Stephen J. Dodd

Detection of Single Mammalian Cells by High-Resolution Magnetic Resonance Imaging

This study reports the detection of single mammalian cells, specifically T cells (T lymphocytes) labeled with dextran-coated superparamagnetic iron oxide particles, using magnetic resonance microscopy. Size amplification due to sequestration of the superparamagnetic particles in vacuoles enhances contrast in localized areas in high-resolution magnetic resonance imaging. Magnetic resonance images of samples containing differing concentrations of T cells embedded in 3% gelatin show a number of dark regions due to the superparamagnetic iron oxide particles, consistent with the number predicted by transmission electron microscopy. Colabeling of T cell samples with a fluorescent dye leads to strong correlations between magnetic resonance and fluorescence microscopic images, showing the presence of the superparamagnetic iron oxide particles at the cell site. This result lays the foundation for our approach to tracking the movement of a specific cell type in live animals and humans.

The contribution of chemical exchange to MRI frequency shifts in brain tissue.

Recent high‐field MRI studies based on resonance frequency contrast have revealed brain structure with unprecedented detail. Although subtle magnetic susceptibility variations caused by iron and myelin seem to be important to this contrast, recent research on protein solutions suggests that chemical exchange between water and macromolecular protons may contribute substantially to the observed gray‐white matter frequency contrast. To investigate this, we performed spectroscopic MRI experiments at 14 T on samples of fixed human visual cortex and fresh pig brain. To allow direct observation of any exchange‐induced frequency shifts, these samples were soaked in reference chemicals (TSP and dioxane) that are assumed not to be involved in exchange. For both fresh and fixed tissues and with both reference chemicals, substantial negative exchange‐induced gray‐white matter frequency contrast (–6.3 to –13.5 ppb) was found, whereas intracortical contrast was negligible. The sign of the gray‐white matter exchange‐induced frequency difference was opposite to the overall gray‐white matter frequency difference observed in vivo. This suggests that exchange contributes to, but is not sufficient to explain, the frequency contrast in vivo and tissue susceptibility differences may have a greater contribution than previously thought. The exchange‐dependent contribution may report on tissue chemical composition and pH. Magn Reson Med, 2010.

Characterization of T2* Heterogeneity in Human Brain White Matter

Recent in vivo MRI studies at 7.0 T have demonstrated extensive heterogeneity of T2* relaxation in white matter of the human brain. In order to study the origin of this heterogeneity, we performed T2* measurements at 1.5, 3.0, and 7.0 T in normal volunteers. Formalin‐fixed brain tissue specimens were also studied using T2*‐weighted MRI, histologic staining, chemical analysis, and electron microscopy. We found that T2* relaxation rate (R2* = 1/T2*) in white matter in living human brain is linearly dependent on the main magnetic field strength, and the T2* heterogeneity in white matter observed at 7.0 T can also be detected, albeit more weakly, at 1.5 and 3.0 T. The T2* heterogeneity exists also in white matter of the formalin‐fixed brain tissue specimens, with prominent differences between the major fiber bundles such as the cingulum (CG) and the superior corona radiata. The white matter specimen with substantial difference in T2* has no significant difference in the total iron content, as determined by chemical analysis. On the other hand, evidence from histologic staining and electron microscopy demonstrates these tissue specimens have apparent difference in myelin content and microstructure. Magn Reson Med, 2009.

MRI of the basement membrane using charged nanoparticles as contrast agents

The integrity of the basement membrane is essential for tissue cellular growth and is often altered in disease. In this work a method for noninvasively detecting the structural integrity of the basement membrane, based on the delivery of cationic iron‐oxide nanoparticles, was developed. Cationic particles accumulate due to the highly negative charge of proteoglycans in the basement membrane. The kidney was used to test this technique because of its highly fenestrated endothelia and well‐established disease models to manipulate the basement membrane charge barrier. After systemic injection of cationic or native ferritin (CF or NF) in rats, ex vivo and in vivo MRI showed selective accumulation of CF, but not NF, causing a 60% reduction in signal intensity in cortex at the location of individual glomeruli. Immunofluorescence and electron microscopy demonstrated that this CF accumulation was localized to the glomerular basement membrane (GBM). In a model of GBM breakdown during focal and segmental glomerulosclerosis, MRI showed reduced single glomerular accumulation of CF, but a diffuse accumulation of CF in the kidney tubules caused by leakage of CF through the glomerulus. Cationic contrast agents can be used to target the basement membrane and detect the breakdown of the basement membrane in disease. Magn Reson Med 60:564–574, 2008.

Direct imaging of macrovascular and microvascular contributions to BOLD fMRI in layers IV-V of the rat whisker-barrel cortex

The spatiotemporal characteristics of the hemodynamic response to increased neural activity were investigated at the level of individual intracortical vessels using BOLD-fMRI in a well-established rodent model of somatosensory stimulation at 11.7 T. Functional maps of the rat barrel cortex were obtained at 150 × 150 × 500 μm spatial resolution every 200 ms. The high spatial resolution allowed separation of active voxels into those containing intracortical macro vessels, mainly vein/venules (referred to as macrovasculature), and those enriched with arteries/capillaries and small venules (referred to as microvasculature) since the macro vessel can be readily mapped due to the fast T2 decay of blood at 11.7 T. The earliest BOLD response was observed within layers IV-V by 0.8s following stimulation and encompassed mainly the voxels containing the microvasculature and some confined macrovasculature voxels. By 1.2s, the BOLD signal propagated to the macrovasculature voxels where the peak BOLD signal was 2-3 times higher than that of the microvasculature voxels. The BOLD response propagated in individual venules/veins far from neuronal sources at later times. This was also observed in layers IV-V of the barrel cortex after specific stimulation of separated whisker rows. These results directly visualized that the earliest hemodynamic changes to increased neural activity occur mainly in the microvasculature and spread toward the macrovasculature. However, at peak response, the BOLD signal is dominated by penetrating venules even at layers IV-V of the cortex.

Deciphering laminar-specific neural inputs with line-scanning fMRI

Using a line-scanning method during functional magnetic resonance imaging (fMRI), we obtained high temporal (50-ms) and spatial (50-μm) resolution information along the cortical thickness and showed that the laminar position of fMRI onset coincides with distinct neural inputs in rat somatosensory and motor cortices. This laminar-specific fMRI onset allowed us to identify the neural inputs underlying ipsilateral fMRI activation in the barrel cortex due to peripheral denervation-induced plasticity.

Thalamocortical Inputs Show Post-Critical Period Plasticity

Experience-dependent plasticity in the adult brain has clinical potential for functional rehabilitation following central and peripheral nerve injuries. Here, plasticity induced by unilateral infraorbital (IO) nerve resection in 4-week-old rats was mapped using MRI and synaptic mechanisms were elucidated by slice electrophysiology. Functional MRI demonstrates a cortical potentiation compared to thalamus 2 weeks after IO nerve resection. Tracing thalamocortical (TC) projections with manganese-enhanced MRI revealed circuit changes in the spared layer 4 (L4) barrel cortex. Brain slice electrophysiology revealed TC input strengthening onto L4 stellate cells due to an increase in postsynaptic strength and the number of functional synapses. This work shows that the TC input is a site for robust plasticity after the end of the previously defined critical period for this input. Thus, TC inputs may represent a major site for adult plasticity in contrast to the consensus that adult plasticity mainly occurs at cortico-cortical connections.

The Use of Silica Coated MnO Nanoparticles to Control MRI Relaxivity in Response to Specific Physiological Changes

MnO nanoparticles have been tested to engineer a delayed increase in MRI T(1) relaxivity caused by cellular uptake via endocytosis into acidic compartments. Various coatings on core-shell structured MnO nanoparticles were tested for those that had the lowest T(1) relaxivity at pH 7.4, a pH where MnO does not dissolve into Mn(2+) ions. The rate of dissolution and release of Mn(2+) of the different coated MnO particles as well as changes in T(1) relaxivity were measured at pH 5, a pH routinely obtained in the endosomal-lysosomal pathway. Of a number of coatings, silica coated MnO (MnO@SiO(2)) had the lowest relaxivity at pH 7.4 (0.29 mm(-1) sec(-1)). About one third of the MnO dissolved within 20 min and the T(1) relaxivity increased to that of free Mn(2+) (6.10 mm(-1) sec(-1)) after three days at pH 5. MRI of MnO@SiO(2) particles injected into the rat brain showed time-dependent signal changes consistent with the in vitro rates. Thalamocortical tract-tracing could be observed due to the released Mn(2+). Intravenous infusion of MnO@SiO(2) particles showed little enhancement in any tissue except gallbladder. The gallbladder enhancement was interpreted to be due to endocytosis by liver cells and excretion of Mn(2+) ions into the gallbladder. The MnO@SiO(2) core-shell nanoparticles show the best potential for delaying the release of MRI contrast until endocytosis into low pH compartments activate MRI contrast. The delayed enhancement may have benefits for targeting MRI contrast to specific cells and surface receptors that are known to be recycled by endocytosis.

The fabrication of uniform cylindrical nanoshells and their use as spectrally tunable MRI contrast agents

A new form of tunable magnetic resonance imaging agent based on precisely dimensioned cylindrical magnetic nanoshells is introduced. Using top-down prepatterned substrates, the nanoshells are fabricated by exploiting what is usually regarded as a detrimental processing side-effect, namely the redeposition of material back-sputtered during ion-milling. The well-resolved nuclear magnetic resonance peaks of the resulting nanostructures attest to the nanoscale fabrication control and the general feasibility of such sputter redeposition for fabrication of a variety of self-supporting, highly monodisperse nanoscale structures.

3D mapping of somatotopic reorganization with small animal functional MRI

There are few in vivo noninvasive methods to study neuroplasticity in animal brains. Functional MRI (fMRI) has been developed for animal brain mapping, but few fMRI studies have analyzed functional alteration due to plasticity in animal models. One major limitation is that fMRI maps are characterized by statistical parametric mapping making the apparent boundary dependent on the statistical threshold used. Here, we developed a method to characterize the location of center-of-mass in fMRI maps that is shown not to be sensitive to statistical threshold. Utilizing centers-of-mass as anchor points to fit the spatial distribution of the BOLD response enabled quantitative group analysis of altered boundaries of functional somatosensory maps. This approach was used to study cortical reorganization in the rat primary somatosensory cortex (S1) after sensory deprivation to the barrel cortex by follicle ablation (F.A.). FMRI demonstrated an enlarged nose S1 representation in the 3D somatotopic functional maps. This result clearly demonstrates that fMRI enables the spatial mapping of functional changes that can characterize multiple regions of S1 cortex and still be sensitive to changes due to plasticity.