This project was funded by the Intramural Research Program at the National Institutes of Health Clinical Center. We acknowledge the helpful discussions with Dr. Thomas Bulea, PhD and Laurie Ohlrich, PT in refining the procedures presented in this protocol. Muyinat W. Osoba and Andrew Gravunder, MS assisted with the animations.

NOTE: This protocol was approved by the Institutional Review Board of the National Institutes of Health (ClinicalTrials.gov identifier: NCT01829724). All participants are given the opportunity to ask questions and provide informed consent prior to their participation. In consideration of changes to the hemodynamic response caused by recent use of vasodilators and vasoconstrictors, participants are asked to refrain from alcohol and caffeine for 24 hr before the experiment3.These animation videos were custom made in our laboratory, but could be recorded with other sounds or images specific to alternative research questions.
1. Set Up the Room Prior to the Participant&#8217;s Arrival.
<ol>
	<li>Calibrate the motion capture cameras relative to a laboratory&#8217;s coordinates according to the motion capture manufacturer&#8217;s specific process. Ensure that camera positions will allow recording of all markers on both the body and head of the participant during the tasks that will be tested. The calibration process ensures accuracy of the motion capture system and is standard practice for any motion laboratory. Use a ten camera system, with an approximate volume of 17 m3 where reflective markers could be identified reliably.</li>
	<li>Connect the trigger from the instruction computer to the motion capture and fNIRS computers&#8217; BNC inputs. Ensure that the trigger is connected to a mouse button, and clicking the mouse completes the circuit and sends a pulse simultaneously to the motion capture/EMG data acquisition board and to the fNIRS data acquisition board as auxillary analog inputs.</li>
	<li>Connect this mouse through the USB port to the computer that run instruction animation videos, such that starting the video will cause a voltage change simultaneously on both data acquisition systems. 
	NOTE: EMG signals are automatically synched and saved by the motion capture software, so additional synching of the EMG system is not needed.</li>
	<li>Set up the screen and projector for instructions to be shown to the participant. Remove any unnecessary items that could be distracters. Place the tripod and digital video camera where they will have full view of the participant&#8217;s movements.</li>
	<li>Verify that reflective markers are securely attached to the top of each optode in the probe.</li>
	<li>Assemble all necessary documents: consent and assent copies, clinical examination sheets, and experimental note sheets, for example.</li>
</ol>
2. Basic Measures
<ol>
	<li>After completing the informed consent process, measure and record participant&#8217;s height, weight, age, and head circumference.</li>
	<li>Administer Edinburgh Handedness Inventory10 and other clinical examinations as indicated. Record participant-reported hair and skin types.</li>
	<li>Place reflective markers on the posterior superior iliac spines (PSIS) bilaterally. Have the participant walk at their comfortable pace across the lab 3 - 5 times, and average the speed across trials to estimate their self selected walking pace.</li>
</ol>
3. Functional Near Infrared Spectroscopy (fNIRS) Setup
NOTE: This can be completed simultaneously with the setup of EMG and motion capture, if there are enough experimenters or research staff to assist, and if the participant is comfortable with several people being close to them at the same time.
<ol>
	<li>Measure the distance between the nasion (Nz) and the inion (Iz), and between the pre-auricular points on the right (Ar) and left (Al) ears. The intersection of the midpoint of these two measures is Cz, which is marked on the scalp using a washable marker.</li>
	<li>If the participant has long hair, section off small portions of the hair using braids or ponytails in order to expose the scalp where optodes will be placed.</li>
	<li>Place the fNIRS probe onto the participant&#8217;s head, taking care to align with Cz, Ar. Then move hair away from under each optode as it is placed it on the scalp. Finally, attach velcro straps to securely hold the optodes in place. 
	NOTE: In this protocol, use a cap that has one strap that goes behind the head, one that goes across the forehead, and one that goes under the chin. Optodes are anchored to this cap with Velcro on a flexible plastic ring that encircles the ear.
	<ol>
		<li>If the participant has short hair (less than approximately 2 inches in length), pull out hair between optodes with a small thin stick or plastic end of a comb.</li>
	</ol>
	</li>
	<li>Verify that all optode cables are lying flat, and that optodes are approximately perpendicular to the surface of the scalp.
	<ol>
		<li>If necessary, place a thin piece of foam under the group of optode cables to promote perpendicular alignment of the optodes.</li>
	</ol>
	</li>
	<li>Check with the participant about comfort of the probes, and adjust if needed. Instruct them to tell the experimenters if their comfort decreases at any point during the experiment.</li>
	<li>Turn on sources and check the signals.
	<ol>
		<li>In this system, ensure a signal that has an intensity of at least 80 dB and a heartbeat clearly visible in the deltaOD (change in optical density) signal, at both 690 and 830 nm wavelengths. When channels have signals not meeting these criteria, confirm that hair is not blocking the optode(s) and then adjust detector gains as needed to maximize the signal intensity. Ensure that motion capture cameras are off during this time. 
		NOTE: Other fNIRS machines may operate at wavelengths different to 690 and 830 nm; in this case, check the wavelengths most appropriate to the machine being used.</li>
	</ol>
	</li>
	<li>Add reflective markers to Nz, Iz, Ar, and Al. Ask the participant to hold still and collect approximately 2 sec&#160;of motion capture data for these and the fNIRS optode markers. Verify that all markers have been recorded, and collect additional trials as necessary. It may require the participant to change head position to improve line of sight between the cameras and the markers. Use these collected three dimensional locations during analysis for probabilistic registration of a participant&#8217;s individual structural MRI if one is available.</li>
	<li>Add a cover with several layers of black felt or other optically absorbent material on top of the fNIRS optodes to protect detectors from interference or saturation from the motion capture cameras. Ensure that the cables and front panel of the fNIRS unit are also well shielded using the same optically absorbent material.</li>
</ol>
4. Surface Electromyography (EMG) Setup
<ol>
	<li>Locate the muscle belly of each targeted muscle using anatomical landmarks, palpation during muscle contraction, and electrode placement guides11. 
	NOTE: The muscles targeted in this protocol include bilateral medial gastrocnemius, tibialis anterior, rectus femoris, vastus lateralis, biceps femoris, extensor carpi radialis, and flexor carpi radialis.</li>
	<li>Prepare for EMG electrode placement over the muscle belly by shaving, removing dead skin cells with tape, and then cleaning with an isopropyl alcohol pad, as recommended by SENIAM12 and wait for skin to dry.</li>
	<li>Place EMG electrode oriented to the direction of the muscle fibers.</li>
	<li>Wrap snugly with a self adherent wrap.</li>
	<li>Check muscle signals on the computer while performing manual muscle testing to ensure proper electrode placement, and clear visualization of signal change when the muscle is active.</li>
</ol>
5. Motion Capture Setup
<ol>
	<li>Place reflective markers at joint landmarks. These include medial and lateral malleolus, medial and lateral knee joint, anterior superior iliac spine (ASIS), posterior superior iliac spine (PSIS), radial styloid, ulnar syloid, medial humeral epicondyl, and lateral humeral epicondyl.</li>
	<li>Place 3 or more markers, or a rigid body cluster of markers, onto each segment of interest, including the foot, shank, thigh, hand, and forearm.</li>
	<li>Collect approximately 2 sec of motion capture data while the participant is standing still in a standardized position, such as standing with arms at 90&#176; shoulder flexion and 90&#176; elbow flexion. Ensure that all markers are clearly visible to the cameras.</li>
</ol>
6. Gait Task
<ol>
	<li>Have the participant transfer to the treadmill. Assist them by supporting the fNIRS optode cables and then secure the cables to the ceiling support after the patient is in position. If patient is at high risk for falls, use a body weight support harness for safety during this task.</li>
	<li>Start the treadmill, slowly building up to the measured self selected walking speed to get the participant comfortable with the set up conditions. Then slow to a stop again.</li>
	<li>Set up the animation file with the auditory feedback that will cue the participant to either rest or move. Review task instructions with the participant, telling them to remain as still and relaxed as possible during &#8220;rest&#8221; periods and to walk at the treadmill&#8217;s set speed during the &#8220;task&#8221; period, while focusing their attention to the small black circle on the screen for the duration of data acquisition.</li>
	<li>Dim the lights, and begin data acquisition on the motion capture computer and the fNIRS computer. Begin recording on the video camera.</li>
	<li>Using the mouse trigger, click the play button on the animation file associated with this task. Make sure that the trigger was received by both the motion capture and the NIRS systems.
	<ol>
		<li>Switch to an image of a black dot located in the participant&#8217;s line of sight, so that they have a focus point for the duration of the trial. 
		NOTE: The overview schematic for each trial is shown in Figure 2.</li>
	</ol>
	</li>
	<li>Monitor participant performance and provide feedback about speed, or extraneous voluntary movements as needed.</li>
	<li>At the end of the instructional animation, stop recording on the motion capture, EMG, and fNIRS systems, as well as the video camera. Give the participant an opportunity to rest or shift positions as necessary.</li>
</ol>
7. Bilateral Lower Extremity Cycling Task
<ol>
	<li>Have the participant transition to a plinth with movable back and leg support, taking care to support the fNIRS optode cables and to not bump or dislodge the motion capture markers or EMG electrodes. Have a foam seat cushion to improve comfort during the experiment.</li>
	<li>Lift the cycle frame into position and secure it to the plinth with a strap.</li>
	<li>Secure the feet into the pedals and adjust the position of the cycle as necessary to promote a comfortable and natural distance to the pedals. At the furthest point in the cycle, maintain their knee in approximately 10&#176; of flexion. 
	NOTE: At this point, the participant will be in a semi-recumbent posture, which provides some trunk support and facilitates relaxation during the rest period.</li>
	<li>Review task instructions with the participant, telling them to remain as still and relaxed as possible during &#8220;rest&#8221; periods and to cycle at approximately 60 rpm during the &#8220;task&#8221; period.</li>
	<li>Repeat steps 6.4 to 6.7. Instead of switching to an image of a dot, project the cartoon animation that will cue the participant to either rest or move through visual and auditory feedback. Maximize the movie window so that the participant is not able to monitor the time that has passed, or is remaining, in the current trial.</li>
</ol>
8. Hand Squeezing Task
<ol>
	<li>After removing the feet from the cycle and the cycle itself, place a bed table in front of the participant, making sure that participant&#8217;s arms are supported on the table at a comfortable position.</li>
	<li>Instruct participant to squeeze a soft object approximately once per sec (1 Hz) during the &#8220;task&#8221; period, and remain as relaxed as possible during &#8220;rest&#8221; periods.</li>
	<li>Repeat step 7.5.</li>
</ol>
9. Ankle Dorsiflexion Task
<ol>
	<li>Remove the bed table, and raise the foot rest portion of the plinth up to bring the feet into the participant&#8217;s view.</li>
	<li>Remove the participant&#8217;s shoe and sock, and replace foot markers in appropriate positions. Support the calf just above their ankle joint with a foam pad to allow ankle joint movement.</li>
	<li>Instruct the participant to dorsiflex their ankle approximately once per sec&#160;(1 Hz) during the &#8220;task&#8221; period, and remain as relaxed as possible during &#8220;rest&#8221; periods.</li>
	<li>Repeat step 7.5.</li>
</ol>
10. Conclusion of Protocol
<ol>
	<li>Remove the cap and inspect skin for areas of pressure or redness.</li>
	<li>Remove all reflective markers and EMG units.</li>
	<li>Thank the participant for their time and invite their input about the subjective experience of the protocol. This can be a formal questionnaire (as used by Garvey and colleagues for transcranial magnetic stimulation13), or an informal discussion to identify common sources of discomfort that could be improved in the future.</li>
</ol>

<ol>
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The authors have nothing to disclose.

Simultaneous collection of brain activity from targeted areas of the cortex and quantitative data about how a person is moving presents tremendous potential for improving our understanding of the neural control of movement, both in a typically developing population as well as those with movement disorders. There is also broad application in terms of ages and movement tasks that could be completed, as participants are not restricted to a supine position as they would be for a functional MRI. The specific equipment items a...

Neural imaging during functional tasks has become more portable and cost-efficient using non-invasive functional near-infrared spectroscopy (fNIRS) to identify areas of brain activity by measuring blood flow dynamics at the cortex. The portability of fNIRS is especially useful in the study of upright and functional tasks such as gait1, which is not possible with other technologies such as functional magnetic resonance imaging (fMRI). This capability is critical in the fields of neurology and neuroscience, and could provide new insights into mechanisms underlying movement disorders in children and adults with cerebral palsy (CP) and other neurological conditions affecting motor control. Understanding mechanisms improves the ability to design efficacious interventions to target the source of impairments and activity limitations.
Many fNIRS studies of motor tasks to date have been with a healthy population of adults where participants are instructed to perform a certain task and monitoring of task performance is limited to visual inspection. This can be sufficient for those with typical movements and a high level of engagement, but is not acceptable when studying participants with movement disorders or those who have difficulty attending to a task for extended periods of time, including typically developing children. In order to inform the analysis of brain activation in these cases, concurrent monitoring of the motor pattern that is actually completed is required.
Comprehensive reviews of fNIRS systems and usages have been presented in the literature2-5 that guide usage and help to demonstrate the accuracy and sensitivity of these systems, but technical issues in the collection, processing and interpretation of fNIRS data still remain. Color and thickness of hair affect quality of the optical signal, with dark thick hair most likely to block or distort optical transmission3,6. This is especially relevant when studying the sensorimotor areas located on the crown area of the head where hair follicle density is the greatest, and some studies report non-responders6,7. The well established International 10/20 system can be used for placement of the optodes, but particularly in the case of those with atypical brain anatomy, co-registration of optode location to a participant&#8217;s anatomical MRI is very useful if not essential to accurately interpret the results.
The use of fNIRS to assess brain activation in childhood-onset brain injury is fairly recent, but gaining traction in the area of unilateral cerebral palsy6,8,9. In consideration of the aforementioned challenges, this protocol combines fNIRS, motion capture, and electromyographic (EMG) monitoring during a number of tasks, including simple single-joint tasks as well as more complex full-body motions. Visual and auditory guidance is used to improve attention and task performance across multiple ages of participants. The goal of the protocol is to identify differences in brain activation patterns in those with unilateral and bilateral childhood-onset brain injury compared to those who are typically developing. We explore a full body movement (gait), a bilateral lower extremity multi-joint movement (cycling), and two unilateral single-joint movements (isolated ankle dorsiflexion, and isolated hand squeezing) to illustrate the variety of applications of the methods. The same or a very similar protocol could be used to study other sensory or movement disorders or other tasks of interest.
Continuous wave near infrared light was emitted and detected at 690 nm and 830 nm over the sensorimotor cortices using the fNIRS system at a rate of 50 Hz, using a custom designed source-detector configuration. EMG data were collected wirelessly at a frequency of 1,000 Hz. Reflective marker 3-D locations were collected by an optical motion capture system at a rate of 100 Hz. Two different computers handled data acquisition, one for the fNIRS and another for the motion capture and EMG. Data were synced using a trigger pulse from a third computer that corresponds to a mouse button press to start the instructional animation for each task. For all tasks except gait, instructional animations were designed to standardize participant performance using visual guidance of the pace of a task (1 Hz), represented by a cartoon animal jumping or kicking, as well as an auditory cue.

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	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Name of Reagent/ Equipment</td>
			<td style="width:107px;">Company</td>
			<td style="width:163px;">Catalog Number</td>
			<td style="width:293px;">Comments/Description</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">CW6</td>
			<td style="width:107px;">TechEn</td>
			<td style="width:163px;">http://nirsoptix.com/</td>
			<td style="width:293px;">fNIRS machine with variable number of sources and detectors, depending on the number of modules included</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">MX system with ten T40-series cameras</td>
			<td style="width:107px;">Vicon Motion Systems Ltd., Oxford, UK</td>
			<td style="width:163px;">http://www.vicon.com/System/TSeries</td>
			<td style="width:293px;">Motion capture cameras</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">reflective 4 mm markers</td>
			<td style="width:107px;">Vicon Motion Systems Ltd., Oxford, UK</td>
			<td style="width:163px;">n/a</td>
			<td style="width:293px;">Markers used by the motion capture cameras to locate fNIRS optodes, Ar, Al, Nz, and hand coordinates.</td>
		</tr>
		<tr height="113">
			<td height="113" style="height:113px;width:216px;">reflective 9.5 mm markers</td>
			<td style="width:107px;">Vicon Motion Systems Ltd., Oxford, UK</td>
			<td style="width:163px;">n/a</td>
			<td style="width:293px;">Markers used by the motion capture cameras to locate arm and leg coordinates. Clusters are used for the limb segments, and markers with offsets are uses for PSIS and Iz to improve reliability in data capture.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Trigno Wireless EMG system</td>
			<td style="width:107px;">Delsys, Inc. Natick, MA</td>
			<td style="width:163px;">http://www.delsys.com/products/wireless-emg/</td>
			<td style="width:293px;">Electromyography</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Bertec split-belt instrumented treadmill</td>
			<td style="width:107px;">Bertec Corporation, Columbus, OH</td>
			<td style="width:163px;">http://bertec.com/products/instrumented-treadmills.html</td>
			<td style="width:293px;">Treadmill</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">ZeroG body-weight support system</td>
			<td style="width:107px;">Aretech, LLC, Ashburn, VA</td>
			<td style="width:163px;">http://www.aretechllc.com/overview.html</td>
			<td style="width:293px;">Track and passive trolley used to support cables, harness can be used for patient safety during gait trials</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">3DS Max 2013</td>
			<td style="width:107px;">Autodesk, Inc., San Francisco, CA&#160;</td>
			<td style="width:163px;">http://www.autodesk.com/</td>
			<td style="width:293px;">3-D animation software used to animate animals for instructional videos</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Windows Movie Maker</td>
			<td style="width:107px;">Microsoft Corporation, Redmond, WA</td>
			<td style="width:163px;">http://windows.microsoft.com/en-us/windows-live/movie-maker</td>
			<td style="width:293px;">software used to combine animation footage with music</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Audacity</td>
			<td style="width:107px;">open source</td>
			<td style="width:163px;">http://audacity.sourceforge.net/</td>
			<td style="width:293px;">Software used to alter musical beat to appropriate cadence</td>
		</tr>
	</tbody>
</table>

functional near infrared spectroscopy of the sensory and motor brain regions with simultaneous kinematic and emg monitoring during motor tasks

There are several advantages that functional near-infrared spectroscopy (fNIRS) presents in the study of the neural control of human movement. It is relatively flexible with respect to participant positioning and allows for some head movements during tasks. Additionally, it is inexpensive, light weight, and portable, with very few contraindications to its use. This presents a unique opportunity to study functional brain activity during motor tasks in individuals who are typically developing, as well as those with movement disorders, such as cerebral palsy. An additional consideration when studying movement disorders, however, is the quality of actual movements performed and the potential for additional, unintended movements. Therefore, concurrent monitoring of both blood flow changes in the brain and actual movements of the body during testing is required for appropriate interpretation of fNIRS results. Here, we show a protocol for the combination of fNIRS with muscle and kinematic monitoring during motor tasks. We explore gait, a unilateral multi-joint movement (cycling), and two unilateral single-joint movements (isolated ankle dorsiflexion, and isolated hand squeezing). The techniques presented can be useful in studying both typical and atypical motor control, and can be modified to investigate a broad range of tasks and scientific questions.

This protocol coordinates concurrent acquisition of 3 modalities to capture brain blood flow, electrical muscle activity, and kinematic movement of joints while a participant performs motor tasks (Figure 1).
<img alt="Figure 1" src="/files/ftp_upload/52391/52391fig1highres.jpg" /> 
Figure 1.&#160;Probe location. The left portion of this figure shows the approximate locations of the sensory areas (in b...

Watch this Scientific Journal Video about Functional Near Infrared Spectroscopy of the Sensory and Motor Brain Regions with Simultaneous Kinematic and EMG Monitoring During Motor Tasks at JoVE.com

Functional Near Infrared Spectroscopy of the Sensory and Motor Brain Regions with Simultaneous Kinematic and EMG Monitoring During Motor Tasks

Monitoring brain activity during upright motor tasks is of great value when investigating the neural source of movement disorders. Here, we demonstrate a protocol that combines functional near infrared spectroscopy with continuous monitoring of muscle and kinematic activity during 4 types of motor tasks.

There are several advantages that functional near-infrared spectroscopy (fNIRS) presents in the study of the neural control of human movement. It is ...

functional-near-infrared-spectroscopy-sensory-motor-brain-regions

Research

JoVE Journal

Behavior

15.0K Views.  National Institutes of Health. Monitoring brain activity during upright motor tasks is of great value when investigating the neural source of movement disorders. Here, we demonstrate a protocol that combines functional near infrared spectroscopy with continuous monitoring of muscle and kinematic activity during 4 types of motor tasks.