the level of anesthesia and often check the physiological parameters of the animal during the study. Moreover, an initial check of the patency of the cannulation must be done to avoid unsuccessful injections. Due to the non-invasive nature of this study, the animal can be recovered after the procedure and decay of the radioactive tracer so that virtually, any other investigation method can be applied.

Otherwise, if the animal is going to be euthanized after PET/CT imaging, all standard ex vivo analysis on excited tissues can be performed.