A study of diffusion isotropy in single neurons by using NMR microscopy

The apparent diffusion coefficient (ADC) of water was measured in single Aplysia californica neurons by using NMR microscopy encoded in each of two perpendicular gradient directions. Comparisons of the mean ADCs of the gross nuclear and cytoplasmic compartments in five cells, and 50 subregions within these cells, showed no significant difference between the diffusion measurements in the majority of cases. Since anisotropic diffusion would make the ADC dependent on the encoding direction, the results indicate that the ADC in these single neurons is isotropic at the spatial and temporal resolutions used in these studies. Consequently, a single scalar ADC measurement is sufficient for characterizing the ADC in these cells, hence reducing the acquisition time and measurement complexity that would have been required had the ADC been anisotropic.     

Three-dimensional diffusion tensor microscopy of fixed mouse hearts.

The relative utility of 3D, microscopic resolution assessments of fixed mouse myocardial structure via diffusion tensor imaging is demonstrated in this study. Isotropic 100-microm resolution fiber orientation mapping within 5.5 degrees accuracy was achieved in 9.1 hr scan time. Preliminary characterization of the diffusion tensor primary eigenvector reveals a smooth and largely linear angular rotation across the left ventricular wall. Moreover, a higher level of structural hierarchy is evident from the organized secondary and tertiary eigenvector fields. These findings are consistent with the known myocardial fiber and laminar structures reported in the literature and suggest an essential role of diffusion tensor microscopy in developing quantitative atlases for studying the structure-function relationships of mouse hearts.     

Microcoil-based MR phase imaging and manganese enhanced microscopy of glial tumor neurospheres with direct optical correlation

Susceptibility differences among tissues were recently used for highlighting complementary contrast in MRI different from the conventional T(1), T(2), or spin density contrasts. This method, based on the signal phase, previously showed improved image contrast of human or rodent neuroarchitecture in vivo, although direct MR phase imaging of cellular architecture was not available until recently. In this study, we present for the first time the ability of microcoil-based phase MRI to resolve the structure of human glioma neurospheres at significantly improved resolutions (10 × 10 μm(2)) with direct optical image correlation. The manganese chloride property to function as a T(1) contrast agent enabled a closer examination of cell physiology with MRI. Specifically the temporal changes of manganese chloride uptake, retention and release time within and from individual clusters were assessed. The optimal manganese chloride concentration for improved MR signal enhancement was determined while keeping the cellular viability unaffected. The presented results demonstrate the possibilities to reveal structural and functional observation of living glioblastoma human-derived cells. This was achieved through the combination of highly sensitive microcoils, high magnetic field, and methods designed to maximize contrast to noise ratio. The presented approach may provide a powerful multimodal tool that merges structural and functional information of submilimeter biological samples.     

Some new observations on pulse sequence dependent diffusion related edge enhancement in MR microscopy

Self-diffusion of nuclear spins has been suggested to cause edge enhancement in images especially on a microscopic scale. According to previously published work, theory suggests that edge enhancement is caused by motional narrowing due to the boundaries and spin self-diffusion during the data acquisition period. More careful examination reveals that edge enhancement due to motional narrowing develops only under a few specific conditions. This lack of generality of motional narrowing theory, as well as experimental observations, indicate that edge enhancement due to effects other than motional narrowing alone can exist. It is found that edge enhancement depends greatly on the data acquisition mode; therefore, the images obtained are different depending on the pulse sequence employed. For example, excessive attenuation of DC components due to diffusion can result in edge enhancement in the spin echo signal. However, in the case of FID-like signals, DC components are preserved while positive high frequency parts are attenuated, thereby degrading resolution. The new phenomenon observed has been termed selective spectral suppression since the observed edge enhancement results from the selective attenuation of certain frequency components in the nuclear signals due to diffusion-dependent signal attenuation for a given pulse sequence.     

Modulation of water diffusion during gonadotropin-induced ovulation: nmr microscopy of the ovarian follicle

The preovulatory rat follicle reaches a diameter of 1 mm with no internal blood vessels. Nutrient supply to the enclosed oocyte depends solely on passive diffusion across the follicular wall and the follicular fluid. Spin-echo and stimulatedecho NMR microscopy experiments were applied here for studying modulations in water diffusion during gonadotropin-induced maturation of perfused rat ovarian follicles (32°C). Two diffusion compartments were observed for the follicular wall. The intracellular water diffusion coefficient, measured at a short diffusion time (9 ms) was 0.28 x 10^?5 cm²/s. Diffusion at long diffusion times was restricted to 16 μm, the size of cells in the follicular wall, and did not change during maturation. In the follicular fluid a transient 26% decrease in the diffusion coefficient was observed 4–7 h after gonadotropin stimulation, a change that is bound to affect the metabolic balance of the oocyte before ovulation.     

Retrospective distortion correction for 3D MR diffusion tensor microscopy using mutual information and Fourier deformations.

Magnetic resonance diffusion tensor imaging (DTI) can be complicated by distortions that contribute to errors in tissue characterization and loss of fine structures. This work presents a correction scheme based on retrospective registration via mutual information (MI), using Fourier transform (FT)-based deformations to enhance the reliability of the entropy-based image registration. The registration methodology is applied to correct distortions in 3D high-resolution DTI datasets, incorporating a complete set of affine deformations. The results demonstrate that the proposed methodology can consistently and significantly reduce the number of misregistered pixels, leading to marked improvement in the visualization of internal brain white matter (WM) structure via DTI. Post-registration analysis revealed that eddy-current effects cannot fully account for the observed image distortions. Combined, these findings support the non-model-based, postprocessing approach for correcting distortions, and demonstrate the advantages of combining FT-based deformations and MI registration to enhance the practical utility of DTI.     

Statistical model for diffusion attenuated MR signal.

A general statistical model that can describe a rather large number of experimental results related to the structure of the diffusion-attenuated MR signal in biological systems is introduced. The theoretical framework relies on a phenomenological model that introduces a distribution function for tissue apparent diffusion coefficients (ADC). It is shown that at least two parameters--the position of distribution maxima (ADC) and the distribution width (sigma)--are needed to describe the MR signal in most regions of a human brain. A substantial distribution width, on the order of 36% of the ADC, was found for practically all brain regions examined. This method of modeling the MR diffusion measurement allows determination of an intrinsic tissue-specific ADC for a given diffusion time independent of the strength of diffusion sensitizing gradients. The model accounts for the previously found biexponential behavior of the diffusion-attenuated MR signal in CNS.     

MR microscopy of chick embryo vasculature.

Six-day-old chick embryos were examined with magnetic resonance microscopy after vascular perfusion fixation and perfusion with gadolinium-doped gelatin to high-light the developing vascular anatomy. Gadolinium gelatin, with its short T1, provided a source of signal contrast within the vessels. The entire embryo was embedded in gelatin to minimize susceptibility artifacts that are prevalent at the high field strength (7.0 T) used. A series of single-section spin-echo images were acquired with various TRs to determine the optimal imaging sequence for a three-dimensional (3D) acquisition. The combination of gadolinium gelatin in the vascular spaces, gelatin embedding of the specimen, and optimal acquisition parameters yielded a 3D stack of high-resolution images that was readily reconstructed and rendered to effectively demonstrate the developing thoracic vessels in the embryo.     

MR microscopy of perfused brain slices

To study the origins of signal changes in clinical MRI we have previously studied isolated single neuronal cells by MR microscopy. To account for the extracellular environment of the cells, we have developed a prototype perfusion chamber for MR microimaging of perfused rat hippocampal brain slices. To demonstrate the utility of this model, brain slices were initially perfused in isotonic solutions and then subjected to osmotic perturbations via perfusate exchange with 20% hypertonic and 20% hypotonic solutions. In diffusion weighted images, signal intensity changes of +16(σ_n-1 = 11)%(hypotonic) and -26(σ_n-1 = 10)% (hypertonic) were observed. No significant variation in response was observed across the slice when several subregions were examined. These observations are consistent with the view that contrast changes are driven primarily by changes in the intra- and extracellular compartmentation of water. This is the first report of MR microimaging of the isolated brain slice. The technique will enable the correlation of MR microimaging measurements with microscopic changes using other modalities and techniques to provide a better understanding of signals in clinical MRI.     

19F MR imaging of ventilation and diffusion in excised lungs.

Perfluorinated gases, particularly C2F6, are potentially suitable alternatives to hyperpolarized noble gases for pulmonary airspace spin density and diffusion MRI. This work focuses mainly on 19F imaging of C2F6 gas in healthy and emphysematous explanted lungs, avoiding regulatory issues of human in vivo measurements. Three-dimensional gradient echo and spin echo spin density images of human lungs can be made in 10 s with adequate signal-to-noise, demonstrating the feasibility for breathing dynamics to be captured during a succession of short breath holds. As expected, the spin echo images have much smaller susceptibility artifacts than the gradient echo images. 19F and 3He images of the same lungs are compared. The apparent diffusion coefficient (ADC) of C2F6 is sensitive to restrictions imposed by the lung microstructure: the average ADC is measured to be 0.018 cm2/s in healthy lungs versus 0.031 cm2/s in emphysematous lungs at a diffusion time Delta=2.2 ms. The low free diffusivity of pure C2F6 (D0=0.033 cm2/s) places it in a regime where the ADC measurement allows the surface-to-volume ratio to be determined in each voxel, a potentially valuable quantitative characterization of regional lung tissue destruction in emphysema.     

High-Field MR microscopy using fast spin-echoes

Fast spin-echo imaging has been investigated with attention to the requirements and opportunities for high-field MR microscopy. Two-and three-dimensional versions were implemented at 2.0 T, 7.1 T, and 9.4 T. At these fields, at least eight echoes were collectable with a 10 ms TE from fixed tissue specimens and living animals, giving an eightfold improvement in imaging efficiency. To reduce the phase-encoding gradient amplitude and its duty cycle, a modified pulse sequence with phase accumulation was developed. Images obtained using this pulse sequence exhibited comparable signal-to-noise (SNR) to those obtained from the conventional fast spin-echo pulse sequences. Signal losses due to imperfections in RF pulses and lack of phase rewinders were offset in this sequence by reduced diffusion losses incurred with the gradients required for MR microscopy. Image SNR, contrast, edge effects and spatial resolution for three k-space sampling schemes were studied experimentally and theoretically. One method of sampling k-space, 4-GROUP FSE, was found particularly useful in producing varied T₂ contrast at high field. Two-dimensional images of tissue specimens were obtained in a total acquisition time of 1 to 2 min with in-plane resolution between 30 to 70 μm, and 3D images with 256³ arrays were acquired from fixed rat brain tissue (isotropic voxel = 70 μm) and a living rat (isotropic voxel = 117 μm) in∼4.5 h.     

In vivo MR microscopy of the human skin

The requirements for imaging the skin are dictated by the organ's layered structure, which extends only a few millimeters from the surface and thus demands extremely high resolution in this direction. While less critical, resolution in the remaining two dimensions determines whether the skin's accessory structures can be resolved. The problem is compounded by short transverse relaxation times, in particular of the dermis, the structure of most clinical interest. In this work images of the normal human skin were obtained in vivo at voxel sizes as small as 19 × 78 × 800 μm³, by means of customized 3D gradient and partial flip-angle spin-echo pulse sequences and very small transmit/receive coils on a 1.5T clinical imager equipped with high-power whole-body gradients. Structures resolved include hair follicles and the sublayers of the dermis. The very short time constant for the major component (91 %) for transverse relaxation in the dermis (T₂* ～10 ms) suggests the potential of substantial gains in achievable signal-to-noise ratio by shortening the echo time.     

MR microscopy of hyaline cartilage: current status

Cartilage degenerative diseases, such as osteoarthritis, affect million of people. Magnetic resonance imaging is presently the most accurate imaging modality in evaluating the state of hyaline cartilage; however, clinical MRI does not accurately reveal early degenerative alterations in cartilage, due mainly to low spatial resolution. Magnetic resonance microscopy (MRM, or microMRI) appears exceptionally well suited to the in vitro or ex vivo study of this heterogeneous tissue, due to its high spatial resolution; however, despite this, further studies are necessary to evaluate the potential of MRM in the detection of early cartilage damage. Herein we briefly review the current applications of MRM in the study of hyaline cartilage. In particular, we review the MR appearance of hyaline cartilage on high-resolution images, the different MRM techniques used to image normal and enzymatically or chemically degraded cartilage and the potential use of contrast agents. The future directions and the relevance of MRM findings for a better understanding of cartilage physiology in health and disease are also discussed.     

Diffusion-weighted MR microscopy with fast spin-echo

A diffusion-weighted fast spin-echo (FSE) imaging sequence for high-field MR microscopy was developed and experimentally validated in a phantom and in a live rat. Pulsed diffusion gradients were executed before and after the initial 180° pulse in the FSE pulse train. This produced diffusion-related reductions in image signal intensity corresponding to gradient (“b”) factors between 1.80 and 1352 s/mm². The degree of diffusion weighting was demonstrated to be independent of echo train length for experiments using trains up to 16 echoes long. Quantitative measurements on a phantom and on a live rat produced diffusion coefficients consistent with literature values. Importantly, the eight- to 16-fold increase in imaging efficiency with FSE was not accompanied by a significant loss of spatial resolution or contrast. This permits acquisition of in vivo three-dimensional data in time periods that are appropriate for evolving biological processes. The combination of accurate diffusion weighting and high spatial resolution provided by FSE makes the technique particularly useful for MR microscopy.     

A simulation environment for diffusion weighted MR experiments in complex media

Simulations of diffusion in neural tissues have traditionally been limited to analytical solutions, to grid‐based solvers, or to small‐scale Monte Carlo simulations. None of these approaches has had the capability to simulate realistic complex neural tissues on the scale of even a single voxel of reasonable (i.e., clinical) size. An approach is described that combines a Monte Carlo Brownian dynamics simulator capable of simulating diffusion in arbitrarily complex polygonal geometries with a signal integrator flexible enough to handle a variety of pulse sequences. Taken together, this package provides a complete and general simulation environment for diffusion MRI experiments. The simulator is validated against analytical solutions for unbounded diffusion and diffusion between parallel plates. Further results are shown for aligned fibers, varying packing density and permeability, and for crossing straight fibers. Magn Reson Med, 2009. © 2009 Wiley‐Liss, Inc.     

Efficient measurement and calculation of MR diffusion anisotropy images using the Platonic variance method.

The Platonic variance method produces MR diffusion anisotropy (DA) images with a minimum amount of computational effort. It can be programmed in a self-contained MR sequence, thus eliminating the need for postprocessing on a separate workstation. The method uses gradient acquisition schemes, based on Platonic solids: the "icosahedric" scheme (N = 6), the "dodecahedric" scheme (N = 10), and combinations thereof. For these schemes the average of the diffusion tensor eigenvalues equals the average of the measured apparent diffusion coefficients (ADCs), and the variance of the eigenvalues equals 5/2 times the variance of the diffusion coefficients. This results in compact expressions for anisotropy measures, directly in terms of the acquired images, i.e., without calculating the eigenvalues or even the tensor elements. The resulting anisotropy images were shown to be identical to the ones traditionally derived. It is expected that this method will considerably promote the routine use of DA imaging.     

MR microscopy of the lung in small rodents

Understanding how the mammalian respiratory system works and how it changes in disease states and under the influence of drugs is frequently pursued in model systems such as small rodents. These have many advantages, including being easily obtained in large numbers as purebred strains. Studies in small rodents are valuable for proof of concept studies and for increasing our knowledge about disease mechanisms. Since the recent developments in the generation of genetically designed animal models of disease, one needs the ability to assess morphology and function in in vivo systems. In this article, we first review previous reports regarding thoracic imaging. We then discuss approaches to take in making use of small rodents to increase MR microscopic sensitivity for these studies and to establish MR methods for clinically relevant lung imaging.     

Accelerating MR diffusion tensor imaging via filtered reduced-encoding projection-reconstruction.

MR diffusion tensor imaging (DTI) is a promising tool for characterizing the microstructure of ordered tissues. However, its practical applications are hampered by relatively low signal-to-noise-ratio and spatial and temporal resolution. Reduced-encoding imaging (REI) via k-space sharing with constrained reconstruction has previously been shown to be effective for accelerating DTI, although the implementation was based on rectilinear k-space sampling. Due to the intrinsic oversampling of central k-space and allowance for isotropic downsampling, projection-reconstruction (PR) imaging may be better suited for REI. In this study, regularization procedures, including radial filtering and baseline signal correction to adequately reconstruct reduced encoded PR imaging data, are investigated. The proposed filtered reduced-encoding projection-reconstruction (FREPR) technique is applied to DTI tissue fiber orientation and fractional anisotropy (FA) measurements. Results show that FREPR offers improved reconstructions of the reduced encoded images and on an equal total scan-time basis provides more accurate fiber orientation and FA measurements compared to rectilinear k-space sampling-based REI methods or a control experiment consisting of only fully encoded images. These findings suggest a potentially significant role of FREPR in accelerating repeated imaging and improving the data acquisition-time efficiency of DTI experiments.