This work is part of the Mouse Imaging Centre (MICe) at the Hospital for Sick Children and the University of Toronto.

1. Multiple-mouse in vivo Brain Imaging:
When imaging live animals, several key features must be present throughout the imaging session: 1) a safe method of anesthesia, 2) environmental control and 3) physiological monitoring. Furthermore, when imaging multiple subjects simultaneously, there are added complexities regarding the ease and speed of preparation and the reproducibility of animal positions to facilitate image registration. Consequently, three major custom components were designed and fabricated: a loading system to insert the mice into RF coils within the MRI, an induction chamber to facilitate preparation and a platform with embedded monitoring leads to standardize position.
The Loading System:
The mouse loading system consists of two major parts: the 'mouse hive' and the 'loading array'. The mouse hive's main function is to position seven Millipede RF coils (Varian NMR Systems, Palo Alto, CA) in a hexagonal array inside the magnet bore. The loading array is designed to hold and transport multiple mice housed in 50 milliliter centrifuge tubes with holes drilled through their tips to allow entry of anesthetic gas. After the mice are anesthetized and interfaced to the monitoring equipment at a preparation area in the vicinity of the magnet, they are inserted into the modified centrifuge tubes and mounted onto the loading array. After mounting all mice, the loading array is transported and inserted into the magnet and positioned on a rail system. The rail system allows the array to couple with the mouse hive when pushed down the bore of the magnet. When fully inserted into the magnet, the centrifuge tubes dock onto the anesthetic delivery system within the RF coils. Isoflurane mixed with oxygen is supplied from the mouse hive end to the specimen through a tube along the axis of each individual coil. This anesthetic gas mixture flows into the tubes, past the mice and is collected by an active scavenging unit attached to the back of the loading array (Figure 1).
The Induction Chamber:
Since imaging times can take up to three hours, minimizing animal preparation time is crucial to limit the mouse's exposure to anesthesia. Hence, we have developed a custom induction chamber to streamline the preparation process (Figure 2). The custom induction chamber creates a single environment for both induction and handling of multiple mice. Constructed from clear acrylic, the induction chamber features selfclosing silicone iris ports to minimize anesthetic leakage and allows the user to access the internal environment without the need for special gloves. Compared to conventional mask and circuits for a single mouse, the induction chamber is large enough to house up to twenty mice and allows for free manipulation of the mice without the attachment of cumbersome tubes and masks. The unit is supplied with a constant flow of anesthetic gas which is collected using a passive scavenging system. Resistive heating elements are used to heat the floor of the chamber to maintain the animals' body temperature during preparation.
The Sled:
One of the most awkward and time-consuming aspects of preparing mice for the MRI are the application of electrocardiograph (ECG) electrodes and rectal temperature probes. In addition, many of the conventional electrodes, such as cuff and needle electrodes, were found to distort the animal's posture making it difficult to standardize positioning. Therefore, we devised a custom form-fitted positioning platform with embedded ECG, respiration and temperature probes called the 'sled' (U.S. Patent 7,146,936) (Figure 3). Motion restraints made from Velcro fasteners were used to limit movement of the head.
Multiple-mouse in vivo brain imaging steps:
<ol>
<li>All mouse research requires local ACC (Animal Care Committee), IACUC (Institutional Animal Care and Use Committee) or equivalent approval for mouse handling procedures.</li>
<li>All procedures such as identification and weighing of animals should be performed under a Biological Safety Cabinet (BSC) and in the MRI Unit. The animals are transferred into an autoclavable plastic container and transported to the MRI induction chamber.</li>
<li>Mice are anesthetized in a prewarmed induction chamber using 4% isoflurane and 4 L/min of oxygen. Animals are fully anesthetized once they fail to respond to paw pinch. Fur from the chest is removed using a hair remover (NAIR) if necessary to provide better contact with ECG and temperature monitoring devices that have been built into a custom sled. Eve salve (Tears Naturale P.M.) is applied to the eyes to prevent drying, and approximately 0.3 mL of saline is administered subcutaneously to maintain hydration.</li>
<li>Gd-DTPA-BMA (Omniscan) may be used if contrast enhancement is desired. If Gd-DTPA is to be used, it will be diluted in saline (final volume 300uL) and administered in a single dose of 1 mmol/kg prior to the MR session via IP.</li>
<li>Mice are then loaded into individual sleds, immobilized with head straps and slide into open-ended 50mL conical tube (Figure 3). Up to 7 live mice can be scanned at one time. Once all animals are loaded with physiological monitoring connected, set the isoflurane level in the magnet bore to 2% and the oxygen level to 8 L/min.</li>
<li>The conical tubes are mounted into a docking system (Figure 1) designed to position mice uniformly into each RF coil positioned in the centre of the magnet bore. Once loaded, isoflurane can be reduced to 0.9-2%. ECG and temperature are monitored on each animal throughout the course of the scan. Animals are kept warm during the course of the scan with warmed air.</li>
<li>The duration of each three dimensional scan is approximately 3 hours. The details are as follows: fast spin echo with TR of 2300 ms and a TEeff of 36 ms. Echo train length of 8 with 1 average. Resulting image resolution is 125 &mu;m (Figure 4).</li>
<li>When the scan is complete, the animals are removed from the magnet and unloaded in a warm induction chamber filled with 100% oxygen. The animals are transferred into a plastic sealed containers and transported under a BSC. They are placed on a warm draft free cage and allowed to recover from anesthesia.</li>
</ol>
2. Multiple-mouse ex vivo Brain Imaging:
Unaffected by motion artifacts, fixed MR imaging achieves higher resolution than live imaging. High-resolution, Three-dimensional datasets provide maximal flexibility in extracting quantitative information and allow for automated image analysis. A custom RF coil array was developed for parallel acquisition of 16 high-resolution MR datasets of fixed in-skull mouse brains in overnight scanning sessions.
The 16-Coil ex vivo Brain Imaging Array:
A custom-built 16-coil solenoid array was created to image 16 samples concurrently. This design improves upon an earlier prototype used to image three samples concurrently in a 60mm insert gradient set. The 8-turn solenoid coils are over wound at the ends to provide uniform sensitivity to within 10% over a length of 26mm and are individually shielded within modular compartments (Idziak and Haeberlen, 1982). The 16 coil compartments are assembled to a frame (Figure 5), which positions the coils within the gradient and clamps it in place using a pneumatic bladder to minimize motion.
Multiple-mouse ex vivo brain imaging steps:
<ol>
<li>Open chest cavity and insert needle (or Safety Winged infusion set, 25G X 3&#8260;4) into the left ventricle of the heart of an anesthetized mouse (via intraperitoneal injection of ketamine (150 mg/kg) and xylazine (10 mg/kg). Cut the right auricle.</li>
<li>Transcardiac perfusion flush with 30 mL room temp 1 X PBS + 1 &mu;L/ mL heparin (1000 USP units/ mL) + 2 mM ProHance at a flow rate of approximately 100 mL/hr.</li>
<li>Fixation pass with 30 mL 4% PFA (room temperature) + 2 mM ProHance at 100 mL/hr.</li>
<li>Decapitate and remove skin, lower jaw, ears, cartilaginous nose tip.</li>
<li>Place remaining skull structure in 4% PFA + 2 mM ProHance overnight at 4&deg;C.</li>
<li>Transfer to 1X PBS + 0.02% sodium azide + 2 mM ProHance. </li>
<li>High resolution three dimensional MRI scanning at 7 Tesla occurs between 4 days and not longer than 2.5 months after perfusion. Brains are placed in a 16-channel solenoid coil array (Figure 5).</li>
<li>Imaging parameters are as follows: fast spin echo with TR 325 ms and a TEeff of 30 ms, echo train length of 6 with 4 averages. The final images have an isotropic resolution of 32 &mu;m (Figure 6) and the scan duration is approximately 12 hours.</li>
<li>After scanning, place the skull in 10% Formalin + 2 mM ProHance for preservation.</li>
</ol>

3. Representative Results:
<img src="/files/ftp_upload/2497/2497fig1.jpg" alt="Figure 1" /> 
Figure 1. The mouse loading system. The "loading array" and "mouse hive" connected with a common fiberglass rail system.
<img src="/files/ftp_upload/2497/2497fig2.jpg" alt="Figure 2" /> 
Figure 2. The induction chamber.
<img src="/files/ftp_upload/2497/2497fig3.jpg" alt="Figure 3" /> 
Figure 3. a) The sled, showing embedded monitoring sensors and head restraint. b) An anesthetized mouse on a sled with the head restraint attached. The sled assembly easily slides into the centrifuge tube.
<img src="/files/ftp_upload/2497/2497fig4.jpg" alt="Figure 4" /> 
Figure 4. Representative in vivo multiple brain images from one 3 hour three dimensional scan.
<img src="/files/ftp_upload/2497/2497fig5.jpg" alt="Figure 5" /> 
Figure 5. 16-channel solenoid coil array for scanning of 16 fixed brain specimens.
<img src="/files/ftp_upload/2497/2497fig6.jpg" alt="Figure 6" /> 
Figure 6. Representative ex-vivo brain image.

<ol>
	<li>Bock, N. A., Konyer, N. B., Henkelman, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple-mouse+MRI.">Multiple-mouse MRI.</a> Magn. Reson. Med. 49, 158-167 (2003).</li><li>Dazai, J., Bock, N. A., Nieman, B. J., Davidson, L. M., Henkelman, R. M., Chen, X. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple+mouse+biological+loading+and+monitoring+system+for+MRI.">Multiple mouse biological loading and monitoring system for MRI.</a> Magn. Reson. Med. 52, 709-715 (2004).</li><li>Spring, S., Lerch, J. P., Henkelman, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sexual+dimorphism+revealed+in+the+structure+of+the+mouse+brain+using+three-dimensional+magnetic+resonance+imaging.">Sexual dimorphism revealed in the structure of the mouse brain using three-dimensional magnetic resonance imaging.</a> NeuroImage. 35, 1424-1433 (2007).</li><li>Idziak, S., Haeberlen, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+and+construction+of+a+high+homogeneity+rf+coil+for+solid-state+multiple-pulse+NMR.">Design and construction of a high homogeneity rf coil for solid-state multiple-pulse NMR.</a> J. Magn. Reson. 50, 281-288 (1982).</li></ol>

No conflicts of interest declared.

Both the in vivo and ex vivo mouse imaging systems image multiple subjects at once to increase throughput of imaging studies considerably without increasing the imaging time. The brain images from both the in vivo and ex vivo multiple-mouse imaging techniques are of high quality and are suitable for phenotyping major and minor structures in the mouse brain respectively.
To minimize animal preparation time of multiple specimens, parallelization of processes is of utmost importance. For example, the development of the induction chamber allowed for the induction of multiple specimens simultaneously, and the sled has synchronized the application of the ECG and temperature probes while standardizing body positioning. Furthermore, our ex vivo imaging system enables us to acquire high resolution three dimensional images of 16 fixed whole brains at one time which is ideal for high-throughput phenotyping studies.
Possible limitations of our in vivo imaging system include the inability to individually control anesthetic and temperature for each mouse. If required individual anesthetic control can be implemented with the addition of exclusive anesthetic vaporizers for each mouse. Another limitation of the in vivo imaging system is that scanning is restricted to mice that are less than approximately 32 grams. However, there is a plan currently to increase coil size to accommodate larger animals.

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Isoflurane, USP (AErrane)</td>
<td>&nbsp;</td>
<td>Baxter</td>
<td>CA2L9108</td>
<td>&nbsp;</td>
<tr>
<td>NAIR</td>
<td>&nbsp;</td>
<td>Church &amp; Dwight Canada Corp.</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Tears Naturale P.M.</td>
<td>&nbsp;</td>
<td>Alcon</td>
<td>DIN 02082519</td>
<td>&nbsp;</td>
<tr>
<td>Omniscan (gadodiamide injection USP) </td>
<td>&nbsp;</td>
<td>GE healthcare</td>
<td>J-110A</td>
<td>&nbsp;</td>
<tr>
<td>Custom Sleds </td>
<td>&nbsp;</td>
<td>Dazai Research Instruments</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>PBS w/o Ca and Mg </td>
<td>&nbsp;</td>
<td>Wisent</td>
<td>311-010-CL</td>
<td>&nbsp;</td>
<tr>
<td>Heparin 10000USP/10ml </td>
<td>&nbsp;</td>
<td>Pharmaceutical Partners of Canada</td>
<td>DIN 02264315</td>
<td>&nbsp;</td>
<tr>
<td>ProHance (gadoteridol injection USP)</td>
<td>&nbsp;</td>
<td>Bracco Diagnostics</td>
<td>11181 </td>
<td>&nbsp;</td>
<tr>
<td>Parafolmadehyde (powder)</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>P-6148-500g</td>
<td>&nbsp;</td>
<tr>
<td>Sodium Azide</td>
<td>&nbsp;</td>
<td>Fisher</td>
<td>S227-100</td>
<td>&nbsp;</td>
<tr>
<td>Formalin 10%</td>
<td>&nbsp;</td>
<td>Fisher </td>
<td>SF100-4</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

multiple-mouse neuroanatomical magnetic resonance imaging

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and evaluating mouse models of human disease. MRI is an excellent modality for investigating genetically altered animals. It is capable of whole brain coverage, can be used in vivo, and provides multiple contrast mechanisms for investigating different aspects of neuranatomy and physiology. The advent of high-field scanners along with the ability to scan multiple mice simultaneously allows for rapid phenotyping of novel mutations.
Effective mouse MRI studies require attention to many aspects of experiment design. In this article, we will describe general methods to acquire quality images for mouse phenotyping using a system that images mice concurrently in shielded transmit/receive radio frequency (RF) coils in a common magnet (Bock et al., 2003). We focus particularly on anatomical phenotyping, an important and accessible application that has shown a high potential for impact in many mouse models at our imaging centre. Before we can provide the detailed steps to acquire such images, there are important practical considerations for both in vivo brain imaging (Dazai et al., 2004) and ex vivo brain imaging (Spring et al., 2007) that should be noted. These are discussed below.

Watch this Scientific Journal Video about Multiple-mouse Neuroanatomical Magnetic Resonance Imaging at JoVE.com

Multiple-mouse Neuroanatomical Magnetic Resonance Imaging

Magnetic resonance imaging (MRI) has become an increasingly popular tool for examining the phenotype of genetically altered mice. This article illustrates the methods necessary to achieve high-throughput phenotyping of genetically altered mice using multiple-mouse MRI.

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and ...

multiple-mouse-neuroanatomical-magnetic-resonance-imaging

Research

JoVE Journal

Neuroscience

15.7K Views.  Hospital for Sick Children. Risonanza magnetica (MRI) è diventato uno strumento sempre più popolare per l'esame del fenotipo di topi geneticamente modificati. Questo articolo illustra i metodi necessari per raggiungere high-throughput fenotipizzazione di topi geneticamente modificati utilizzando più-mouse risonanza magnetica.

Video: Multiple-mouse Neuroanatomical Magnetic Resonance Imaging

Magnetic Resonance Spectroscopy of live Drosophila melanogaster using Magic Angle Spinning

High-Resolution Magic Angle Spinning (HRMAS) proton magnetic resonance spectroscopy (1H-MRS) is a novel non-destructive technique that improves spectral line-widths and allows high-resolution spectra to be obtained from extracts, intact cells, cell cultures, and more importantly intact tissue to investigate relationships between metabolites and cellular processes. In vivo HRMAS 1H-MRS studies have yet to be reported in the live fruit fly Drosophila melanogaster. Drosophila, as a simpler genetic organism, allows the multiple biological functions and various evolutionarily conserved signaling pathways to be examined at the whole organism level and it is a useful model for investigating genetics and physiology. To this end, we developed and implemented an in vivo HRMAS 1H-MRS method to investigate live Drosophila at 14.1 T. Here, we outline an HRMAS 1H-MRS protocol for the molecular characterization of Drosophila with a conventional MR spectrometer equipped with an HRMAS probe. This technique is a novel, in vivo, non-destructive Drosophila metabolite measurement approach, which enables the identification of disease biomarkers and thus may contribute to novel therapeutic development.

Magnetic Resonance Spectroscopy of live Drosophila melanogaster using Magic Angle Spinning

This technique enables the use of high-resolution magic angle spinning proton MR spectroscopy (HRMAS 1H-MRS) for molecular characterization of live Drosophila melanogaster with a conventional 14.1 tesla spectrometer equipped with an HRMAS probe.

High-Resolution Magic Angle Spinning (HRMAS) proton magnetic resonance spectroscopy (1H-MRS) is a novel non-destructive technique that improves spectral ...

High-resolution Functional Magnetic Resonance Imaging Methods for Human Midbrain

Functional MRI (fMRI) is a widely used tool for non-invasively measuring correlates of human brain activity. However, its use has mostly been focused upon measuring activity on the surface of cerebral cortex rather than in subcortical regions such as midbrain and brainstem. Subcortical fMRI must overcome two challenges: spatial resolution and physiological noise. Here we describe an optimized set of techniques developed to perform high-resolution fMRI in human SC, a structure on the dorsal surface of the midbrain; the methods can also be used to image other brainstem and subcortical structures.
High-resolution (1.2 mm voxels) fMRI of the SC requires a non-conventional approach. The desired spatial sampling is obtained using a multi-shot (interleaved) spiral acquisition1. Since, T2* of SC tissue is longer than in cortex, a correspondingly longer echo time (TE ~ 40 msec) is used to maximize functional contrast. To cover the full extent of the SC, 8-10 slices are obtained. For each session a structural anatomy with the same slice prescription as the fMRI is also obtained, which is used to align the functional data to a high-resolution reference volume.
In a separate session, for each subject, we create a high-resolution (0.7 mm sampling) reference volume using a T1-weighted sequence that gives good tissue contrast. In the reference volume, the midbrain region is segmented using the ITK-SNAP software application2. This segmentation is used to create a 3D surface representation of the midbrain that is both smooth and accurate3. The surface vertices and normals are used to create a map of depth from the midbrain surface within the tissue4.
Functional data is transformed into the coordinate system of the segmented reference volume. Depth associations of the voxels enable the averaging of fMRI time series data within specified depth ranges to improve signal quality. Data is rendered on the 3D surface for visualization.
In our lab we use this technique for measuring topographic maps of visual stimulation and covert and overt visual attention within the SC1. As an example, we demonstrate the topographic representation of polar angle to visual stimulation in SC.

This article describes techniques to perform high-resolution functional magnetic resonance imaging with 1.2 mm sampling in human midbrain and subcortical structures using a 3T scanner. Use of these techniques to resolve topographic maps of visual stimulation in the human superior colliculus (SC) is given as an example.

Functional MRI (fMRI) is a widely used tool for non-invasively measuring correlates of human brain activity. However, its use has mostly been focused upon ...

The Use of Magnetic Resonance Spectroscopy as a Tool for the Measurement of Bi-hemispheric Transcranial Electric Stimulation Effects on Primary Motor Cortex Metabolism

Transcranial direct current stimulation (tDCS) is a neuromodulation technique that has been increasingly used over the past decade in the treatment of neurological and psychiatric disorders such as stroke and depression. Yet, the mechanisms underlying its ability to modulate brain excitability to improve clinical symptoms remains poorly understood 33. To help improve this understanding, proton magnetic resonance spectroscopy (1H-MRS) can be used as it allows the in vivo quantification of brain metabolites such as &#947;-aminobutyric acid (GABA) and glutamate in a region-specific manner 41. In fact, a recent study demonstrated that 1H-MRS is indeed a powerful means to better understand the effects of tDCS on neurotransmitter concentration 34. This article aims to describe the complete protocol for combining tDCS (NeuroConn MR compatible stimulator) with 1H-MRS at 3 T using a MEGA-PRESS sequence. We will describe the impact of a protocol that has shown great promise for the treatment of motor dysfunctions after stroke, which consists of bilateral stimulation of primary motor cortices 27,30,31. Methodological factors to consider and possible modifications to the protocol are also discussed.

This article aims to describe a basic protocol for combining transcranial direct current stimulation (tDCS) with proton magnetic resonance spectroscopy (1H-MRS) measurements to investigate the effects of bilateral stimulation on primary motor cortex metabolism.

Transcranial direct current stimulation (tDCS) is a neuromodulation technique that has been increasingly used over the past decade in the treatment of ...

High-resolution In Vivo Manual Segmentation Protocol for Human Hippocampal Subfields Using 3T Magnetic Resonance Imaging

The human hippocampus has been broadly studied in the context of memory and normal brain function and its role in different neuropsychiatric disorders has been heavily studied. While many imaging studies treat the hippocampus as a single unitary neuroanatomical structure, it is, in fact, composed of several subfields that have a complex three-dimensional geometry. As such, it is known that these subfields perform specialized functions and are differentially affected through the course of different disease states. Magnetic resonance (MR) imaging can be used as a powerful tool to interrogate the morphology of the hippocampus and its subfields. Many groups use advanced imaging software and hardware (&#62;3T) to image the subfields; however this type of technology may not be readily available in most research and clinical imaging centers. To address this need, this manuscript provides a detailed step-by-step protocol for segmenting the full anterior-posterior length of the hippocampus and its subfields: cornu ammonis (CA) 1, CA2/CA3, CA4/dentate gyrus (DG), strata radiatum/lacunosum/moleculare (SR/SL/SM), and subiculum. This protocol has been applied to five subjects (3F, 2M; age 29-57, avg. 37). Protocol reliability is assessed by resegmenting either the right or left hippocampus of each subject and computing the overlap using the Dice&#39;s kappa metric. Mean Dice&#39;s kappa (range) across the five subjects are: whole hippocampus, 0.91 (0.90-0.92); CA1, 0.78 (0.77-0.79); CA2/CA3, 0.64 (0.56-0.73); CA4/dentate gyrus, 0.83 (0.81-0.85); strata radiatum/lacunosum/moleculare, 0.71 (0.68-0.73); and subiculum 0.75 (0.72-0.78). The segmentation protocol presented here provides other laboratories with a reliable method to study the hippocampus and hippocampal subfields in vivo using commonly available MR tools.

High-resolution In Vivo Manual Segmentation Protocol for Human Hippocampal Subfields Using 3T Magnetic Resonance Imaging

The goal of this manuscript is to study the hippocampus and hippocampal subfields using MRI. The manuscript describes a protocol for segmenting the hippocampus and five hippocampal substructures: cornu ammonis (CA) 1, CA2/CA3, CA4/dentate gyrus, strata radiatum/lacunosum/moleculare, and subiculum.

The human hippocampus has been broadly studied in the context of memory and normal brain function and its role in different neuropsychiatric disorders has ...

High-resolution Structural Magnetic Resonance Imaging of the Human Subcortex In Vivo and Postmortem

The focus of this study was to test the resolution limits of structural MRI of a postmortem brain compared to living human brains. The resolution of structural MRI in vivo is ultimately limited by physiological noise, including pulsation, respiration and head movement. Although imaging hardware continues to improve, it is still difficult to resolve structures on the millimeter scale. For example, the primary visual sensory pathways synapse at the lateral geniculate nucleus (LGN), a visual relay and control nucleus in the thalamus that normally is organized into six interleaved monocular layers. Neuroimaging studies have not been able to reliably distinguish these layers due their small size that are less than 1 mm thick.
The resolving limit of structural MRI, in a postmortem brain was tested using multiple images averaged over a long duration (~24 h). The purpose was to test whether it was possible to resolve the individual layers of the LGN in the absence of physiological noise. A proton density (PD)1 weighted pulse sequence was used with varying resolution and other parameters to determine the minimum number of images necessary to be registered and averaged to reliably distinguish the LGN and other subcortical regions. The results were also compared to images acquired in living human brains. In vivo subjects were scanned in order to determine the additional effects of physiological noise on the minimum number of PD scans needed to differentiate subcortical structures, useful in clinical applications.

High-resolution Structural Magnetic Resonance Imaging of the Human Subcortex In Vivo and Postmortem

Here we present a protocol to determine the minimum number images that needed to be registered and averaged to resolve subcortical structures and test whether the individual layers of the LGN could be resolved in the absence of physiological noise.

The focus of this study was to test the resolution limits of structural MRI of a postmortem brain compared to living human brains. The resolution of ...

A Magnetic Resonance Imaging Protocol for Stroke Onset Time Estimation in Permanent Cerebral Ischemia

MRI provides a sensitive and specific imaging tool to detect acute ischemic stroke by means of a reduced diffusion coefficient of brain water. In a rat model of ischemic stroke, differences in quantitative T1 and T2 MRI relaxation times (qT1 and qT2) between the ischemic lesion (delineated by low diffusion) and the contralateral non-ischemic hemisphere increase with time from stroke onset. The time dependency of MRI relaxation time differences is heuristically described by a linear function and thus provides a simple estimate of stroke onset time. Additionally, the volumes of abnormal qT1 and qT2 within the ischemic lesion increase linearly with time providing a complementary method for stroke timing. A (semi)automated computer routine based on the quantified diffusion coefficient is presented to delineate acute ischemic stroke tissue in rat ischemia. This routine also determines hemispheric differences in qT1 and qT2 relaxation times and the location and volume of abnormal qT1 and qT2 voxels within the lesion. Uncertainties associated with onset time estimates of qT1 and qT2 MRI data vary from &plusmn; 25 min to &plusmn; 47 min for the first 5 hours of stroke. The most accurate onset time estimates can be obtained by quantifying the volume of overlapping abnormal qT1 and qT2 lesion volumes, termed 'Voverlap' (&plusmn; 25 min) or by quantifying hemispheric differences in qT2 relaxation times only (&plusmn; 28 min). Overall, qT2 derived parameters outperform those from qT1. The current MRI protocol is tested in the hyperacute phase of a permanent focal ischemia model, which may not be applicable to transient focal brain ischemia.

A protocol for stroke onset time estimation in a rat model of stroke exploiting quantitative magnetic resonance imaging (qMRI) parameters is described. The procedure exploits diffusion MRI for delineation of the acute stroke lesion and quantitative T1 and T2 (qT1 and qT2) relaxation times for timing of stroke.

MRI provides a sensitive and specific imaging tool to detect acute ischemic stroke by means of a reduced diffusion coefficient of brain water. In a rat ...

Diffusion Tensor Magnetic Resonance Imaging in Chronic Spinal Cord Compression

Chronic spinal cord compression is the most common cause of spinal cord impairment in patients with nontraumatic spinal cord damage. Conventional magnetic resonance imaging (MRI) plays an important role in both confirming the diagnosis and evaluating the degree of compression. However, the anatomical detail provided by conventional MRI is not sufficient to accurately estimate neuronal damage and/or assess the possibility of neuronal recovery in chronic spinal cord compression patients. In contrast, diffusion tensor imaging (DTI) can provide quantitative results according to the detection of water molecule diffusion in tissues. In the present study, we develop a methodological framework to illustrate the application of DTI in chronic spinal cord compression disease. DTI fractional anisotropy (FA), apparent diffusion coefficients (ADCs), and eigenvector values are useful for visualizing microstructural pathological changes in the spinal cord. Decreased FA and increases in ADCs and eigenvector values were observed in chronic spinal cord compression patients compared to healthy controls. DTI could help surgeons understand spinal cord injury severity and provide important information regarding prognosis and neural functional recovery. In conclusion, this protocol provides a sensitive, detailed, and noninvasive tool to evaluate spinal cord compression.

Here, we present a protocol for the application of diffusion tensor imaging parameters to evaluate spinal cord compression.

Chronic spinal cord compression is the most common cause of spinal cord impairment in patients with nontraumatic spinal cord damage. Conventional magnetic ...

Standardized Data Acquisition for Neuromelanin-Sensitive Magnetic Resonance Imaging of the Substantia Nigra

The dopaminergic system plays a crucial role in healthy cognition (e.g., reward learning and uncertainty) and neuropsychiatric disorders (e.g., Parkinson's disease and schizophrenia). Neuromelanin is a byproduct of dopamine synthesis that accumulates in dopaminergic neurons of the substantia nigra. Neuromelanin-sensitive magnetic resonance imaging (NM-MRI) is a noninvasive method for measuring neuromelanin in those dopaminergic neurons, providing a direct measure of dopaminergic cell loss in the substantia nigra and a proxy measure of dopamine function. Although NM-MRI has been shown to be useful for studying various neuropsychiatric disorders, it is challenged by a limited field-of-view in the inferior-superior direction resulting in the potential loss of data from the accidental exclusion of part of the substantia nigra. In addition, the field is lacking a standardized protocol for the acquisition of NM-MRI data, a critical step in facilitating large-scale multisite studies and translation into the clinic. This protocol describes a step-by-step NM-MRI volume placement procedure and online quality control checks to ensure the acquisition of good-quality data covering the entire substantia nigra.

This protocol shows how to acquire neuromelanin-sensitive magnetic resonance imaging data of the substantia nigra.

The dopaminergic system plays a crucial role in healthy cognition (e.g., reward learning and uncertainty) and neuropsychiatric disorders (e.g., ...

Acquisition of Resting-State Functional Magnetic Resonance Imaging Data in the Rat

Resting-state functional magnetic resonance imaging (rs-fMRI) has become an increasingly popular method to study brain function in a resting, non-task state. This protocol describes a preclinical survival method for obtaining rs-fMRI data. Combining low dose isoflurane with continuous infusion of the &#945;2 adrenergic receptor agonist dexmedetomidine provides a robust option for stable, high-quality data acquisition while preserving brain network function. Furthermore, this procedure allows for spontaneous breathing and near-normal physiology in the rat. Additional imaging sequences can be combined with resting-state acquisition creating experimental protocols with anesthetic stability of up to 5 h using this method. This protocol describes the setup of equipment, monitoring of rat physiology during four distinct phases of anesthesia, acquisition of resting-state scans, quality assessment of data, recovery of the animal, and a brief discussion of post-processing data analysis. This protocol can be used across a wide variety of preclinical rodent models to help reveal the resulting brain network changes that occur at rest.

This protocol describes a method for obtaining stable resting-state functional magnetic resonance imaging (rs-fMRI) data from a rat using low dose isoflurane in combination with low dose dexmedetomidine.

Resting-state functional magnetic resonance imaging (rs-fMRI) has become an increasingly popular method to study brain function in a resting, non-task ...

Endovascular Perforation Model for Subarachnoid Hemorrhage Combined with Magnetic Resonance Imaging (MRI)

The endovascular filament perforation model to mimic subarachnoid hemorrhage (SAH) is a commonly used model - however, the technique can cause a high mortality rate as well as an uncontrollable volume of SAH and other intracranial complications such as stroke or intracranial hemorrhage. In this protocol, a standardized SAH mouse model is presented, induced by endovascular filament perforation, combined with magnetic resonance imaging (MRI) 24 h after operation to ensure the correct bleeding site and exclude other relevant intracranial pathologies. Briefly, C57BL/6J mice are anesthetized with an intraperitoneal ketamine/xylazine (70 mg/16 mg/kg body weight) injection and placed in a supine position. After midline neck incision, the common carotid artery (CCA) and carotid bifurcation are exposed, and a 5-0 non-absorbable monofilament polypropylene suture is inserted in a retrograde fashion into the external carotid artery (ECA) and advanced into the common carotid artery. Then, the filament is invaginated into the internal carotid artery (ICA) and pushed forward to perforate the anterior cerebral artery (ACA). After recovery from surgery, mice undergo a 7.0 T MRI 24 h later. The volume of bleeding can be quantified and graded via postoperative MRI, enabling a robust experimental SAH group with the option to perform further subgroup analyses based on blood quantity.

Endovascular Perforation Model for Subarachnoid Hemorrhage Combined with Magnetic Resonance Imaging MRI

Here we present a standardized SAH mouse model, induced by endovascular filament perforation, combined with magnetic resonance imaging (MRI) 24 h after operation to ensure the correct bleeding site and exclude other relevant intracranial pathologies.

The endovascular filament perforation model to mimic subarachnoid hemorrhage (SAH) is a commonly used model - however, the technique can cause a high ...