Beyond Cytoarchitectonics: The Internal and External Connectivity Structure of the Caudate Nucleus

While there is ample evidence on the functional and connectional differentiation of the caudate nucleus (CN), less is known about its potential microstructural subdivisions. However, this latter aspect is critical to the local information processing capabilities of the tissue. We applied diffusion MRI, a non-invasive in vivo method that has great potential for the exploration of the brain structure-behavior relationship, in order to characterize the local fiber structure in gray matter of the CN. We report novel evidence of a functionally meaningful structural tri-partition along the anterior-posterior axis of this region. The connectivity of the CN subregions is in line with connectivity evidence from earlier invasive studies in animal models. In addition, histological validation using polarized light imaging (PLI) confirms these results, corroborating the notion that cortico-subcortico-cortical loops involve microstructurally differentiated regions in the caudate nucleus. Methodologically speaking, the comparison with advanced analysis of diffusion MRI shows that diffusion tensor imaging (DTI) yields a simplified view of the CN fiber architecture which is refined by advanced high angular resolution imaging methods.     

In vivo High Angular Resolution Diffusion-Weighted Imaging of Mouse Brain at 16.4 Tesla

Magnetic Resonance Imaging (MRI) of the rodent brain at ultra-high magnetic fields (> 9.4 Tesla) offers a higher signal-to-noise ratio that can be exploited to reduce image acquisition time or provide higher spatial resolution. However, significant challenges are presented due to a combination of longer T ₁ and shorter T ₂/T₂* relaxation times and increased sensitivity to magnetic susceptibility resulting in severe local-field inhomogeneity artefacts from air pockets and bone/brain interfaces. The Stejskal-Tanner spin echo diffusion-weighted imaging (DWI) sequence is often used in high-field rodent brain MRI due to its immunity to these artefacts. To accurately determine diffusion-tensor or fibre-orientation distribution, high angular resolution diffusion imaging (HARDI) with strong diffusion weighting (b >3000 s/mm²) and at least 30 diffusion-encoding directions are required. However, this results in long image acquisition times unsuitable for live animal imaging. In this study, we describe the optimization of HARDI acquisition parameters at 16.4T using a Stejskal-Tanner sequence with echo-planar imaging (EPI) readout. EPI segmentation and partial Fourier encoding acceleration were applied to reduce the echo time (TE), thereby minimizing signal decay and distortion artefacts while maintaining a reasonably short acquisition time. The final HARDI acquisition protocol was achieved with the following parameters: 4 shot EPI, b = 3000 s/mm², 64 diffusion-encoding directions, 125×150 μm² in-plane resolution, 0.6 mm slice thickness, and 2h acquisition time. This protocol was used to image a cohort of adult C57BL/6 male mice, whereby the quality of the acquired data was assessed and diffusion tensor imaging (DTI) derived parameters were measured. High-quality images with high spatial and angular resolution, low distortion and low variability in DTI-derived parameters were obtained, indicating that EPI-DWI is feasible at 16.4T to study animal models of white matter (WM) diseases.     

Cranial MRI of small rodents using a clinical MR scanner

Increasing numbers of small animal models are in use in the field of neuroscience research. Magnetic resonance imaging (MRI) provides an excellent method for non-invasive imaging of the brain. Using three-dimensional (3D) MR sequences allows lesion volumetry, e.g. for the quantification of tumor size. Specialized small-bore animal MRI scanners are available for high-resolution MRI of small rodents' brain, but major drawbacks of this dedicated equipment are its high costs and thus its limited availability. Therefore, more and more research groups use clinical MR scanners for imaging small animal models. But to achieve a reasonable spatial resolution at an acceptable signal-to-noise ratio with these scanners, some requirements concerning sequence parameters have to be matched. Thus, the aim of this paper was to present in detail a method how to perform MRI of small rodents brain using a standard clinical 1.5 T scanner and clinically available radio frequency coils to keep material costs low and to circumvent the development of custom-made coils.     

Bremsstrahlung radiation detection for small animal imaging using a CCD detector

The use of optical methods for the detection of radionuclides is becoming an established tool for preclinical molecular imaging experiments. In this paper we present a set of proof of principle experiments showing that planar bremsstrahlung radiation images can be detected with an intensifying screen using a small animal optical imager based on charge coupled device detector.We develop a bremsstrahlung source using a ³²P-ATP vial placed in a Plexiglas box, the source with an intensifying screen on top was placed inside a small animal optical imaging system. Bremsstrahlung radiation images were produced with the ³²P-ATP source only and also with a pair of pliers placed between the source and the screen. We found that the pair of pliers absorption image matches the shape of the object.Spatial resolution measurements were not performed however, the bremsstrahlung image of the pliers show that the resolution is relatively poor due to a large penumbra effect.We conclude that it is possible to produce planar bremsstrahlung images using optical imaging devices.     

Development of a dynamic imaging method for gravitropism in pea sprouts using clinical magnetic resonance imaging system

Although magnetic resonance imaging (MRI) is a useful technique, only a few studies have investigated the dynamic behavior of small subjects using MRI owing to constraints such as experimental space and signal amount. In this study, to acquire high-resolution continuous three-dimensional gravitropism data of pea (Pisum sativum) sprouts, we developed a small-bore MRI signal receiver coil that can be used in a clinical MRI and adjusted the imaging sequence. It was expected that such an arrangement would improve signal sensitivity and improve the signal-to-noise ratio (SNR) of the acquired image. All MRI experiments were performed using a 3.0-T clinical MRI scanner. An SNR comparison using an agarose gel phantom to confirm the improved performance of the small-bore receiver coil and an imaging experiment of pea sprouts exhibiting gravitropism were performed. The SNRs of the images acquired with a standard 32-channel head coil and the new small-bore receiver coil were 5.23±0.90 and 57.75±12.53, respectively. The SNR of the images recorded using the new coil was approximately 11-fold higher than that of the standard coil. In addition, when the accuracy of MR imaging that captures the movement of pea sprout was verified, the difference in position information from the optical image was found to be small and could be used for measurements. These results of this study enable the application of a clinical MRI system for dynamic plant MRI. We believe that this study is a significant first step in the development of plant MRI technique.     

A new technique for in vivo imaging of specific GLP-1 binding sites: first results in small rodents

EXPERIMENTAL OBJECTIVES: In vivo imaging of GLP-1 receptor-positive tissues may allow examination of physiologic and pathophysiologic processes. Based on the GLP-1 analog Exendin 4, we have developed a radiolabeled compound specifically targeting the GLP-1 receptor (DTPA-Lys40-Exendin 4). This work aims to detect GLP-1 receptor-positive tissues by biodistribution studies and in vivo small animal imaging studies. For in vivo imaging, a high-resolution multi-pinhole SPECT (single photon emission computed tomography) system was used in conjunction with an MRI (magnetic resonance imaging) system for image fusion. RESULTS: DTPA-Lys40-Exendin 4 can be labeled with 111In to high specific activity (40 GBq/micromol). The radiochemical purity reliably exceeded 95%. Using this compound for in vivo small animal imaging of rats and mice as well as for biodistribution studies, specific GLP-1 binding sites could be detected in stomach, pancreas, lung, adrenals, and pituitary. Receptor-positive tissues were visualized with a high-resolution SPECT system with a resolution of less than 1 mm. CONCLUSIONS: The new technique using DTPA-Lys40-Exendin 4 allows highly sensitive imaging of GLP-1 receptor-positive tissues in vivo. Therefore, intra-individual follow-up studies of GLP-1 receptor-positive tissue could be conducted in vivo.     

Design of a multimodal fibers optic system for small animal optical imaging

Small animals optical imaging systems are widely used in pre-clinical research to image in vivo the bio-distribution of light emitting probes using fluorescence or bioluminescence modalities. In this work we presented a set of simulated results of a novel small animal optical imaging module based on a fibers optics matrix, coupled with a position sensitive detector, devoted to acquire bioluminescence and Cerenkov images. Simulations were performed using GEANT 4 code with the GAMOS architecture using the tissue optics plugin. Results showed that it is possible to image a 30 × 30 mm region of interest using a fiber optics array containing 100 optical fibers without compromising the quality of the reconstruction. The number of fibers necessary to cover an adequate portion of a small animal is thus quite modest. This design allows integrating the module with magnetic resonance (MR) in order to acquire optical and MR images at the same time. A detailed model of the mouse anatomy, obtained by segmentation of 3D MRI images, will improve the quality of optical 3D reconstruction.     

Magnetic resonance imaging using a clinical whole body system: an introduction to a useful technique in small animal experiments.

A clinical whole body magnetic resonance imaging (MRI) system with high resolution coils was used to obtain non-invasive images of the living rat. The results demonstrate the feasibility of the set-up and the advantages of this new imaging technique: detailed information, no extra costs, longitudinal studies without killing animals and simple anaesthesia. It is concluded that in small animal experimentation, this use of high resolution coils in whole body magnetic resonance systems may be particularly helpful in establishing effects of experimental procedures. Whenever non-invasive visualization is required, especially in longitudinal animal studies, e.g. biomaterial research or tumour investigation, this use of MRI will offer challenging possibilities.     

Current advances in molecular imaging: noninvasive in vivo bioluminescent and fluorescent optical imaging in cancer research

Recently, there has been tremendous interest in developing techniques such as MRI, micro-CT, micro-PET, and SPECT to image function and processes in small animals. These technologies offer deep tissue penetration and high spatial resolution, but compared with noninvasive small animal optical imaging, these techniques are very costly and time consuming to implement. Optical imaging is cost-effective, rapid, easy to use, and can be readily applied to studying disease processes and biology in vivo. In vivo optical imaging is the result of a coalescence of technologies from chemistry, physics, and biology. The development of highly sensitive light detection systems has allowed biologists to use imaging in studying physiological processes. Over the last few decades, biochemists have also worked to isolate and further develop optical reporters such as GFP, luciferase, and cyanine dyes. This article reviews the common types of fluorescent and bioluminescent optical imaging, the typical system platforms and configurations, and the applications in the investigation of cancer biology.     

Magnetization Transfer Prepared Gradient Echo MRI for CEST Imaging

Chemical exchange saturation transfer (CEST) is an emerging MRI contrast mechanism that is capable of noninvasively imaging dilute CEST agents and local properties such as pH and temperature, augmenting the routine MRI methods. However, the routine CEST MRI includes a long RF saturation pulse followed by fast image readout, which is associated with high specific absorption rate and limited spatial resolution. In addition, echo planar imaging (EPI)-based fast image readout is prone to image distortion, particularly severe at high field. To address these limitations, we evaluated magnetization transfer (MT) prepared gradient echo (GRE) MRI for CEST imaging. We proved the feasibility using numerical simulations and experiments in vitro and in vivo. Then we optimized the sequence by serially evaluating the effects of the number of saturation steps, MT saturation power (B1), GRE readout flip angle (FA), and repetition time (TR) upon the CEST MRI, and further demonstrated the endogenous amide proton CEST imaging in rats brains (n = 5) that underwent permanent middle cerebral artery occlusion. The CEST images can identify ischemic lesions in the first 3 hours after occlusion. In summary, our study demonstrated that the readily available MT-prepared GRE MRI, if optimized, is CEST-sensitive and remains promising for translational CEST imaging.     

Ultrasound Imaging-guided Intracardiac Injection to Develop a Mouse Model of Breast Cancer Brain Metastases Followed by Longitudinal MRI

Breast cancer brain metastasis, occurring in 30% of breast cancer patients at stage IV, is associated with high mortality. The median survival is only 6 months. It is critical to have suitable animal models to mimic the hemodynamic spread of the metastatic cells in the clinical scenario. Here, we are introducing the use of small animal ultrasound imaging to guide an accurate injection of brain tropical breast cancer cells into the left ventricle of athymic nude mice. Longitudinal MRI is used to assessing intracranial initiation and growth of brain metastases. Ultrasound-guided intracardiac injection ensures not only an accurate injection and hereby a higher successful rate but also significantly decreased mortality rate, as compared to our previous manual procedure. In vivo high resolution MRI allows the visualization of hyperintense multifocal lesions, as small as 310 µm in diameter on T₂-weighted images at 3 weeks post injection. Follow-up MRI reveals intracranial tumor growth and increased number of metastases that distribute throughout the whole brain.     

An In Vivo Three-Dimensional Magnetic Resonance Imaging-Based Averaged Brain Collection of the Neonatal Piglet (Sus scrofa)

Due to the fact that morphology and perinatal growth of the piglet brain is similar to humans, use of the piglet as a translational animal model for neurodevelopmental studies is increasing. Magnetic resonance imaging (MRI) can be a powerful tool to study neurodevelopment in piglets, but many of the MRI resources have been produced for adult humans. Here, we present an average in vivo MRI-based atlas specific for the 4-week-old piglet. In addition, we have developed probabilistic tissue classification maps. These tools can be used with brain mapping software packages (e.g. SPM and FSL) to aid in voxel-based morphometry and image analysis techniques. The atlas enables efficient study of neurodevelopment in a highly tractable translational animal with brain growth and development similar to humans.     

Merging computational fluid dynamics and 4D Flow MRI using proper orthogonal decomposition and ridge regression

Time resolved phase-contrast magnetic resonance imaging 4D-PCMR (also called 4D Flow MRI) data while capable of non-invasively measuring blood velocities, can be affected by acquisition noise, flow artifacts, and resolution limits. In this paper, we present a novel method for merging 4D Flow MRI with computational fluid dynamics (CFD) to address these limitations and to reconstruct de-noised, divergence-free high-resolution flow-fields. Proper orthogonal decomposition (POD) is used to construct the orthonormal basis of the local sampling of the space of all possible solutions to the flow equations both at the low-resolution level of the 4D Flow MRI grid and the high-level resolution of the CFD mesh. Low-resolution, de-noised flow is obtained by projecting in vivo 4D Flow MRI data onto the low-resolution basis vectors. Ridge regression is then used to reconstruct high-resolution de-noised divergence-free solution. The effects of 4D Flow MRI grid resolution, and noise levels on the resulting velocity fields are further investigated. A numerical phantom of the flow through a cerebral aneurysm was used to compare the results obtained using the POD method with those obtained with the state-of-the-art de-noising methods. At the 4D Flow MRI grid resolution, the POD method was shown to preserve the small flow structures better than the other methods, while eliminating noise. Furthermore, the method was shown to successfully reconstruct details at the CFD mesh resolution not discernible at the 4D Flow MRI grid resolution. This method will improve the accuracy of the clinically relevant flow-derived parameters, such as pressure gradients and wall shear stresses, computed from in vivo 4D Flow MRI data.     

First combined in vivo X-ray tomography and high-resolution molecular electron paramagnetic resonance (EPR) imaging of the mouse knee joint taking into account the disappearance kinetics of the EPR probe

Although laboratory data clearly suggest a role for oxidants (dioxygen and free radicals derived from dioxygen) in the pathogenesis of many age-related and degenerative diseases (such as arthrosis and arthritis), methods to image such species in vivo are still very limited. This methodological problem limits physiopathologic studies about the role of those species in vivo, the effects of their regulation using various drugs, and the evaluation of their levels for diagnosis of degenerative diseases. In vivo electron paramagnetic resonance (EPR) imaging and spectroscopy are unique, noninvasive methods used to specifically detect and quantify paramagnetic species. However, two problems limit their application: the anatomic location of the EPR image in the animal body and the relative instability of the EPR probes. Our aim is to use EPR imaging to obtain physiologic and pathologic information on the mouse knee joint. This article reports the first in vivo EPR image of a small tissue, the mouse knee joint, with good resolution (≈ 160 μm) after intra-articular injection of a triarylmethyl radical EPR probe. It was obtained by combining EPR and x-ray micro-computed tomography for the first time and by taking into account the disappearance kinetics of the EPR probe during image acquisition to reconstruct the image. This multidisciplinary approach opens the way to high-resolution EPR imaging and local metabolism studies of radical species in vivo in different physiologic and pathologic situations.     

Interpolated Compressed Sensing for 2D Multiple Slice Fast MR Imaging

Sparse MRI has been introduced to reduce the acquisition time and raw data size by undersampling the k-space data. However, the image quality, particularly the contrast to noise ratio (CNR), decreases with the undersampling rate. In this work, we proposed an interpolated Compressed Sensing (iCS) method to further enhance the imaging speed or reduce data size without significant sacrifice of image quality and CNR for multi-slice two-dimensional sparse MR imaging in humans. This method utilizes the k-space data of the neighboring slice in the multi-slice acquisition. The missing k-space data of a highly undersampled slice are estimated by using the raw data of its neighboring slice multiplied by a weighting function generated from low resolution full k-space reference images. In-vivo MR imaging in human feet has been used to investigate the feasibility and the performance of the proposed iCS method. The results show that by using the proposed iCS reconstruction method, the average image error can be reduced and the average CNR can be improved, compared with the conventional sparse MRI reconstruction at the same undersampling rate.     

Anesthesia and other considerations for in vivo imaging of small animals

The use of small animal imaging is increasing in biomedical research thanks to its ability to localize altered biochemical and physiological processes in the living animal and to follow these processes longitudinally and noninvasively. In contrast to human studies, however, imaging of small animals generally requires anesthesia, and anesthetic agents can have unintended effects on animal physiology that may confound the results of the imaging studies. In addition, repeated anesthesia, animal preparation for imaging, exposure to ionizing radiation, and the administration of contrast agents may affect the processes under study. We discuss this interplay of factors for small animal imaging in the context of four common imaging modalities for small animals: positron emission tomography (PET) and single photon emission computed tomography (SPECT), computed tomography (CT), magnetic resonance imaging (MRI), and optical imaging. We discuss animal preparation for imaging, including choice of animal strain and gender, the role of fasting and diet, and the circadian cycle. We review common anesthesias used in small animal imaging, such as pentobarbital, ketamine/xylazine, and isoflurane, and describe techniques for monitoring the respiration and circulation of anesthetized animals that are being imaged as well as developments for imaging conscious animals. We present current imaging literature exemplifying how anesthesia and animal handling can influence the biodistribution of PET tracers. Finally, we discuss how longitudinal imaging studies may affect animals due to repeated injections of radioactivity or other substrates and the general effect of stress on the animals. In conclusion, there are many animal handling issues to consider when designing an imaging experiment. Reproducible experimental conditions require clear, consistent reporting, in the study design and throughout the experiment, of the animal strain and gender, fasting, anesthesia, and how often individual animals were imaged.     

In vivo19F MRI for Cell Tracking

In vivo¹⁹F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. Here, we describe a general protocol for cell labeling, imaging, and image processing. The technique is applicable to various cell types and animal models, although here we focus on a typical mouse model for tracking murine immune cells. The most important issues for cell labeling are described, as these are relevant to all models. Similarly, key imaging parameters are listed, although the details will vary depending on the MRI system and the individual setup. Finally, we include an image processing protocol for quantification. Variations for this, and other parts of the protocol, are assessed in the Discussion section. Based on the detailed procedure described here, the user will need to adapt the protocol for each specific cell type, cell label, animal model, and imaging setup. Note that the protocol can also be adapted for human use, as long as clinical restrictions are met.     

Current issues and perspectives in small rodent magnetic resonance imaging using clinical MRI scanners

Small rodents such as mice and rats are frequently used in animal experiments for several reasons. In the past, animal experiments were frequently associated with invasive methods and groups of animals had to be killed to perform longitudinal studies. Today's modern imaging techniques such as magnetic resonance imaging (MRI) allow non-invasive longitudinal monitoring of multiple parameters. Although only a few institutions have access to dedicated small animal MR scanners, most institutions carrying out animal experiments have access to clinical MR scanners. Technological advances and the increasing field strength of clinical scanners make MRI a broadly available and viable technique in preclinical in vivo research. This review provides an overview of current concepts, limitations, and recent studies dealing with small animal imaging using clinical MR scanners.