This work was supported by the National Institute of Neurological Disorders and Stroke (R01NS104116; R01NS134944; Principal Investigator: Christos Papadelis).

Integrating MEG and HD-EEG in Pediatric Epilepsy Localization

The authors report no disclosures.

In this study, we illustrate the experimental setup to record simultaneous MEG and HD-EEG in children with DRE while resting/sleeping, performing a task, or receiving stimuli, and propose a methodological framework for localizing the irritative zone, SOZ, and eloquent brain areas using EMSI, as well as individual MSI and ESI. We further provide technical recommendations for merging MEG and HD-EEG data from different commercially available products that present unique features. We present data from three cases to enhance the clinical utility of EMSI in the localization of epileptogenic and eloquent brain areas. The findings here indicate that EMSI results outperform those obtained by either modality alone, most likely due to the additive value of the complementary properties of MEG and EEG signals in the combined solution and possibly due to the increased number of sensors used for recording the data (&#62;550 sensors). Particularly, EMSI noninvasively localized the irritative and SOZs with concordant findings as ESI on iEEG gold standard, which confirmed the clinical observations.
The proposed methodology includes the following critical steps: (i) high-quality acquisition of simultaneous MEG and HD-EEG (i.e., high SNR) recordings with high spatial sampling of sensors (&#62;550 sensors) covering the entire brain of interictal and ictal activities, as well as visual-, motor-, auditory-, and somatosensory-evoked fields and potentials, from children with DRE (steps 3.1-3.2); (ii) temporal synchronization and spatial co-registration of MEG and HD-EEG signals recorded with different acquisition systems (step 3.12); (iii) careful preprocessing and selection of data portions containing interictal activity (steps 4.1.1-4.1.7), ictal onset activity (steps 4.2.1-4.2.7), and event-related responses (steps 4.3.1-4.3.6), respectively; and (iv) accurate source localization of the irritative zone, SOZ, and eloquent brain areas of interest using reliable source localization methods (e.g., ECDs with clustering and dSPM) (steps 4.1.8-4.1.9, 4.2.8-4.2.9, and 4.3.7-4.3.9, respectively).
The most critical step when performing simultaneous MEG and HD-EEG recordings is to spatially (alignment between coordinate spaces) and temporally (correction of the linear clock drift) synchronize the data recorded by the two acquisition systems. Such synchronization is crucial to ensure the correct identification of interictal, ictal, and visual/motor/auditory/tactile events that simultaneously occur in the MEG and HD-EEG signals. Errors in the timepoint selection of these events may affect the source localization results and identify areas of the brain that are not necessarily involved in the generation of these events.
MEG systems often offer compatible 32-, 64-, and 128-channel EEG systems incorporated into the product for performing simultaneous MEG and EEG measurements. In these cases, there is no need to temporally synchronize the data by sending common trigger signals. Similarly, most EEG systems are nowadays compatible with all MEG systems. Despite these advances in hardware, only few epilepsy centers perform simultaneous MEG and HD-EEG recordings as part of the presurgical evaluation. Here, we took advantage of such integrability and combined the 306-channel MEG and 256-channel EEG systems to simultaneously record the brain activity with &#62;550 sensors covering the subject&#39;s head. So far, few software for advanced analysis of MEG, HD-EEG, and iEEG data (e.g., Brainstorm, CURRY, EEGLab, FieldTrip, MNE, or NUTMEG) are available. Future studies are therefore necessary to validate the proposed methodology with new neuroimaging analysis software. Finally, the combination of MSI and ESI into a unique solution (EMSI) increased the computational complexity of the data analysis.
The described method presents a few limitations that should be addressed in future studies. We manually selected IEDs occurring on both MEG and HD-EEG data of two representative patients while ignoring interictal spikes that occurred in only one of the two signals (either MEG or EEG). Manual selection of spikes can be a time-consuming and subjective approach that can be simplified using automated approaches for detecting IEDs developed during the last decades57,58,59. However, visual inspection is always recommended for careful analysis and refined detection of each IED. Furthermore, we used the SOZ as an approximator of the EZ. Yet, the SOZ does not always predict surgical outcomes60,61,62,63. Future studies can, therefore, use the surgical outcome as ground truth for more precise delineation of the EZ13,14,15,16,17,19,20. Although seizures can be successfully captured using simultaneous MEG and EEG and localized using appropriate source localization techniques44,64, it is relatively rare to record such ictal events in clinical practice, especially from outpatients on ASMs. This is mostly due to the limited duration of MEG recordings and the excessive body movements occurring during seizures (e.g., the patient&#39;s head slipped out the dewar), which may cause biological artifacts that can severely affect source localization findings. In a recent review, Stefan et al. reported the occurrence of seizures during MEG recordings in 7%-24% of patients, with an average recording time of 30 minutes up to 5.7 h across different studies65.&#160;At CCMC, 18 out of 89 (20.2%) patients had ictal events captured during simultaneous MEG and HD-EEG recordings performed within the past ~2 years. However, only 8 out of the 18 (44.4%) patients were successfully analyzed. In cases where interictal MEG recordings show normal or inconclusive findings, ictal MEG or HD-EEG may be used to localize the EZ with high precision. Yet, technical and logistical requirements for these recordings should be addressed. In addition, the representative data for the eloquent cortex localization via EMSI were not compared with any gold standards for localization of these functional brain areas, such as noninvasive fMRI or intraoperative electrocortical stimulation. Further investigations may, therefore, integrate EMSI and fMRI towards a multimodal noninvasive imaging tool to improve the localization accuracy of these eloquent brain areas in children with DRE. This work may also be extended to localize other functional brain areas, such as the language-eloquent regions. Localization of language functions is of critical importance during the presurgical evaluation of patients with DRE to determine their surgical candidacy, plan the extent of surgical resection, and prevent permanent postoperative functional deficits66. Several noninvasive studies have shown that language mapping using MEG can provide concordant results, similar to the invasive Wada test, which is often regarded as the gold standard for identifying the dominant language hemisphere67,68,69,70. A recent study has proposed a multimodal approach in which the combination of different techniques (i.e., cortical stimulation mapping, high-gamma electrocorticography, fMRI, and transcranial magnetic stimulation) can provide mutual, confirmatory, and complementary information for presurgical language mapping71. Despite these advantages, mapping language areas is still challenging in pediatric patients who have cognitive, intellectual, and language barriers due to their age. Thus, more age-specific tasks and child-friendly setups should be developed in the near future. In this work, we analyzed MEG and HD-EEG data using software that is not certified for clinical purposes. Although these tools have been proven to be valuable and effective, they carry liability issues that should be considered when reporting presurgical evaluation findings for clinical use. Here, we describe procedures for HD-EEG recordings using only sponge-based EEG electrode systems. Alternative systems using gel-based EEG electrodes are widely used in both clinical and research settings. Although they provide higher SNR EEG recordings, they required longer preparation time (~40-60 min) and thus are less suitable for pediatric use. Alternatively, several laboratories use low-density gel-based EEG systems during the MEG recordings, which are advantageous in terms of preparation time (compared to HD-EEG systems), but they offer significantly lower spatial resolution due to the reduced number of electrodes covering the entire scalp12,16,72,73.
At present, the localization of the epileptogenic brain areas in patients with epilepsy is still mainly achieved with iEEG monitoring. Moreover, the methodology for the precise localization of eloquent brain areas is poorly defined, and the experiment setups currently used in MEG laboratories are inappropriate for pediatric patients, while the use of HD-EEG for this purpose is very limited. Accurate localization of these areas may facilitate the presurgical evaluation and augment the surgical planning for either resection or iEEG electrode placement. So far, several studies investigated the contribution of either ESI or MSI in the presurgical evaluation of patients with DRE and focal epilepsies for the identification of the EZ12,13,14,15,16,17,18,19&#160;and eloquent areas of the somatosensory cortex41, respectively. Few studies have shown better source localization results and outcome prediction performance using EMSI compared to either MSI or ESI alone13,31,42. Despite these findings, recording of MEG and EEG is rarely performed simultaneously, and MSI and ESI are implemented in only a few epilepsy centers worldwide. To our best knowledge, this is the first study that provides suggestions for collecting and analyzing simultaneous MEG and HD-EEG data, as well as performing EMSI in pediatric epilepsy for the noninvasive identification of the irritative zone, SOZ, and eloquent brain areas, namely primary visual, motor, auditory, and somatosensory cortices.
Here, we performed EMSI on interictal spikes and ictal events detected on simultaneous noninvasive data from two patients with DRE (Cases 1 and 2) and achieved a source localization error of ~9 mm and ~12 mm from the SOZ, respectively, in line with previous studies42. Impressively, such a method achieved a localization accuracy comparable to the intracranial findings (i.e., ESI on iEEG data), with clustered dipoles localized in the brain area pinpointed as epileptogenic by the clinical observations (Figure 3C and Figure 4B). Using noninvasive data from a third representative patient with DRE (Case 3), we also performed EMSI on visual-, motor-, auditory-, and somatosensory-evoked activities and found prominent source activation patterns in the corresponding eloquent brain areas (i.e., visual, motor, auditory, and somatosensory cortices) (Figure 5C, Figure 6C, Figure 7C, and Figure 8C).
Our results were derived from the fusion of complementary information captured from MEG and EEG modalities that may improve localization accuracy. EEG is well known to reflect all intracranial currents, whereas MEG is mostly sensitive to tangential sources and blind to deep brain sources29,74. As shown in this study, combining MEG and EEG can, therefore, overcome the limitations of each modality, provide superior localization results, and identify epileptogenic and eloquent brain areas that either ESI or MSI may have missed if used alone. Moreover, we present an alternative noninvasive approach for mapping eloquent brain areas using EMSI in those patients who did not undergo fMRI during their presurgical evaluation.
The localization of epileptogenic and eloquent brain areas using noninvasive techniques, such as simultaneous MEG and EEG, is an essential step during the presurgical evaluation of children with DRE for complete removal or disconnection of the EZ while preserving eloquent cortical areas. The proposed methodology offers a detailed description of the acquisition and analysis of simultaneous MEG and EEG data that supports its application not only in the presurgical epilepsy evaluation but also in cognitive neurosciences for exploring physiological functions of the healthy brain in both typically developing children as well as healthy adults, as well as morphological and functional brain changes associated with epilepsy or other neurological disorders. Future studies investigating epileptogenic brain networks may also assess whether network hubs (i.e., highly connected brain regions) estimated noninvasively using EMSI on simultaneous MEG and HD-EEG data can more accurately localize the EZ in children with DRE than those estimated using MSI and/or ESI alone75,76,77. Furthermore, the noninvasive mapping of spatiotemporal propagations of spikes and ripples (i.e., high-frequency oscillations, &#62;80 Hz), estimated through EMSI, can help to better understand the pathophysiological mechanisms of propagating epileptiform activity and noninvasively assess the onset generator of these propagations that is a precise biomarker of the EZ78,79. The presented protocol may help to further investigate the complementarity of MEG and EEG systems by examining the sensitivity of MEG and EEG sensor arrays to sources of different orientations. Such analysis may provide insights into the electrophysiological properties of the brain while performing simultaneous MEG and HD-EEG.

Epilepsy is one of the most common and disabling neurological disorders characterized by recurrent and unprovoked seizures that can be either focal or generalized in nature. Despite the availability of several effective pharmacologic therapies (e.g., antiseizure medications [ASMs]), about 20-30% of these patients are unable to control their seizures and suffer from drug-resistant epilepsy (DRE)1. For these patients, epilepsy surgery is the most effective treatment to eliminate seizures; a successful surgery can be achieved through the complete resection (or ablation/disconnection) of the epileptogenic zone (EZ), defined as the minimal area indispensable for the generation of seizures2. Accurate delineation and resection (or ablation/disconnection) of the EZ while preserving the eloquent cortex are crucial factors in ensuring seizure freedom. To establish surgical candidacy, several noninvasive diagnostic tools are used by a multidisciplinary team to define different cortical areas (i.e., irritative zone, seizure onset zone [SOZ], functional deficit zone, and epileptogenic lesion), which serve as indirect approximators of the EZ3. Extra-operative monitoring with intracranial EEG (iEEG) is required when none of these methods unequivocally identify the EZ. The role of iEEG is to define precisely the EZ by localizing the SOZ (i.e., the brain area where clinical seizures generate) and map eloquent brain areas. Yet, it presents serious limitations due to its invasiveness4,5,6, it offers limited spatial coverage, and it needs a clear presurgical localization hypothesis7. As a result, the actual focus and extent of the SOZ may be missed, leading to unsuccessful surgery. Also, its interpretation requires the recording of multiple stereotyped clinical seizures during several days of hospitalization, which increases the chances of complications (e.g., infection and/or bleeding)5. Hence, there is an unmet need to develop reliable and noninvasive localization methods that can provide clinically useful information and overall improve the presurgical evaluation of children with DRE.
Over the last decades, electric and magnetic source imaging (ESI and MSI) have been increasingly utilized in the presurgical evaluation of patients with DRE for the delineation of epileptogenic as well as functional brain areas. Particularly, ESI and MSI allow the reconstruction of neural sources from noninvasive recordings, such as high-density EEG (HD-EEG) and magnetoencephalography (MEG), to help guide surgical planning or iEEG electrode placement. ESI and MSI can be applied for localizing either interictal epileptiform discharges (IEDs), such as spikes and sharp waves, or ictal (seizure) activity. It may further be used for the localization of different functional brain areas involved in sensory, motor, auditory, and cognitive functions. The reconstruction of electrophysiological events, such as IEDs and seizures, allows the identification of the irritative zone (i.e., the brain area where IEDs originate) and the SOZ, respectively, which are considered a valid surrogate for the EZ localization. The localization of the eloquent cortex (i.e., the brain areas indispensable for defined cortical functions)3 allows instead to map the location and extent of eloquent areas with respect to the planned resection and, therefore, reduce in advance potential functional deficits that may be expected from epilepsy surgery8,9,10,11. Several studies investigated the clinical utility of ESI and/or MSI in the presurgical evaluation of epilepsy, and they showed promising findings in the delineation of the EZ12,13,14,15,16,17,18,19. For example, Mouthaan et al.14 performed an extensive meta-analysis using noninvasive data of 11 prospective and retrospective epilepsy studies and reported that these source localization techniques can overall identify the EZ with high sensitivity (82%) and low specificity (53%). Other studies also showed that MSI and ESI can correctly localize the epileptic focus within the resected area in epileptic patients having a normal magnetic resonance imaging (MRI)19,20,21. These localization results are particularly important for those patients who are ineligible for epilepsy surgery due to inconclusive clinical or imaging findings. In summary, ESI and MSI can contribute significantly to the presurgical mapping of epileptogenic as well as functional brain areas in patients with DRE.
Despite these encouraging findings, such techniques are currently performed in only a few tertiary epilepsy centers on a regular basis and are often underutilized in pediatric populations. Moreover, HD-EEG and MEG are rarely recorded simultaneously, although they provide both confirmatory and complementary information. MEG is sensitive to detect superficial sources with tangential orientation but is blind to radially oriented sources located at the gyri or deeper areas of the brain22,23,24,25,26. Furthermore, MEG provides better spatial resolution (millimeters) compared to EEG16,22,25. Unlike EEG signals, MEG signals are reference-free and are essentially unaffected by different conductivities of brain tissues (i.e., meninges, cerebrospinal fluid, skull, and scalp)25,27 providing undistorted measurements of the magnetic fields produced by the brain. On the other hand, EEG can detect sources of all orientations, but it offers lower spatial resolution than MEG and is more susceptible to artifacts26,28. Due to these complementary sensitivities to source orientation and depth, approximately 30% of the epileptiform activity (e.g., IEDs) can only be recorded on MEG but not on EEG, and vice versa26,29,30,31,32. Contrary to EEG, which allows prolonged recordings, capturing clinical seizures with MEG is challenging due to the restricted recording time that is usually insufficient to record ictal events in most patients. Furthermore, artifacts caused by seizure-related head movements can often interfere with the quality of MEG recordings29,33,34,35. On the other hand, MEG recordings are faster and easier compared to EEG, especially in children since there is no need to attach sensors over the children&#39;s head35.
Advances in hardware have made it possible to record simultaneously MEG and HD-EEG data with a high number of sensors (over 550 sensors) covering the whole head. Moreover, modern developments in EEG technologies have minimized the preparation time for HD-EEG to less than a quarter of an hour36. This is particularly important for pediatric populations with challenging behaviors who are unable to stay still for prolonged periods. Furthermore, advancements in software technologies have allowed the combination of ESI and MSI into a single solution, namely electromagnetic source imaging (EMSI), performed on simultaneous HD-EEG and MEG recordings. Several theoretical and empirical studies reported higher source localization accuracy with EMSI than either modality alone13,30,31,37,38,39,40,41. Using different source localization approaches to reconstruct the activity in response to sensory stimuli, Sharon et al.37 found that EMSI had consistently better localization results than either ESI or MSI alone compared to functional MRI&#160;(fMRI), which serves as noninvasive benchmark of precise localization accuracy. The authors suggested that this improved localization is due to the increased number of sensors for solving the inverse solution and the different sensitivity patterns of the two imaging modalities37. Similarly, Yoshinaga et al.31 performed dipole analysis on simultaneous EEG and MEG data of patients with intractable localization-related epilepsy and showed that EMSI provided information that would be not obtainable by using only one modality alone and led to successful localization for epilepsy surgery in one of the patients analyzed. In a prospective blinded study, Duez et al.13 showed that EMSI achieved a significantly higher odds ratio (i.e., probability of becoming seizure-free) compared to ESI and MSI, a localization accuracy &#8805;52%, and a concordance &#8805;53% and &#8805;36% with the irritative and SOZ, respectively. A more recent study from our group42 has shown that EMSI provided superior localization estimates and better outcome prediction performance than either ESI or MSI alone, with localization errors from resection and SOZ of ~8 mm and ~15 mm, respectively. Despite these promising findings, there is a lack of studies that provide the methodological framework regarding EMSI in children with DRE.
This study illustrates the experimental setup for performing simultaneous MEG and HD-EEG recordings as well as the methodological framework for analyzing these data aiming to localize the irritative zone, SOZ, and eloquent brain areas in children with DRE. More specifically, the experimental setups are presented for (i) recording and localizing interictal and ictal epileptiform activity during sleep and (ii) recording visual-, motor-, auditory-, and somatosensory-evoked responses and mapping relevant eloquent brain areas (i.e., visual, motor, auditory, and somatosensory) during a visuomotor task, as well as auditory and somatosensory stimulations. Detailed steps of the data analysis pipeline are further presented for performing EMSI as well as individual ESI and MSI using equivalent current dipole (ECD) and dynamic statistical parametric mapping (dSPM).

<table><tbody><tr><td>AIRSTIM unit</td><td>SD Instruments</td><td>N/A</td><td>The SDI AIRSTIM system is an alternative unconditioned stimulus to shock</td></tr><tr><td>Baby Shampoo&amp;nbsp;</td><td>Johnson&#039;s</td><td>N/A</td><td>Baby Shampoo is as gentle to the eyes as pure water and is specially designed to gently cleanse baby&amp;rsquo;s delicate hair and scalp.&amp;nbsp;</td></tr><tr><td>Control III disinfectant cleaning solution</td><td>Maril Products, Inc.</td><td>http://www.controlthree.com/</td><td>Disinfectant and germicide solution formulated for hospitals</td></tr><tr><td>Elekta Neuromag</td><td>TRIUX</td><td>NM24132A</td><td>Comprehensive bioelectromagnetic measurement system characterized by 306-channel neuromagnetometer for functional brain studies</td></tr><tr><td>FASTRAK</td><td>Polhemus technology</td><td>NS-7806</td><td>Using A/C electromagnetic technology, FASTRAK delivers accurate position and orientation data, with virtually no latency. With a single magnetic source, FASTRAK delivers data for up to four sensors. The source emits an electromagnetic field, sensors within the field of range are tracked in full 6DOF (6 Degrees-Of-Freedom). Setup is simple and intuitive, with no user calibration required.</td></tr><tr><td>Genuine Grass Reusable Cup EEG Electrodes</td><td>Natus Medical, Inc.</td><td>N/A</td><td>Each Genuine Grass EEG Electrode undergoes rigorous mechanical and electrical testing to assure long life for unsurpassed recording clarity and dependability.</td></tr><tr><td>Geodesic Sensor Net</td><td>Electrical Geodesics, Inc.</td><td>S-MAN-200-GSNR-001</td><td>32 to 256 electrodes to place on the human head to aquire dense-array electroencephalography data</td></tr><tr><td>GeoScan Sensor Digitization System</td><td>Electrical Geodesics, Inc.</td><td>8100550-03</td><td>Handheld Scanner and Software for 3D electrode position registration</td></tr><tr><td>Natus Xltek NeuroWorks</td><td>Natus Medical, Inc.</td><td>https://natus.com/</td><td>The Natus NeuroWorks platform simplifies the process of collecting, monitoring and managing data for routine EEG testing, ambulatory EEG, long-term monitoring, ICU monitoring, and research studies.</td></tr><tr><td>Natus NeuroWorks EEG Software</td><td>Natus Medical, Inc.</td><td>https://natus.com/neuro/neuroworks-eeg-software/</td><td>NeuroWorks EEG software simplifies the process of collecting, monitoring, trending and managing EEG testing data, allowing care providers to save time and focus on delivering the best care.</td></tr><tr><td>ROSA ONE Brain</td><td>Zimmer Biomet</td><td>https://www.zimmerbiomet.com/en/products-and-solutions/zb-edge/robotics/rosa-brain.html</td><td>ROSA ONE Brain is a robotic solution to assist surgeons in planning and performing complex neurosurgical procedures through a small drill hole in the skull.&amp;nbsp;</td></tr><tr><td>Ten20 Conductive Paste</td><td>Weaver and company</td><td>N/A</td><td>Ten20 contains the right balance of adhesiveness and conductivity, enabling the electrodes to remain in place while allowing the transmittance of electrical signals.</td></tr></tbody></table>

electromagnetic source imaging in presurgical evaluation of children with drug-resistant epilepsy

For children with drug-resistant epilepsy (DRE), seizure freedom relies on the delineation and resection (or ablation/disconnection) of the epileptogenic zone (EZ) while preserving the eloquent brain areas. The development of a reliable and noninvasive localization method that provides clinically useful information for the localization of the EZ is, therefore, crucial to achieving successful surgical outcomes. Electric and magnetic source imaging (ESI and MSI) have been increasingly utilized in the presurgical evaluation of these patients showing promising findings in the delineation of epileptogenic as well as eloquent brain areas. Moreover, the combination of ESI and MSI into a single solution, namely electromagnetic source imaging (EMSI), performed on simultaneous high-density electroencephalography (HD-EEG) and magnetoencephalography (MEG) recordings has shown higher source localization accuracy than either modality alone. Despite these encouraging findings, such techniques are performed in only a few tertiary epilepsy centers, are rarely recorded simultaneously, and are underutilized in pediatric cohorts. This study illustrates the experimental setup for recording simultaneous MEG and HD-EEG data as well as the methodological framework for analyzing these data aiming to localize the irritative zone, the seizure onset zone, and eloquent brain areas in children with DRE. More specifically, the experimental setups are presented for (i) recording and localizing interictal and ictal epileptiform activity during sleep and (ii) recording visual-, motor-, auditory-, and somatosensory-evoked responses and mapping relevant eloquent brain areas (i.e., visual, motor, auditory, and somatosensory) during visuomotor task, as well as auditory and somatosensory stimulations. Detailed steps of the data analysis pipeline are further presented for performing EMSI as well as individual ESI and MSI using equivalent current dipole (ECD) and dynamic statistical parametric mapping (dSPM).

Author Spotlight: Advancing Pediatric Epilepsy Surgery in Children Through Novel Biomarkers and Enhanced Localization

Magnetoencephalography (MEG) and high-density electroencephalography (HD-EEG) are rarely recorded simultaneously, although they yield confirmatory and complementary information. Here, we illustrate the experimental setup for recording simultaneous MEG and HD-EEG and the methodology for analyzing these data aiming to localize epileptogenic and eloquent brain areas in children with drug-resistant epilepsy.

Electromagnetic Source Imaging in Presurgical Evaluation of Children with Drug-Resistant Epilepsy

For children with drug-resistant epilepsy (DRE), seizure freedom relies on the delineation and resection (or ablation/disconnection) of the epileptogenic ...

electromagnetic-source-imaging-presurgical-evaluation-children-with

Research

JoVE Journal

Neuroscience

2.5K Views.  Cook Children’s Health Care System. Magnetoencephalography (MEG) and high-density electroencephalography (HD-EEG) are rarely recorded simultaneously, although they yield confirmatory and complementary information. Here, we illustrate the experimental setup for recording simultaneous MEG and HD-EEG and the methodology for analyzing these data aiming to localize epileptogenic and eloquent brain areas in children with drug-resistant epilepsy.

Video: Author Spotlight: Advancing Pediatric Epilepsy Surgery in Children Through Novel Biomarkers and Enhanced Localization

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.

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 ...

Forebrain Electrophysiological Recording in Larval Zebrafish

Epilepsy affects nearly 3 million people in the United States and up to 50 million people worldwide. Defined as the occurrence of spontaneous unprovoked seizures, epilepsy can be acquired as a result of an insult to the brain or a genetic mutation. Efforts to model seizures in animals have primarily utilized acquired insults (convulsant drugs, stimulation or brain injury) and genetic manipulations (antisense knockdown, homologous recombination or transgenesis) in rodents. Zebrafish are a vertebrate model system1-3 that could provide a valuable alternative to rodent-based epilepsy research. Zebrafish are used extensively in the study of vertebrate genetics or development, exhibit a high degree of genetic similarity to mammals and express homologs for ~85% of known human single-gene epilepsy mutations. Because of their small size (4-6 mm in length), zebrafish larvae can be maintained in fluid volumes as low as 100 &mu;l during early development and arrayed in multi-well plates. Reagents can be added directly to the solution in which embryos develop, simplifying drug administration and enabling rapid in vivo screening of test compounds4. Synthetic oligonucleotides (morpholinos), mutagenesis, zinc finger nuclease and transgenic approaches can be used to rapidly generate gene knockdown or mutation in zebrafish5-7. These properties afford zebrafish studies an unprecedented statistical power analysis advantage over rodents in the study of neurological disorders such as epilepsy. Because the "gold standard" for epilepsy research is to monitor and analyze the abnormal electrical discharges that originate in a central brain structure (i.e., seizures), a method to efficiently record brain activity in larval zebrafish is described here. This method is an adaptation of conventional extracellular recording techniques and allows for stable long-term monitoring of brain activity in intact zebrafish larvae. Sample recordings are shown for acute seizures induced by bath application of convulsant drugs and spontaneous seizures recorded in a genetically modified fish.

A simple method to record extracellular field potentials in the larval zebrafish forebrain is described. The method provides a robust in vivo read-out of seizure-like activity. This technique can be used with genetically modified zebrafish larvae carrying epilepsy-related genes or seizures evoked by administration of convulsant drugs.

Epilepsy affects nearly 3 million people in the United States and up to 50 million people worldwide. Defined as the occurrence of spontaneous unprovoked ...

Deep Brain Stimulation with Simultaneous fMRI in Rodents

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for using blood oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI) to image rodents with simultaneous DBS. DBS fMRI presents a number of technical challenges, including accuracy of electrode implantation, MR artifacts created by the electrode, choice of anesthesia and paralytic to minimize any neuronal effects while simultaneously eliminating animal motion, and maintenance of physiological parameters, deviation from which can confound the BOLD signal.&#160;Our laboratory has developed a set of procedures that are capable of overcoming most of these possible issues. For electrical stimulation, a homemade tungsten bipolar microelectrode is used, inserted stereotactically at the stimulation site in the anesthetized subject. In preparation for imaging, rodents are fixed on a plastic headpiece and transferred to the magnet bore. For sedation and paralysis during scanning, a cocktail of dexmedetomidine and pancuronium is continuously infused, along with a minimal dose of isoflurane; this preparation minimizes the BOLD ceiling effect of volatile anesthetics. In this example experiment, stimulation of the subthalamic nucleus (STN) produces BOLD responses which are observed primarily in ipsilateral cortical regions, centered in motor cortex. Simultaneous DBS and fMRI allows the unambiguous modulation of neural circuits dependent on stimulation location and stimulation parameters, and permits observation of neuronal modulations free of regional bias. This technique may be used to explore the downstream effects of modulating neural circuitry at nearly any brain region, with implications for both experimental and clinical DBS.

This protocol describes a standard method for simultaneous functional magnetic resonance imaging and deep brain stimulation in the rodent. The combined use of these experimental tools allows for the exploration of global downstream activity in response to electrical stimulation at virtually any brain target.

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for ...

Brain Source Imaging in Preclinical Rat Models of Focal Epilepsy using High-Resolution EEG Recordings

Electroencephalogram (EEG) has been traditionally used to determine which brain regions are the most likely candidates for resection in patients with focal epilepsy. This methodology relies on the assumption that seizures originate from the same regions of the brain from which interictal epileptiform discharges (IEDs) emerge. Preclinical models are very useful to find correlates between IED locations and the actual regions underlying seizure initiation in focal epilepsy. Rats have been commonly used in preclinical studies of epilepsy1; hence, there exist a large variety of models for focal epilepsy in this particular species. However, it is challenging to record multichannel EEG and to perform brain source imaging in such a small animal. To overcome this issue, we combine a patented-technology to obtain 32-channel EEG recordings from rodents2 and an MRI probabilistic atlas for brain anatomical structures in Wistar rats to perform brain source imaging. In this video, we introduce the procedures to acquire multichannel EEG from Wistar rats with focal cortical dysplasia, and describe the steps both to define the volume conductor model from the MRI atlas and to uniquely determine the IEDs. Finally, we validate the whole methodology by obtaining brain source images of IEDs and compare them with those obtained at different time frames during the seizure onset.

This video introduces the preparation, recording, and source analysis procedures of high-resolution EEG on sedated rats with a particular preclinical model of focal epilepsy under noninvasive conditions.

Electroencephalogram (EEG) has been traditionally used to determine which brain regions are the most likely candidates for resection in patients with ...

Medicine

A Familiarization Protocol Facilitates the Participation of Children with ASD in Electrophysiological Research

This paper includes a detailed description of a familiarization protocol, which is used as an integral component of a larger research protocol to collect electroencephalography (EEG) data and Event-Related Potentials (ERPs). At present, the systems available for the collection of high-quality EEG/ERP data make significant demands on children with developmental disabilities, such as those with an Autism Spectrum Disorder (ASD). Children with ASD may have difficulty adapting to novel situations, tolerating uncomfortable sensory stimuli, and sitting quietly. This familiarization protocol uses Evidence-Based Practices (EBPs) to increase research participants&#39; knowledge and understanding of the specific activities and steps of the research protocol. The tools in this familiarization protocol are a social narrative, a visual schedule, the Premack principle, role-playing, and modeling. The goal of this familiarization protocol is to increase understanding and agency and to potentially reduce anxiety for child participants, resulting in a greater likelihood of the successful completion of the research protocol for the collection of EEG/ERP data.

Electrophysiological research is an important tool for identifying biomarkers of developmental disorders, including Autism Spectrum Disorders (ASD), but data collection in these populations remains challenging. This work presents a familiarization protocol to accompany research that includes electroencephalography (EEG) to improve the likelihood of collecting EEG data from children with ASD.

This paper includes a detailed description of a familiarization protocol, which is used as an integral component of a larger research protocol to collect ...

Analyzing Neural Activity and Connectivity Using Intracranial EEG Data with SPM Software

Measuring neural activity and connectivity associated with cognitive functions at high spatial and temporal resolutions is an important goal in cognitive neuroscience. Intracranial electroencephalography (EEG) can directly record electrical neural activity and has the unique potential to accomplish this goal. Traditionally, averaging analysis has been applied to analyze intracranial EEG data; however, several new techniques are available for depicting neural activity and intra- and inter-regional connectivity. Here, we introduce two analytical protocols we recently applied to analyze intracranial EEG data using the Statistical Parametric Mapping (SPM) software: time-frequency SPM analysis for neural activity and dynamic causal modeling of induced responses for intra- and inter-regional connectivity. We report our analysis of intracranial EEG data during the observation of faces as representative results. The results revealed that the inferior occipital gyrus (IOG) showed gamma-band activity at very early stages (110 ms) in response to faces, and both the IOG and amygdala showed rapid intra- and inter-regional connectivity using various types of oscillations. These analytical protocols have the potential to identify the neural mechanisms underlying cognitive functions with high spatial and temporal profiles.

We present two analytical protocols that can be used to analyze intracranial electroencephalography data using the Statistical Parametric Mapping (SPM) software: time-frequency statistical parametric mapping analysis for neural activity, and dynamic causal modeling of induced responses for intra- and inter-regional connectivity.

Measuring neural activity and connectivity associated with cognitive functions at high spatial and temporal resolutions is an important goal in cognitive ...

Behavioral And Physiological Analysis In A Zebrafish Model Of Epilepsy

Epilepsy represents one of the most common neurological disorders, affecting an estimated 50 million people worldwide. Recent advances in genetic research have uncovered a large spectrum of genes implicated in various forms of epilepsy, highlighting the heterogeneous nature of this disorder. Appropriate animal models are essential for investigating the pathological mechanisms triggered by genetic mutations implicated in epilepsy and for developing specialized, targeted therapies. In recent years, zebrafish has emerged as a valuable vertebrate organism for modeling epilepsies, with the use of both genetic manipulation and exposure to known epileptogenic drugs, such as pentylenetetrazole (PTZ), to identify novel anti-epileptic therapeutics. Deleterious mutations in the mTOR regulator DEPDC5 have been associated with various forms of focal epilepsies and knock-down of the zebrafish orthologue causes hyperactivity associated with spontaneous seizure-like episodes, as well as enhanced electrographic activity and characteristic turn wheel swimming. Here, we described the method involved in generating the DEPDC5 loss-of-function model and illustrate the protocol for assessing motor activity at 28 and 48 h post fertilization (hpf), as well as a method for recording field activity in the zebrafish optic tectum. An illustration of the effect of the epileptogenic drug PTZ on neuronal activity over time is also provided.

Here, we present a protocol for the development and the characterization of a zebrafish model of epilepsy resulting from the transient inhibition of the DEPDC5 gene.

Epilepsy represents one of the most common neurological disorders, affecting an estimated 50 million people worldwide. Recent advances in genetic research ...

Studying Brain Function in Children Using Magnetoencephalography

Magnetoencephalography (MEG) is a non-invasive neuroimaging technique which directly measures magnetic fields produced by the electrical activity of the human brain. MEG is quiet and less likely to induce claustrophobia compared with magnetic resonance imaging (MRI). It is therefore a promising tool for investigating brain function in young children. However, analysis of MEG data from pediatric populations is often complicated by head movement artefacts which arise as a consequence of the requirement for a spatially-fixed sensor array that is not affixed to the child's head. Minimizing head movements during MEG sessions can be particularly challenging as young children are often unable to remain still during experimental tasks. The protocol presented here aims to reduce head movement artefacts during pediatric MEG scanning. Prior to visiting the MEG laboratory, families are provided with resources that explain the MEG system and the experimental procedures in simple, accessible language. An MEG familiarization session is conducted during which children are acquainted with both the researchers and the MEG procedures. They are then trained to keep their head still whilst lying inside an MEG simulator. To help children feel at ease in the novel MEG environment, all of the procedures are explained through the narrative of a space mission. To minimize head movement due to restlessness, children are trained and assessed using fun and engaging experimental paradigms. In addition, children's residual head movement artefacts are compensated for during the data acquisition session using a real-time head movement tracking system. Implementing these child-friendly procedures is important for improving data quality, minimizing participant attrition rates in longitudinal studies, and ensuring that families have a positive research experience.

This article introduces a child-friendly research protocol designed to improve data quality by reducing head movement during pediatric magnetoencephalography (MEG). We familiarize families with the MEG environment, train children to remain still using an MEG simulator, and correct for residual head movement artefacts using a real-time head movement detection system.

Magnetoencephalography (MEG) is a non-invasive neuroimaging technique which directly measures magnetic fields produced by the electrical activity of the ...

Advanced Diffusion Imaging in The Hippocampus of Rats with Mild Traumatic Brain Injury

Mild traumatic brain injury (mTBI) is the most common type of acquired brain injury. Since patients with traumatic brain injury show a tremendous variability and heterogeneity (age, gender, type of trauma, other possible pathologies, etc.), animal models play a key role in unraveling factors that are limitations in clinical research. They provide a standardized and controlled setting to investigate the biological mechanisms of injury and repair following TBI. However, not all animal models mimic the diffuse and subtle nature of mTBI effectively. For example, the commonly used controlled cortical impact (CCI) and lateral fluid percussion injury (LFPI) models make use of a craniotomy to expose the brain and induce widespread focal trauma, which are not commonly seen in mTBI. Therefore, these experimental models are not valid to mimic mTBI. Thus, an appropriate model should be used to investigate mTBI. The Marmarou weight drop model for rats induces similar microstructural alterations and cognitive impairments as seen in patients who sustain mild trauma; therefore, this model was selected for this protocol. Conventional computed tomography and magnetic resonance imaging (MRI) scans commonly show no damage following a mild injury, because mTBI induces often only subtle and diffuse injuries. With diffusion weighted MRI, it is possible to investigate microstructural properties of brain tissue, which can provide more insight into the microscopic alterations following mild trauma. Therefore, the goal of this study is to obtain quantitative information of a selected region-of-interest (i.e., hippocampus) to follow up disease progression after obtaining a mild and diffuse brain injury.

The overall goal of this procedure is to obtain quantitative microstructural information of the hippocampus in a rat with mild traumatic brain injury. This is done using an advanced diffusion-weighted magnetic resonance imaging protocol and region-of-interest based analysis of parametric diffusion maps.

Mild traumatic brain injury (mTBI) is the most common type of acquired brain injury. Since patients with traumatic brain injury show a tremendous ...

Detecting Pre-Stimulus Source-Level Effects on Object Perception with Magnetoencephalography

Pre-stimulus oscillatory brain activity influences upcoming perception. The characteristics of this pre-stimulus activity can predict whether a near-threshold stimulus will be perceived or not perceived, but can they also predict which one of two competing stimuli with different perceptual contents is perceived? Ambiguous visual stimuli, which can be seen in one of two possible ways at a time, are ideally suited to investigate this question. Magnetoencephalography (MEG) is a neurophysiological measurement technique that records magnetic signals emitted as a result of brain activity. The millisecond temporal resolution of MEG allows for a characterization of oscillatory brain states from as little as 1 second of recorded data. Presenting an empty screen around 1 second prior to the ambiguous stimulus onset therefore provides a time window in which one can investigate whether pre-stimulus oscillatory activity biases the content of upcoming perception, as indicated by participants&#39;&#160;reports. The spatial resolution of MEG is not excellent, but sufficient to localise sources of brain activity at the centimetre scale. Source reconstruction of MEG activity then allows for testing hypotheses about the oscillatory activity of specific regions of interest, as well as the time- and frequency-resolved connectivity between regions of interest. The described protocol enables a better understanding of the influence of spontaneous, ongoing brain activity on visual perception.

This article describes how to set up an experiment that allows detecting pre-stimulus source-level influences on object perception using magnetoencephalography (MEG). It covers stimulus material, experimental design, MEG recording, and data analysis.