This work describes a dynamic magnetic resonance imaging (MRI) technique for local scanning of the human body with use of a handheld receive coil or coil array. Real-time MRI is based on highly undersampled radial gradient-echo sequences with joint reconstructions of serial images and coil sensitivity maps by regularized nonlinear inversion (NLINV). For this proof-of-concept study, a fixed slice position and field-of-view (FOV) were predefined from the operating console, while a local receive coil (array) is moved across the body-for the sake of simplicity by the subject itself. Experimental realizations with a conventional 3 T magnet comprise dynamic anatomic imaging of the head, thorax and abdomen of healthy volunteers. Typically, the image resolution was 0.75 to 1.5 mm with 3 to 6 mm section thickness and acquisition times of 33 to 100 ms per frame. However, spatiotemporal resolutions and contrasts are highly variable and may be adjusted to clinical needs. In summary, the proposed FLASHlight MRI method provides a robust acquisition and reconstruction basis for future diagnostic strategies that mimic the usage of ultrasound. Necessary extensions for this vision require remote control of all sequence parameters by a person at the scanner as well as the design of more flexible gradients and magnets.

BACKGROUND:Pharmaceutical intervention in the CNS is hampered by the shielding function of the blood-brain barrier (BBB). To induce clinical anesthesia, general anesthetics such as isoflurane readily penetrate the BBB. Here, we investigated whether isoflurane can be utilized for therapeutic drug delivery. METHODS:Barrier function in primary endothelial cells was evaluated by transepithelial/transendothelial electrical resistance, and nanoscale STED and SRRF microscopy. In mice, BBB permeability was quantified by extravasation of several fluorescent tracers. Mouse models including the GL261 glioma model were evaluated by MRI, immunohistochemistry, electron microscopy, western blot, and expression analysis. RESULTS:Isoflurane enhances BBB permeability in a time- and concentration-dependent manner. We demonstrate that, mechanistically, isoflurane disturbs the organization of membrane lipid nanodomains and triggers caveolar transport in brain endothelial cells. BBB tightness re-establishes directly after termination of anesthesia, providing a defined window for drug delivery. In a therapeutic glioblastoma trial in mice, simultaneous exposure to isoflurane and cytotoxic agent improves efficacy of chemotherapy. CONCLUSIONS:Combination therapy, involving isoflurane-mediated BBB permeation with drug administration has far-reaching therapeutic implications for CNS malignancies.

OBJECTIVES:The purpose of this work was to improve the clinical versatility of high-speed real-time MRI studies of temporomandibular joint (TMJ) dynamics by simultaneous recordings of multiple MRI movies in different sections.METHODS:Real-time MRI at 3 T was realized using highly undersampled radial FLASH acquisitions and image reconstruction by regularized nonlinear inversion (NLINV). Multi-slice real-time MRI of two, three or four slices at 0.75 mm resolution and 6 to 8 mm thickness was accomplished at 50.0 ms, 33.3 ms or 25.5 ms temporal resolution, respectively, yielding simultaneous movies at 2 × 10, 3 × 10 or 4 × 10 frames per second in a frame-interleaved acquisition mode. Real-time MRI movies were evaluated by three blinded raters for visibility of the anterior and posterior border of disc, shape of the disk body and condyle head as well as movement of the disc and condyle (1 = excellent, 5 = no visibility).RESULTS:Effective delineation of the disk atop the mandibular condyle was achieved by T1-weighted images with opposed-phase water-fat contrast. Compared to 8 mm sections, multi-slice recordings with 6 mm thickness provided sharper delineation of relevant structures as confirmed by inter-rater evaluation. Respective dual-slice and triple-slice recordings of a single TMJ as well as dual-slice recordings of both joints (one slice per TMJ) received the highest visibility ratings of ≤ 2 corresponding to high confidence in diagnostic content.CONCLUSIONS:The improved access to TMJ dynamics by multi-slice real-time MRI will contribute to more effective treatment of temporomandibular disorders.

BACKGROUND Optimal control of minimally invasive interventions by hyperthermia requires dynamic temperature mapping at high temporal resolution. METHODS Based on the temperature-dependent shift of the proton resonance frequency (PRF), this work developed a method for real-time MRI thermometry which relies on highly undersampled radial FLASH MRI sequences with iterative image reconstruction by regularized nonlinear inversion (NLINV). As a first step, the method was validated with use of a temperature phantom and ex vivo organs (swine kidney) subjected to heating by warm water or a pulsed laser source. RESULTS The temperature maps obtained by real-time PRF MRI demonstrate good accuracy as independently controlled by fiber-optic temperature sensors. Moreover, the dynamic results demonstrate both excellent sensitivity to single laser pulses (20 ms duration, 6 J energy output) and high temporal resolution, i.e., 200 ms acquisition times per temperature map corresponding to a rate of 5 frames per second. In addition, future extensions to in vivo applications were prepared by addressing the breathing-related motion problem by a pre-recorded library of reference images representative of all respiratory states. CONCLUSIONS The proposed method for real-time MRI thermometry now warrants further developments towards in vivo MRI monitoring of thermal interventions in animals.

This article provides an overview of in vivo magnetic resonance (MR) imaging contrasts obtained for mammalian brain in relation to histological knowledge. Emphasis is paid to the (1) significance of high spatial resolution for the optimization of T1, T2, and magnetization transfer contrast, (2) use of exogenous extra- and intracellular contrast agents for validating endogenous contrast sources, and (3) histological structures and biochemical compounds underlying these contrasts and (4) their relevance to neuroradiology. Comparisons between MR imaging at subnanoliter resolution and histological data indicate that (a) myelin sheaths, (b) nerve cells, and (c) the neuropil are most responsible for observed MR imaging contrasts, while (a) diamagnetic macromolecules, (b) intracellular paramagnetic ions, and (c) extracellular free water, respectively, emerge as the dominant factors. Enhanced relaxation rates due to paramagnetic ions, such as iron and manganese, have been observed for oligodendrocytes, astrocytes, microglia, and blood cells in the brain as well as for nerve cells. Taken together, a plethora of observations suggests that the delineation of specific structures in high-resolution MR imaging of mammalian brain and the absence of corresponding contrasts in MR imaging of the human brain do not necessarily indicate differences between species but may be explained by partial volume effects. Second, paramagnetic ions are required in active cells in vivo which may reduce the magnetization transfer ratio in the brain through accelerated T1 recovery. Third, reductions of the magnetization transfer ratio may be more sensitive to a particular pathological condition, such as astrocytosis, microglial activation, inflammation, and demyelination, than changes in relaxation. This is because the simultaneous occurrence of increased paramagnetic ions (i.e., shorter relaxation times) and increased free water (i.e., longer relaxation times) may cancel T1 or T2 effects, whereas both processes reduce the magnetization transfer ratio.

Using proton MRS and MRI of mouse brain at 9.4T, this work provides the first in vivo evidence of pH-dependent concurrent changes of three amide signals and related metabolic responses to hypercapnia and hypothermia. During hypercapnia, amide proton MRS signals of glutamine at 6.8-6.9ppm and 7.6ppm as well as of unspecific compounds at 8.1-8.3ppm increase by at least 50% both at 37°C and 22°C. These changes reflect a reduced proton exchange with water. They are strongly correlated with intracellular pH which ranges from 6.75±0.10 to 7.13±0.06 as determined from a shift in creatine phosphokinase equilibrium. In MRI, saturation transfer from aliphatic as well as aromatic and/or amide protons alters slightly during hypercapnia and significantly during hypothermia. The asymmetry in magnetization transfer ratios decreased slightly during hypercapnia and hypothermia. Regardless of pH or temperature, saturation transfer from aliphatic protons between -2 and -4ppm frequency offset to water protons is significantly greater than that from aromatic/amide protons at corresponding offsets between +2 and +4ppm. Irradiation of aliphatic compounds at -3.5ppm frequency offset from water predominantly saturates lipids and water associated with myelin. Taken together, the results indicate that, for the B1 power used in this study, dipolar coupling between aliphatic and water protons rather than proton exchange is the dominant factor in Z-spectra and magnetization transfer ratio asymmetry of the brain in vivo.

Using T 1 -weighted MRI at two different magnetic field strengths, the enhanced longitudinal relaxivity due to paramagnetic manganese ions in mouse brain in vivo is shown to reflect reduced intracellular mobility. One day after systemic administration of manganese chloride, increases of the longitudinal relaxation rate ∆ R 1 in several brain regions are significantly higher at 2.35 T than at 9.4 T. The corresponding relaxivity ratios 100 r 1 / 400 r 1 = 100 ∆ R 1 / 400 ∆ R 1 range from 2.4 (striatum) to 4.4 (cerebellar cortex). In contrast, the ∆ R 1 values after intraventricular administration of gadolinium-DTPA (Gd-DTPA) are not significantly different between both field strengths yielding 100 r 1 / 400 r 1 ratios from 1.0 to 1.1. The same observation holds true for manganese and Gd-DTPA relaxivities in aqueous solution. The pronounced field strength dependence of manganese relaxivities indicates a reduced mobility of manganese ions in vivo by confinement to a viscous fluid compartment and/or due to macromolecular binding. Moreover, preferential enhancement of nerve cell assemblies by manganese ions and the observation of additional contrast enhancement by magnetization transfer suggest an intracellular localization of manganese. This is further supported by a slow release of manganese from nerve cells postmortem , which occurs despite a high permeability of damaged cellular membranes as demonstrated by a rapid uptake of extracellular Gd-DTPA.

Contrast-enhanced T₁- and T₂-weighted MRI at 9.4 T and in-plane resolutions of 25 and 30 µm has been demonstrated to differentiate between neural tissues in mouse brain in vivo, including granule cell layers, principal cell layers, general neuropil, specialized neuropil and white matter. In T₁-weighted MRI of the olfactory bulb, hippocampus and cerebellum, contrast obtained by the intracranial administration of gadopentetate dimeglumine (Gd-DTPA) reflects the extra- and intracellular spaces of gray matter in agreement with histological data. General neuropil areas are highlighted, whereas other tissues present with lower signal intensities. The induced contrast is similar to that in plain T₂-weighted MRI, but offers a 16-30-fold higher contrast-to-noise ratio. Systemic administration of manganese chloride increases the signal-to-noise ratio in T₁-weighted MRI to a significantly greater extent in principal cell layers and specialized neuropil than in granule cell layers, whereas gadolinium-enhanced MRI indicates no larger intracellular spaces in these tissues. Granule cell layers are enhanced no more than general neuropil by manganese, whereas gadolinium-enhanced MRI indicates significantly larger intracellular spaces in the cell layers. These discrepancies suggest that the signal increase after manganese administration reflects cellular activity which is disproportionate to the intracellular space. As a result, principal cell layers and specialized neuropil become highlighted, whereas granule cell layers, general neuropil and white matter present with lower signal intensities.

This work compares magnetization transfer (MT) MRI of living mice with contrast-enhanced MT MRI using intraventricular administration of gadopentetate dimeglumine (Gd-DTPA), systemic administration of MnCl2, and both. In MT MRI at 9.4T, the contrast-to-noise ratio (CNR) between white matter (WM) and gray matter (GM) increased by 85% after Gd-DTPA injection into the lateral ventricle. When applied in conjunction with manganese-enhanced MT MRI (117μm isotropic resolution, 6min measuring time), Gd-DTPA boosted the CNR increase from +56% to +117%. Additional T1 measurements at 2.35T revealed that intraventricular Gd-DTPA shortens the T1 of GM much more than that of WM, which corresponds to estimated extracellular spaces of 26% in GM and only 15% in WM. These results explain the additional MT contrast enhancement by Gd-DTPA and demonstrate that the T1 shortening by intracellular Mn2+ is well complemented by extracellular Gd-DTPA. The data suggest a high myelin and low water content to hinder access of hydrophilic paramagnetic agents, so that the resulting differential accumulation effectively reduces the MT saturation in water-rich tissues and thereby facilitates the mapping of myelin-rich tissues. Finally, a 156% CNR increase between GM and WM for contrast-enhanced MT MRI at 9.4T using both Gd-DTPA and manganese allowed for 60μm isotropic resolution (102min measuring time), which delineated myelinated fibers and layers even within GM areas such as the thalamus and cerebellar cortex. Improved MT contrasts were also seen in the cervical spinal cord.

Halogenated volatile anesthetics (HVA) are widely used in medicine and research but their effects on brain metabolism in intact organisms are still largely unknown. Here, localized proton magnetic resonance spectroscopy (MRS) of anesthetized mice was applied to evaluate HVA effects on cerebral metabolites in vivo. Experimental protocols combined different concentrations of isoflurane, halothane, sevoflurane, and desflurane with known modulators of adrenergic, GABAergic, and glutamatergic neurotransmission. As a most striking finding, brain lactate increased in individual mice from 1.0 ± 0.6 mM (awake state) to 6.2 ± 1.5 mM (1.75% isoflurane). In addition, relative to total creatine, there were significant isoflurane-induced increases of alanine by 111%, GABA by 20%, choline-containing compounds by 20%, and myo-inositol by 10% which were accompanied by significant decreases of glucose by 51% and phosphocreatine by 9%. The elevation of lactate was most pronounced in the striatum. The HVA effects correlated with the respective minimal alveolar concentrations and were mostly reversible within minutes. The observed alterations are best explained by an HVA-induced stimulation of adrenergic pathways in conjunction with an inhibition of the respiratory chain. Apart from casting new light on cerebral energy metabolism, the present results challenge brain studies of HVA-anesthetized animals.

Introduction: General anesthesia using isoflurane is a most common procedure in medicine and research. A series of possible mechanisms of isoflurane action has been described, but it is largely unknown how brain metabolism and signaling is changed in the intact organism by exposure to isoflurane. Such knowledge may be important to reduce adverse effects and to correctly interpret results from animal experiments conducted under such anesthesia. We performed localized proton MR spectroscopy in mice with and without exposure to isoflurane. To further understand metabolic changes observed, exposure to isoflurane was combined with known modulators of the adrenergic system.

Horizontal T2-weighted images of adult C57/bl6 mice revealed a 5-layer structure of the cerebral cortex (Figure A and B, yellow bars). This layering partly correlates with the local cellular density as shown by a corresponding cresyl violet staining of a 10 months old C57/bl6 mouse. In contrast, young adult mice (Figure D) present with a small layer of higher signal intensity inside the two darker layers (Figure D, arrows), which presumably reflects a not yet fully completed myelination. Pax6cKO mice (Figure E) are characterized by an almost complete loss of cortical layers II, III, and IV and an ectopic superficial location of an enlarged set of pyramidal neurons. These histologically proven alterations nicely correspond to the observed 3-layer structure by MRI (Figure E, yellow bars). Concluding remarks High-resolution MRI with pronounced T2 contrast allows for the detection of cortical layer-like structures in mouse brain in vivo, which resemble the histologically defined 6-layer structure of the mammalian cortex. For the first time developmental changes in the cortical architecture of healthy mice as well as layer alterations in Pax6cKO mice could be visualized by in vivo MRI. Most likely the observed contrast depicts both cell density and myelin content. Although it cannot be expected that MRI contrasts completely match to the histological mapping of the cortex, the present findings further enhance the scientific potential of in vivo MRI techniques for studying experimental animals.

Objectives: Volatile anesthetics like isoflurane are widely used in animal research and human medicine. An isoflurane-induced increase in lactate (Lac) was reported from cell culture studies [1]. On the other hand, isoflurane has been discussed as a neuroprotective agent in stroke [2]. Here, we evaluated the influence of isoflurane on brain Lac concentrations in comparison to intravenous anesthetic drugs using localized proton MRS of healthy mice.

OBJECTIVE:The aim of this study was to investigate the role of voltage-dependent calcium channels (VDCCs) in axon degeneration during autoimmune optic neuritis.METHODS:Calcium ion (Ca(2+)) influx into the optic nerve (ON) through VDCCs was investigated in a rat model of optic neuritis using manganese-enhanced magnetic resonance imaging and in vivo calcium imaging. After having identified the most relevant channel subtype (N-type VDCCs), we correlated immunohistochemistry of channel expression with ON histopathology. In the confirmatory part of this work, we performed a treatment study using omega-conotoxin GVIA, an N-type specific blocker.RESULTS:We observed that pathological Ca(2+) influx into ONs during optic neuritis is mediated via N-type VDCCs. By analyzing the expression of VDCCs in the inflamed ONs, we detected an upregulation of alpha(1B), the pore-forming subunit of N-type VDCCs, in demyelinated axons. However, high expression levels were also found on macrophages/activated microglia, and lower levels were detected on astrocytes. The relevance of N-type VDCCs for inflammation-induced axonal degeneration and the severity of optic neuritis was corroborated by treatment with omega-conotoxin GVIA. This blocker led to decreased axon and myelin degeneration in the ONs together with a reduced number of macrophages/activated microglia. These protective effects were confirmed by analyzing the spinal cords of the same animals.INTERPRETATION:We conclude that N-type VDCCs play an important role in inflammation-induced axon degeneration via two mechanisms: First, they directly mediate toxic Ca(2+) influx into the axons; and second, they contribute to macrophage/microglia function, thereby promoting secondary axonal damage. Ann Neurol 2009;66:81-93.

The impact of developmental ablation of Pax6 function on morphology and functional connectivity of the adult cerebrum was studied in cortex-specific Pax6 knockout mice (Pax6cKO) using structural magnetic resonance imaging (MRI), manganese-enhanced MRI, and diffusion tensor MRI in conjunction with fiber tractography. Mutants presented with decreased volumes of total brain and olfactory bulb, reduced cortical thickness, and altered layering of the piriform cortex. Tracking of major neuronal fiber bundles revealed a disorganization of callosal fibers with an almost complete lack of interhemispheric connectivity. In Pax6cKO mice intrahemispheric callosal fibers as well as intracortical fibers were predominantly directed along a rostrocaudal orientation instead of a left-right and dorsoventral orientation found in controls. Fiber disorganization also involved the septohippocampal connection targeting mostly the lateral septal nucleus. The hippocampus was rostrally extended and its volume was increased relative to that of the forebrain and midbrain. Manganese-induced MRI signal enhancement in the CA3 region suggested a normal function of hippocampal pyramidal cells. Noteworthy, several morphologic disturbances in gray and white matter of Pax6cKO mice were similar to observations in human aniridia patients. The present findings indicate an important role of Pax6 in the development of both the cortex and cerebral fiber connectivity.