Afonso C. Silva

In vivo detection of neuroarchitecture in the rodent brain using manganese-enhanced MRI

Visualizing brain anatomy in vivo could provide insight into normal and pathophysiology. Here it is demonstrated that neuroarchitecture can be detected in the rodent brain using MRI after systemic MnCl2. Administration of MnCl2 leads to rapid T1 enhancement in the choroid plexus and circumventricular organs, which spreads to the CSF space in ventricles and periventricular tissue. After 1 day, there was MRI enhancement throughout the brain with high intensity in the pituitary, olfactory bulb, cortex, basal forebrain, hippocampus, basal ganglia, hypothalamus, amygdala, and cerebellum. Contrast obtained enabled visualization of specific features of neuroarchitecture. The arrowhead structure of the dentate gyrus as well as the CA1-CA3 region of the hippocampus and layers in cortex, cerebellum, as well as the olfactory bulb could be readily observed. Preliminary assignments of olfactory bulb layers, cortical layers in frontal and somatosensory cortex, and cerebellum were made. Systemic MnCl2 leads to MRI visualization of neuroarchitecture nondestructively.

Laminar specificity of functional MRI onset times during somatosensory stimulation in rat

The blood oxygenation level-dependent (BOLD) response to somatosensory stimulation was measured in α-chloralose-anesthetized rats. BOLD fMRI was obtained at 40-ms temporal resolution and spatial resolution of 200 × 200 × 2,000 μm3 by using a gated activation paradigm in an 11.7 T MRI. Results show a consistent heterogeneity of fMRI onset times and amplitudes. The earliest onset time (0.59 ± 0.17 s, n = 9) corresponded anatomically to layer IV, with superficial and deeper layers starting significantly later (1.27 ± 0.43 s in layers I–III, and 1.11 ± 0.45 s in layer VI). The amplitude of BOLD signal changes also varied with the cortical depth from the pial surface. Changes in the supragranular layers (8.3%) were 44% bigger than changes in the intermediate layers (5.5%), located only ≈700 μm below, and 144% larger than the bottom layer (3.5%), located ≈1.4 mm below the pial surface. The data presented demonstrate that BOLD signal changes have distinct amplitude and temporal characteristics, which vary spatially across cortical layers.

Sensitivity of MRI resonance frequency to the orientation of brain tissue microstructure

Recent advances in high-field (≥7 T) MRI have made it possible to study the fine structure of the human brain at the level of fiber bundles and cortical layers. In particular, techniques aimed at detecting MRI resonance frequency shifts originating from local variation in magnetic susceptibility and other sources have greatly improved the visualization of these structures. A recent theoretical study [He X, Yablonskiy DA (2009) Proc Natl Acad Sci USA 106:13558–13563] suggests that MRI resonance frequency may report not only on tissue composition, but also on microscopic compartmentalization of susceptibility inclusions and their orientation relative to the magnetic field. The proposed sensitivity to tissue structure may greatly expand the information available with conventional MRI techniques. To investigate this possibility, we studied postmortem tissue samples from human corpus callosum with an experimental design that allowed separation of microstructural effects from confounding macrostructural effects. The results show that MRI resonance frequency does depend on microstructural orientation. Furthermore, the spatial distribution of the resonance frequency shift suggests an origin related to anisotropic susceptibility effects rather than microscopic compartmentalization. This anisotropy, which has been shown to depend on molecular ordering, may provide valuable information about tissue molecular structure.

Early temporal characteristics of cerebral blood flow and deoxyhemoglobin changes during somatosensory stimulation.

The close correspondence between neural activity in the brain and cerebral blood flow (CBF) forms the basis for modern functional neuroimaging methods. Yet, the temporal characteristics of hemodynamic changes induced by neuronal activity are not well understood. Recent optical imaging observations of the time course of deoxyhemoglobin (HbR) and oxyhemoglobin have suggested that increases in oxygen consumption after neuronal activation occur earlier and are more spatially localized than the delayed and more diffuse CBF response. Deoxyhemoglobin can be detected by blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI). In the present study, the temporal characteristics of CBF and BOLD changes elicited by somatosensory stimulation in rat were investigated by high-field (9.4 T) MRI. With use of high-temporal-resolution fMRI, it was found that the onset time of the CBF response in the somatosensory cortex was 0.6 ± 0.4 seconds (n = 10). The CBF changes occurred significantly earlier than changes in HbR concentration, which responded after 1.1 ± 0.3 seconds. Furthermore, no early increases in HbR (early negative BOLD signal changes) were observed. These findings argue against the occurrence of an early loss of hemoglobin oxygenation that precedes the rise in CBF and suggest that CBF and oxygen consumption increases may be dynamically coupled in this animal model of neural activation.

Functional Reactivity of Cerebral Capillaries

The spatiotemporal evolution of cerebral microcirculatory adjustments to functional brain stimulation is the fundamental determinant of the functional specificity of hemodynamically weighted neuroimaging signals. Very little data, however, exist on the functional reactivity of capillaries, the vessels most proximal to the activated neuronal population. Here, we used two-photon laser scanning microscopy, in combination with intracranial electrophysiology and intravital video microscopy, to explore the changes in cortical hemodynamics, at the level of individual capillaries, in response to steady-state forepaw stimulation in an anesthetized rodent model. Overall, the microcirculatory response to functional stimulation was characterized by a pronounced decrease in vascular transit times (20% ± 8%), a dilatation of the capillary bed (10.9% ± 1.2%), and significant increases in red blood cell speed (33.0% ± 7.7%) and flux (19.5% ± 6.2%). Capillaries dilated more than the medium-caliber vessels, indicating a decreased heterogeneity in vessel volumes and increased blood flow-carrying capacity during neuronal activation relative to baseline. Capillary dilatation accounted for an estimated ˜18% of the total change in the focal cerebral blood volume. In support of a capacity for focal redistribution of microvascular flow and volume, significant, though less frequent, local stimulation-induced decreases in capillary volume and erythrocyte speed and flux also occurred. The present findings provide further evidence of a strong functional reactivity of cerebral capillaries and underscore the importance of changes in the capillary geometry in the hemodynamic response to neuronal activation.

Manganese-enhanced magnetic resonance imaging of mouse brain after systemic administration of MnCl2: dose-dependent and temporal evolution of T1 contrast.

Manganese is a useful contrast agent for MRI of animals. Previously, it has been shown that systemic doses of MnCl2 provide unique contrast in the rodent brain, enabling visualization of neuroarchitecture. The present work investigates the dose and temporal dependence of brain enhancement after i.v. administration of MnCl2. Varying doses of MnCl2 (9–175 mg/kg) were administered to mice from 0 to 24 h prior to T1‐weighted manganese‐enhanced MRI (MEMRI) at 11.7 T. Pre‐MnCl2 T1 values measured in different brain regions ranged from 1.17 ± 0.03 to 1.76 ± 0.01 s. Post‐MnCl2 T1 measured 24 hr after administration of MnCl2 were significantly decreased, even after the lowest dose of MnCl2. The largest decreases occurred in the pituitary gland, where post‐MnCl2 T1 ranged from 231 ± 23 ms following the lowest dose to 143 ± 43 ms after the highest dose, while the smallest decreases were observed in cortex (post‐MnCl2 T1 = 1060 ± 5 ms for low dose and 637 ± 5 ms for high dose). The contrast resulting after 14 hr did not change up to 24 hr. Enhancement first occurred in subarachnoid spaces, followed by ventricles and periventricular tissues, and finally reached the remainder of the brain. Cortical layers were detected at higher doses (>88 mg/kg) and olfactory bulb layers were detected with the lowest dose (9 mg/kg). Temporal evolution of the enhancement of the olfactory bulb layers was observed. In some regions of the brain, such as hippocampus and thalamus, the changes in contrast detected between 2 and 14 hr used very specific pathways. These results demonstrate that both the dose and the time after MnCl2 can be manipulated to optimize brain contrast in a region‐specific manner. Published 2005 Wiley‐Liss, Inc.

Manganese-Enhanced MRI: An Exceptional Tool in Translational Neuroimaging

The metal manganese is a potent magnetic resonance imaging (MRI) contrast agent that is essential in cell biology. Manganese-enhanced magnetic resonance imaging (MEMRI) is providing unique information in an ever-growing number of applications aimed at understanding the anatomy, the integration, and the function of neural circuits both in normal brain physiology as well as in translational models of brain disease. A major drawback to the use of manganese as a contrast agent, however, is its cellular toxicity. Therefore, paramount to the successful application of MEMRI is the ability to deliver Mn2+ to the site of interest using as low a dose as possible while preserving detectability by MRI. In the present work, the different approaches to MEMRI in translational neuroimaging are reviewed and challenges for future identified from a practical standpoint.

Superficial white matter fiber systems impede detection of long-range cortical connections in diffusion MR tractography

Significance It is widely recognized that studying the detailed anatomy of the human brain is of great importance for neuroscience and medicine. The principal means for achieving this goal is presently diffusion magnetic resonance imaging (dMRI) tractography, which uses the local diffusion of water throughout the brain to estimate the course of long-range anatomical projections. Such projections connect gray matter regions through axons that travel in the deep white matter. The present study combines dMRI tractography with histological analysis to investigate where in the brain this method succeeds and fails. We conclude that certain superficial white matter systems pose challenges for measuring cortical connections that must be overcome for accurate determination of detailed neuroanatomy in humans. In vivo tractography based on diffusion magnetic resonance imaging (dMRI) has opened new doors to study structure–function relationships in the human brain. Initially developed to map the trajectory of major white matter tracts, dMRI is used increasingly to infer long-range anatomical connections of the cortex. Because axonal projections originate and terminate in the gray matter but travel mainly through the deep white matter, the success of tractography hinges on the capacity to follow fibers across this transition. Here we demonstrate that the complex arrangement of white matter fibers residing just under the cortical sheet poses severe challenges for long-range tractography over roughly half of the brain. We investigate this issue by comparing dMRI from very-high-resolution ex vivo macaque brain specimens with histological analysis of the same tissue. Using probabilistic tracking from pure gray and white matter seeds, we found that ∼50% of the cortical surface was effectively inaccessible for long-range diffusion tracking because of dense white matter zones just beneath the infragranular layers of the cortex. Analysis of the corresponding myelin-stained sections revealed that these zones colocalized with dense and uniform sheets of axons running mostly parallel to the cortical surface, most often in sulcal regions but also in many gyral crowns. Tracer injection into the sulcal cortex demonstrated that at least some axonal fibers pass directly through these fiber systems. Current and future high-resolution dMRI studies of the human brain will need to develop methods to overcome the challenges posed by superficial white matter systems to determine long-range anatomical connections accurately.

The contribution of myelin to magnetic susceptibility-weighted contrasts in high-field MRI of the brain.

T(2)*-weighted gradient-echo MRI images at high field (≥ 7T) have shown rich image contrast within and between brain regions. The source for these contrast variations has been primarily attributed to tissue magnetic susceptibility differences. In this study, the contribution of myelin to both T(2)* and frequency contrasts is investigated using a mouse model of demyelination based on a cuprizone diet. The demyelinated brains showed significantly increased T(2)* in white matter and a substantial reduction in gray-white matter frequency contrast, suggesting that myelin is a primary source for these contrasts. Comparison of in-vivo and in-vitro data showed that, although tissue T(2)* values were reduced by formalin fixation, gray-white matter frequency contrast was relatively unaffected and fixation had a negligible effect on cuprizone-induced changes in T(2)* and frequency contrasts.

Dynamic activity-induced manganese-dependent contrast magnetic resonance imaging (DAIM MRI)

Activity‐induced manganese‐dependent contrast (AIM) MRI is a hemodynamic‐independent functional MRI method that used manganese ion as an MR‐detectable contrast agent. In AIM, MnCl2 is infused intra‐arterially after the blood‐brain barrier (BBB) is opened with a hyperosmolar agent. Upon functional stimulation of the brain, Mn2+ accumulates in the active region(s) by entering active cells through voltage‐gated Ca2+ channels, causing local signal increases in T1‐weighted images. The contrast of AIM MRI depends strongly on the depth of anesthesia, and the low levels used in somatosensory stimulation studies can lead to significant nonspecific accumulation of manganese ion throughout the brain. The purpose of this study was to produce an AIM functional map of somatosensory stimulation, which separates the stimulation‐specific signal increase from the nonspecific activation due to light anesthesia. A dynamic AIM (DAIM) paradigm was developed, which used sequential MR scans during MnCl2 infusion, prior to and following functional stimulation of the brain. Stimulation‐specific functional maps were produced using time‐course analysis. The new method was tested during glutamate administration and electric stimulation of the rat forepaw. It was shown that DAIM maps are better confined to the specific region of brain activated by somatosensory stimulation as compared to AIM MRI. Magn Reson Med 48:927–933, 2002.