Name: A Standardized Protocol for Functional Motor Mapping Using Navigated Transcranial Magnetic Stimulation (Video) | JoVE Journal
Uploaded: 2026-02-27T13:30:00.000+00:00
Duration: 10 min 27 s
Description: Learn navigated transcranial magnetic stimulation combined with diffusion tensor imaging for motor mapping and corticospinal tract assessment. This standardized protocol enables precise preoperative functional mapping for neurosurgical planning.

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This work was supported by the Independent Research Fund Denmark (Grant number: 3165-00230B), Aage &#38; Johanne Louis-Hansens Foundation (Grant number: 25-1-17926), and Muskelsvindfonden (Grant number: 2025-0010)

The authors have nothing to disclose.

In this paper, we present a standardized and reproducible protocol for functional motor cortical mapping with nTMS directly applicable to preoperative surgical planning. By combining neuronavigation with the subject&#39;s anatomical brain reconstruction, this standardized protocol makes it possible to identify and delineate motor-eloquent cortical regions during an exam lasting less than 90 min, depending on the number of muscles studied. This approach is particularly relevant in patients with motor-eloquent tumors, where anatomical reconstruction of the CST is often limited by two factors: (i) anatomical displacement due to mass effect and/or edema and (ii) functional reorganization of motor representations. Anatomical seeding tractography based on fixed anatomical landmarks can therefore be misleading in localizing the cortical origin and propagating errors throughout the fiber tracking. Functional motor cortical mapping addresses this issue by using nTMS-positive sites as cortical seeds, thereby anchoring tractography to the patient&#39;s current motor map that drives corticospinal output. During post-processing analysis, the cortical ROIs derived from the motor maps should be enlarged by 2-3 mm to mitigate fusion-associated mismatch and to standardize ROI volume (0.9 &#177; 0.1 cm3), reducing operator and between-subject variability and improving CST tractography comparability59. Compared with landmark-based tractography, nTMS-seeded tractography yields more plausible and somatotopically consistent CST reconstructions, with fewer aberrant streamlines and lower inter-rater variability27,61,62. Compared with fMRI-based seeding, nTMS-based tractography also produces more plausible reconstructions and higher interrater consistency in patients with tumors adjacent to the CST25. It also allows the extraction of several metrics from the nTMS-motor mapping and the nTMS-seeded CST, which may serve as a predictive factor of postoperative motor outcome. At the cortical level, the presence of nTMS-responsive sites within the tumor has been associated with an increased risk of motor deficit, with a positive predictive value ranging from 50-90%30,63,64,65. In contrast, resection of nTMS-negative sites is considered safe, with a high negative predictive value ranging from 90-100%30,31,65. At the subcortical level, a tumor-to-tract distance &#60;8-12 mm has been identified as a critical threshold associated with an elevated risk of post-operative deficit, as long as the tumor does not invade the precentral gyrus66,67,68,69,70,71. Additionally, microstructural alterations of the nTMS-seeded CST (decreased Fractional Anisotropy with increased Mean Diffusivity) have also been proposed as further risk factors for post-operative deficit70. Finally, the use of nTMS-based tractography has been associated with a greater extent of resection and prolonged survival while preserving motor function, supporting its integration into preoperative planning72.
During motor mapping, a key parameter that strongly influences the spatial distribution of MEPs and the interpretability of motor maps is the stimulation intensity (SI). Higher SI increases response probability and spatial spread (risking false positive responses), whereas insufficient SI increases the risk of false negative responses. To minimize this bias, the SI should be scaled relative to the RMT and, when possible, adjusted to maintain a stable target EF. In practice, near-threshold SI strikes a balance between sensitivity and specificity and provides conservative maps close to direct electrical stimulation mapping. On the other hand, choosing a supra-threshold SI (e.g., 120% RMT) can be justified when clinical safety prioritizes sensitivity at the map margins, acknowledging that higher SI systematically expands the motor map73. In the context of mapping multiple muscles, the use of a single SI may bias the mapping toward the lowest-threshold muscle, as adjacent muscles could have different excitability profiles. Accordingly, RMT should be estimated for each muscle74. On the other hand, significant changes in cortical excitability, reflected by unexpected changes in MEP amplitudes, may occur during a motor mapping session, requiring re-estimation of the RMT and adjustment of the SI.
The use of stimulation grids during motor mapping helps standardize spacing and facilitates map quantification (i.e., by counting active squares). However, grid size directly shapes the results: large squares may overestimate map size, whereas small squares increase the risk of undersampling. Recent evidence suggests that nTMS mapping can be performed without grids, using an anatomy-guided approach with denser stimuli near the anatomical landmarks and map edges75.
Several quantitative parameters can be derived from motor mapping, such as the center of gravity (CoG), motor map area, and volume. The CoG is defined as the amplitude-weighted location in coordinates which represents the center of the motor representation58. Serial examinations have shown shifts in CoG in brain tumor patients76,77,78, capturing evidence of functional reorganization over time in the motor cortex. Motor map area and volume represent the spatial extent of the motor representation. Area is commonly derived either by counting the active squares on a stimulation grid or by using spline interpolation in grid-free stimulation, which connects the positive stimulation points with smooth polynomial curves to generate continuous surface or volume56. These metrics can be monitored longitudinally (follow-up study or assessment of an intervention) or compared to the contralesional hemisphere to investigate cortical motor plasticity79,80,81,82. Quantitative motor mapping metrics have the potential to be extended beyond neuro-oncology, providing biomarkers of the motor system integrity and disease-related plasticity in neurological diseases55,83.
Although nTMS is now well established for preoperative motor mapping, several limitations should be acknowledged. First, the accuracy of co-registration and cortical mapping remains partly operator dependent. Proper training in coil handling, head-tracker stability, and prompt adjustment of stimulation are required to ensure reliability and reproducibility of the technique, although previous studies have shown that nTMS provides reliable motor topography with good inter-operator agreement between expert and novice examiners84. A second limitation relates to the influence of perilesional edema and mass effect on tractography. Excessive perilesional edema can reduce the accuracy of nTMS-based CST reconstruction, particularly in voxels adjacent to the lesion85. Similarly, discrepancies between preoperative datasets and the real intraoperative anatomy may occur due to intraoperative brain shift86,87. Because brain shift cannot be fully prevented - especially in tumors with important mass effect - the accuracy of nTMS-derived motor regions (both cortical and subcortical) may decrease during the later stages of resection. Several strategies can mitigate these inaccuracies, including limiting unnecessary cortical exposure, repeatedly checking superficial anatomical landmarks88, and using intraoperative imaging such as MRI, ultrasound, or CT, combined with brain deformation correction89,90,91,92. Finally, regarding safety, nTMS has demonstrated a favorable safety profile in patients with tumor-related epilepsy. In large series, stimulation-induced seizures are rare or absent during preoperative mapping93, supporting the safety of this technique when appropriate precautions are taken.
Overall, nTMS provides clinically useful functional information to surgical planning and opens the path to longitudinal studies of motor-system plasticity in various neurological or psychiatric diseases.

Maximizing the extent of resection in motor-eloquent brain tumors while minimizing postoperative motor deficits remains a central challenge in neurosurgery. Intraoperative direct electrical stimulation (DES) mapping is the &#34;gold standard&#34; technique to provide reliable anatomo-functional information regarding the cortical and subcortical representation of motor pathways1,2,3,4,5. However, for preoperative planning, risk stratification, and optimal patient counselling, it is crucial to delineate the individual functional anatomy before surgery. The relationship between anatomy and function in cortical motor areas cannot be inferred from conventional structural brain magnetic resonance imaging (MRI), as brain tumors may induce significant anatomical distortion or plastic reorganization of motor networks.
Transcranial magnetic stimulation (TMS) was introduced as a non-invasive method to probe the motor cortex6 and was later adapted for functional mapping of the motor cortex7,8, including in preoperative testing by recording motor evoked potentials (MEPs) from different muscles with surface electromyography9,10,11. Early non-navigated TMS protocols were technically demanding and lacked anatomical accuracy. Subsequent integration with individual MRI data and electric-field-based navigation enabled precise guidance of stimulation sites, improving anatomo-functional accuracy12,13,14 and reproducibility15,16. By directly eliciting MEPs, navigated TMS (nTMS) provides millisecond-scale temporal resolution and sub-centimeter spatial localization of corticospinal output with good concordance with intraoperative DES17,18,19. Image-guided nTMS is safe, well tolerated20,21, and approved by the Food and Drug Administration (FDA) for presurgical functional mapping of the motor cortex for more than 15 years22.
In motor mapping, cortical representations are delineated by sampling MEP amplitudes across targeted stimulation sites to construct patient-specific motor maps23. Compared with task-based functional MRI (fMRI), nTMS shows closer spatial agreement with intraoperative DES24,25,26. While intraoperative decisions ultimately rely on DES when lesions abut or invade the motor areas, preoperative nTMS provides valuable complementary information by exporting stimulation-positive sites as seeds for diffusion tensor imaging (DTI) reconstruction of the corticospinal tract (CST). This approach is particularly useful to assess corticospinal integrity when tumors mainly affect the motor tracts in the subcortical white matter27,28. Moreover, preoperative nTMS motor mapping has shown good positive predictive value29,30 and high negative predictive value29,30,31, with improved surgical outcomes17,18,19,32. It has also recently been proven as an effective tool to assess postoperative motor function31,33. For these reasons, nTMS motor mapping is increasingly used for both preoperative evaluation and postoperative follow-up in neurosurgery. Methodological recommendations for cortical mapping with nTMS have been published in 201734. In light of these recent studies and the integration of modern imaging techniques, this methodology can now be refined to provide more accurate guidance for clinical and research practice.
In this paper, we present a standardized protocol for performing motor mapping with nTMS, combining different techniques to evaluate preoperative cortical and subcortical representations of motor pathways for tumor resection planning under real-life clinical conditions.

<table><tbody><tr><td>Elements software</td><td>BrainLAB AG, Munich, Germany</td><td></td><td>Image processing software and OR neuronavigation software</td></tr><tr><td>Neuronavigation TMS system&amp;nbsp;</td><td>Nexstim, Helsinki, Finland</td><td>NBS 5.1 system</td><td>Navigated TMS system with figure-of-eight coil and EMG amplifier</td></tr><tr><td>Surface electrodes for EMG recording&amp;nbsp;</td><td>Natus, Middleton, WI, USA</td><td>9013L0453</td><td>For EMG recording</td></tr></tbody></table>

a standardized protocol for functional motor mapping using navigated transcranial magnetic stimulation

A Standardized Protocol for Functional Motor Mapping Using Navigated Transcranial Magnetic Stimulation

Navigated transcranial magnetic stimulation (nTMS) is based on the integration of individual brain imaging data to determine the precise positioning of the stimulation coil, thus enabling anatomically guided stimulation of cortical targets. The interest of neuronavigation systems is well recognized in the optimization of coil positioning during repetitive TMS (rTMS) treatments. Furthermore, nTMS is increasingly applied for functional mapping of brain regions in different applications, such as the identification and delineation of eloquent motor and language areas before tumor resection. Besides its usefulness in optimizing neurosurgical procedures, nTMS mapping can also be a tool for monitoring cortical plasticity and quantifying the integrity of the motor system in various neurological diseases. This methodological paper presents a standardized protocol for motor mapping using nTMS, in combination with diffusion tensor imaging (DTI)-based reconstruction of the corticospinal tract (CST). This approach allows for precise delineation of eloquent motor cortical regions and their subcortical projections, and the detection of functional reorganizations in patients with adjacent lesions. When integrated into presurgical planning, this method provides guidance for individualized surgical strategies aimed at maximizing lesion resection while preserving motor function. The protocol presented here is reproducible, clinically applicable, and suitable for integration into routine workflows. It constitutes a promising tool for neuroplasticity research and rehabilitation planning.

We present representative steps and the results of motor mapping obtained in different healthy subjects and in patients who underwent motor mapping in a clinical environment, using our neuronavigated TMS system. CST reconstruction was performed using an imaging-processing software suitable for neurosurgery planning, capable of multimodal image registration and DTI-based tractography. The neuronavigation system integrates a navigated figure-of-eight coil, a stereotaxic camera, an EMG amplifier, and provides real-time visualization of the induced Electrical Field on the 3D brain reconstruction using an individualized multi-sphere head model.
Figure 5 displays the RMT determination at the hotspot determined from the coarse mapping. The coil position and orientation are maintained at the exact same location throughout the entire procedure with the help of the neuronavigation target. Figure6 displays a motor mapping of a healthy subject. The left lower limb (thigh, leg, foot), upper limb (shoulder, forearm, hand), and face were mapped. Positive stimulation sites (color-coded by MEP amplitude) and negative sites (gray) delineate the motor cortical representation. Figure7 displays the motor mapping and CST reconstruction in a patient with lung cancer metastasis involving the premotor region and revealed by an upper limb motor deficit.
<img alt="MRI and EEG brain scan analysis, diagram showing neural activity and imaging alignment." class="xfigimg" src="/files/ftp_upload/69776/69776fig5.jpg" data-altupdatedat="1778558422000" data-alt="Figure 5"> 
Figure 5: Coarse mapping and RMT determination at the hotspot (First Interosseus Dorsalis) in a healthy subject, using neuronavigated TMS. The hotspot, identified using coarse mapping (lower left panel), is selected as the target for the RMT determination. The coil position and orientation are maintained at the exact same location throughout the entire procedure, with the help of the neuronavigation target (lower right panel). Motor-evoked potentials (MEPs) are acquired with continuous EMG traces and epoch responses. <a href="https://www.jove.com/files/ftp_upload/69776/69776fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Functional brain mapping diagram, showing neural activity through multicolored electrode placement." class="xfigimg" src="/files/ftp_upload/69776/69776fig6.jpg" data-altupdatedat="1778558422000" data-alt="Figure 6"> 
Figure 6: Motor cortex mapping of lower limb, upper limb, and face muscles using neuronavigated TMS. Muscles recorded in the lower limbs: quadriceps femoris (green), tibialis anterior (orange), abductor hallucis (yellow). Muscles recorded in the upper limbs: abductor digiti minimi (green), flexor carpi radialis (orange), deltoid (yellow).Muscles recorded in the face: Nasalis (blue), Triangularis (purple). <a href="https://www.jove.com/files/ftp_upload/69776/69776fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Brain MRI showing fiber tracking and tumor visualization, diagram for neurological analysis." class="xfigimg" src="/files/ftp_upload/69776/69776fig7v2.jpg" data-altupdatedat="1778558422000" data-alt="Figure 7"> 
Figure 7: Motor cortical mapping and CST reconstruction for neurosurgical planning. nTMS-motor mapping (left panel) and nTMS-guided reconstruction of the corticospinal tracts (right panel) in a patient with brain metastasis (white) from lung cancer. Muscles recorded: abductor hallucis (purple), tibialis anterior (blue), deltoid (yellow), flexor carpi radialis (red), first interosseus dorsalis (green), orbicularis (cyan). <a href="https://www.jove.com/files/ftp_upload/69776/69776fig7v2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Here, we describe a standardized protocol for motor mapping using nTMS combined with diffusion tensor imaging (DTI)-based reconstruction of the corticospinal tract (CST). The protocol is reproducible, clinically feasible, and readily integrable into routine clinical workflows, providing a robust and valuable framework for motor pathway assessment, neuroplasticity research, and rehabilitation planning.

Navigated transcranial magnetic stimulation (nTMS) is based on the integration of individual brain imaging data to determine the precise positioning of ...

We present a standardized protocol for motor mapping and corticospinal tract assessment using navigated TMS combined with DTI suitable for neurosurgical planning and neuroplasticity research. While intraoperative direct electrical stimulation is the gold standard for motor mapping, existing preoperative methods are limited. Improved preoperative motor mapping methods are required to help neurosurgeons improve their surgical planning and better assess surgical risks.This protocol uses navigated TMS for accurate motor pathway mapping and improved surgical planning. To begin, import high-resolution anatomical brain images of the subject into the neuronavigation system to generate a three-dimensional brain reconstruction. Mark the key anatomical points on the magnetic resonance imaging within the neuronavigation software by identifying the nasion, right ear, and left ear.Now, position the subject on a comfortable armchair with a slight recline to reduce back tension. Adjust the headrest to support the head and neck at the inion. Place the head tracker on the subject's forehead.Using the digitizing pen, coregister the key anatomical points on the subject with the imported image in the neuronavigation software and refine the registration by digitizing additional scalp points for scalp surface matching. Validate the coregistration, ensuring a coregistration error below three millimeters. Next, give the subject a pair of earplugs and wear protective earmuffs during stimulation.Then prepare the skin over the target muscle by gently scraping it using alcohol pads and subsequently attach surface electrodes. Place surface electrodes on the muscles of interest using a belly-tendon montage and the ground electrode on a neutral site. Connect all electrodes to the electromyography amplifier.Start electromyography acquisition and verify that the muscles are at rest. On the rendered brain volume within the neuronavigation software, adjust the peeling depth to between 15 and 25 millimeters from the scalp to optimally visualize the cortical anatomy based on individual case characteristics, then start the stimulator unit. Position the stimulation coil tangentially to the scalp and stabilize it using one hand on the handle and the other on the coil surface to maintain firm contact during repositioning.Stimulate at an intensity sufficient to elicit motor revoked potentials within the amplitude range of 100 to 500 microvolts. Now, adjust the coil orientation based on the limb being mapped. For the upper limb and the face, maintain the coil perpendicular to the central sulcus to ensure a posterior-to-anterior induced current.For the lower limb, orient the coil perpendicular to the sagittal midline to generate a mid-to-lateral current direction. Perform stimulations across the target region. Space the stimulation points one to two millimeters apart and sample three parallel lines along the gyrus.Space each stimulation by at least 1.5 seconds. Stop the course mapping once 20 to 30 motor evoked potentials per muscle have been recorded and review all motor evoked potentials. Next, display the recordings using a normalized color scale to identify the hotspot for each muscle, defined as the stimulation point that evokes the largest motor evoked potential amplitude.Locate the area containing the highest amplitude responses and select the single response with the largest amplitude within that region. For each muscle, select the hotspot to determine the resting motor threshold to store the coil's position and orientation for consistent use during the threshold measurement and determine the resting motor threshold for each muscle. Ensure that the subject remains fully relaxed without any involuntary muscle contractions.For each muscle, perform stimulation at 105%to 110%of its resting motor threshold. Using the same coil orientation as during course mapping, reduce the spacing between stimulation points for higher resolution. Delineate functional motor maps as cortical areas, where navigated transcranial magnetic stimulation generates motor evoked potentials of 50 microvolts or higher.Perform stimulation until the motor maps are bordered by one or two consecutive lines of negative sites that fail to elicit motor evoked potentials. Ensure the motor maps are elliptic, with few negative sites inside. For any negative stimulation points within the motor map, perform additional stimulations at different times to account for transient changes in motor cortex excitability.Open the motor evoked potential review panel or signal viewer in the neuronavigation software. Inspect each recorded motor evoked potential to correct amplitude and latency and adjust markers if needed. Exclude artifactual or abnormal stimulation points from the dataset and display the motor map for each muscle in a binary format.Export the positive stimulation points at 15, 20, and 25-millimeter depths in binarized DICOM format. Use these files for fiber tracking, employing the positive stimulation points as seed points for corticospinal tract reconstruction. For analysis of motor mapping, import the DICOM files of the motor maps into an image analysis software compatible with neurosurgical neuronavigation for brain tumor removal.Register the anatomical T1-weighted image with the motor map DICOMs and diffusion-weighted imaging files. Generate objects from the motor map DICOMs and expand them by one to two millimeters to enhance sensitivity. Crop the motor maps to exclude the ears and nasion in order to avoid incorrect fiber reconstruction during tractography.Manually draw an ending region of interest at the inferior pontine level on the same side as the mapped hemisphere. Perform fiber tracking using the motor map regions of interest as seed points and the pontine region of interest as the endpoint. Select a suitable tractography algorithm, such as deterministic streamlined tracking or probabilistic tractography, and adjust tracking parameters according to the specific case.Finally, segment the brain tumor and create a corresponding object within the analysis software. Display the corticospinal tract either separated by limb part using different colors or as a unified tract from the entire motor mapping. The resting motor threshold was determined at the hotspot of the first interosseous dorsalis muscle identified through course mapping in a healthy subject, and the coil position and orientation were maintained at the same location during the procedure using a neuronavigation target.Motor mapping of a healthy subject revealed cortical representations for the left lower limb, upper limb, and face, with positive stimulation sites color coded by motor evoked potential amplitude and negative sites shown in gray. Motor cortical mapping and reconstruction of the corticospinal tracts were performed in a patient with brain metastasis from lung cancer involving the premotor gyrus with an upper limb motor deficit. By combining neuronavigation with the subject's anatomical brain, this NTMS protocol provides precise identification and delineation of motor eloquent cortical regions in under 19 minutes.Careful selection of stimulation intensity is critical, as it affect motor evoked potential determination and motor map interpretation. Net result level provide conservative map approximating direct electrical stimulation. This technique was first developed to provide clinically useful functional information for surgical planning.In addition, we know now that this technique is able to be applied for longitudinal assessment of motor plasticity in various neurological or psychiatric disorders.

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Source: Charalambos, C. C., et al. Bilateral Assessment of the Corticospinal Pathways of the Ankle Muscles Using Navigated Transcranial Magnetic Stimulation. J. Vis. Exp. (2019)This video demonstrates the method to assess corticospinal pathways of the leg muscles using transcranial magnetic stimulation (TMS). Rest and active muscle responses are recorded from one brain hemisphere to evaluate the contralateral corticomotor function of leg muscles.

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A Standardized Protocol for Functional Motor Mapping Using Navigated Transcranial Magnetic Stimulation

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