This work was funded by the Excellence Initiative of the German federal state and governments (ERS Seed Fund, OPSF449). The Hitachi NIRS system was supported by a funding of the German Research Foundation DFG (INST 948/18-1 FUGG).

Prior to participation, all parents / children provided informed consent / assent. The study was approved by the ethics committee of the Medical Faculty of RWTH Aachen University.
1. Preparation before the Participant Arrives
<ol>
	<li>Prepare NIRS caps.
	<ol>
		<li>Choose the cap sizes the same size or slightly larger than the participant&#8217;s head circumference.</li>
		<li>Cut 15 holes with a diameter of approximately 15 mm each, arranged in a horizontal 3x5 grid, into the forehead area...

<ol>
	<li>Babiloni, F., Astolfi, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Social+neuroscience+and+hyperscanning+techniques:+past,+present+and+future.">Social neuroscience and hyperscanning techniques: past, present and future.</a> Neuroscience &amp; Biobehavioral Reviews. 44, 76-93 (2014).</li><li>Hari, R., Henriksson, L., Malinen, S., Parkkonen, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Centrality+of+social+interaction+in+human+brain+function.">Centrality of social interaction in human brain function.</a> Neuron. 88 (1), 181-193 (2015).</li><li>Funane, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synchronous+activity+of+two+people's+prefrontal+cortices+during+a+cooperative+task+measured+by+simultaneous+near-infrared+spectroscopy.">Synchronous activity of two people's prefrontal cortices during a cooperative task measured by simultaneous near-infrared spectroscopy.</a> Journal of Biomedical Optics. 16 (7), 077011 (2011).</li><li>Lindenberger, U., Li, S. -. C., Gruber, W., M&#252;ller, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brains+swinging+in+concert:+cortical+phase+synchronization+while+playing+guitar.">Brains swinging in concert: cortical phase synchronization while playing guitar.</a> BMC Neuroscience. 10, 22 (2009).</li><li>Jiang, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+synchronization+during+face-to-face+communication.">Neural synchronization during face-to-face communication.</a> Journal of Neuroscience. 32 (45), 16064-16069 (2012).</li><li>Dikker, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brain-to-brain+synchrony+tracks+real-world+dynamic+group+interactions+in+the+classroom.">Brain-to-brain synchrony tracks real-world dynamic group interactions in the classroom.</a> Current Biology. 27 (9), 1375-1380 (2017).</li><li>Liu, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NIRS-based+hyperscanning+reveals+inter-brain+neural+synchronization+during+cooperative+Jenga+game+with+face-to-face+communication.">NIRS-based hyperscanning reveals inter-brain neural synchronization during cooperative Jenga game with face-to-face communication.</a> Frontiers in Human Neuroscience. 10, 82 (2016).</li><li>Hoshi, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+near-infrared+spectroscopy:+current+status+and+future+prospects.">Functional near-infrared spectroscopy: current status and future prospects.</a> Journal of Biomedical Optics. 12 (6), 062106 (2007).</li><li>Lloyd-Fox, S., Blasi, A., Elwell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Illuminating+the+developing+brain:+the+past,+present+and+future+of+functional+near+infrared+spectroscopy.">Illuminating the developing brain: the past, present and future of functional near infrared spectroscopy.</a> Neuroscience &amp; Biobehavioral Reviews. 34 (3), 269-284 (2010).</li><li>Huppert, T. 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J., Diamond, S. G., Franceschini, M. A., Boas, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=HomER:+a+review+of+time-series+analysis+methods+for+near-infrared+spectroscopy+of+the+brain.">HomER: a review of time-series analysis methods for near-infrared spectroscopy of the brain.</a> Applied Optics. 48 (10), D280-D298 (2009).</li><li>Santosa, H., Zhai, X., Fishburn, F., Huppert, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+NIRS+Brain+AnalyzIR+Toolbox.">The NIRS Brain AnalyzIR Toolbox.</a> Algorithms. 11 (5), 73 (2018).</li><li>Tak, S., Uga, M., Flandin, G., Dan, I., Penny, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensor+space+group+analysis+for+fNIRS+data.">Sensor space group analysis for fNIRS data.</a> Journal of Neuroscience Methods. 264, 103-112 (2016).</li><li>Scholkmann, F., Spichtig, S., Muehlemann, T., Wolf, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+to+detect+and+reduce+movement+artifacts+in+near-infrared+imaging+using+moving+standard+deviation+and+spline+interpolation.">How to detect and reduce movement artifacts in near-infrared imaging using moving standard deviation and spline interpolation.</a> Physiological Measurement. 31 (5), 649-662 (2010).</li><li>van der Kant, A., Biro, S., Levelt, C., Huijbregts, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Negative+affect+is+related+to+reduced+differential+neural+responses+to+social+and+non-social+stimuli+in+5-to-8-month-old+infants:+a+functional+near-infrared+spectroscopy-study.">Negative affect is related to reduced differential neural responses to social and non-social stimuli in 5-to-8-month-old infants: a functional near-infrared spectroscopy-study.</a> Developmental Cognitive Neuroscience. 30, 23-30 (2018).</li><li>Bastos, A. 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In this protocol, we show how to conduct fNIRS hyperscanning experiments and one possible way to analyze brain-to-brain synchrony, measuring concentration changes of oxy-Hb and deoxy-Hb at frontal brain regions of two subjects simultaneously. FNIRS hyperscanning is relatively easy to apply: a single NIRS device is sufficient to measure brain activities of both subjects by splitting the optodes between them. Thus, no synchronization between different devices is necessary1. Moreover, since fNIRS doe...

In recent years, neuroscientists have started to study social interactions by recording the brain activities of two or more persons simultaneously, an approach termed hyperscanning1. This technique opens new opportunities to elucidate the neurobiological mechanisms underlying these interactions. To fully understand social interactions, it may not be sufficient to study single brains in isolation but rather the joint activities of brains of interacting persons2. Using different neuroimaging techniques, hyperscanning studies have shown that brain activities of interacting persons or groups synchronize, e.g., while they coordinate their actions3, make music4, communicate5, engage in classroom activities6 or cooperate7.
The article presents a protocol for conducting simultaneous recordings with functional near-infrared spectroscopy (fNIRS). Similar to functional magnetic resonance imaging (fMRI), fNIRS measures the hemodynamic response to brain activation. Changes in oxygenated and deoxygenated hemoglobin (oxy-Hb and deoxy-Hb) are calculated based on the amount of diffusively transmitted near-infrared light through tissue8. fNIRS is well suited for conducting hyperscanning experiments, especially with children, because it can be applied in less constrained and more natural settings than fMRI. Moreover, it is less prone to movement artifacts than both, fMRI and EEG9. In addition, fNIRS data can be acquired at high sampling frequencies (e.g., 10 Hz), thus it highly oversamples the relatively slow hemodynamic response and thereby potentially provides a more complete temporal picture of the brain hemodynamics10.
This protocol was developed within the study of Reindl et al.11 and has been slightly modified (in particular with respect to the channel placement and bad channel identification) more recently. The aim of the study was to examine synchronized brain activity of parent-child dyads. Using fNIRS hyperscanning, we assessed brain-to-brain synchrony in prefrontal brain areas of children (aged five to nine years) and their parents, mostly mothers, during a cooperative and a competitive computer task. Prefrontal brain regions were targeted as they had been identified as important regions for social interactive processes in previous hyperscanning studies1. The cooperative and competitive task were originally developed by Cui et al.12&#160;and recently employed by several previous studies13,14,15. For the study of Reindl et al.11, the tasks were modified to be suitable for children. Participants were instructed to either respond jointly via button presses in response to a target (cooperation) or to respond faster than the other player (competition). Each child performed each task once with the parent and once with an adult stranger of the same sex as the parent. Within each child-adult dyad, wavelet coherence was calculated for the oxy-Hb signals of corresponding channels as a measure of brain-to-brain synchrony.
This protocol describes the procedures to collect fNIRS hyperscanning data of parent and child during the cooperative and competitive game. The overall procedure, however, is not specific to this research design but is appropriate for different populations (e.g., adult strangers, romantic partners, siblings, etc.) and can be adapted for a number of different experimental tasks. This protocol also outlines one possible analytical procedure, which covers necessary and optional data analysis steps, including fNIRS data preprocessing, bad channel detection, wavelet coherence analysis and validation by random pair analysis.

<table border="0" cellpadding="0" cellspacing="0" style="width:872px;" width="873">
	<tbody>
		<tr height="40">
			<td height="40" style="height:40px;width:222px;">NIRS measurement system with probe sets and probe holder grids</td>
			<td style="width:125px;">Hitachi Medical Corporation, Tokyo, Japan</td>
			<td style="width:355px;">ETG-4000 Optical Topography System&#160;</td>
			<td style="width:171px;">The current study protocol requires an optional second adult probe set for 52 channels of measurement in total as well as two 3x5 probe holder grids.&#160;</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">raw EEG caps</td>
			<td>EASYCAP GmbH, Herrsching, Germany</td>
			<td>C-SCMS-56; C-SCMS-58</td>
			<td>Caps must be provided with holes for NIRS probes by the experimenter. Choose cap size the same size or slightly larger than participant&#39;s head circumference.</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Technical computing software</td>
			<td>The MathWorks, Inc., Natick, MA</td>
			<td>MATLAB R2014a (or later versions)</td>
			<td>Serves as base for Psychophysics Toolbox extensions (stimulus presentation), SPM for fNIRS toolbox&#160; (fNIRS data analysis), and ASToolbox (WTC computation).</td>
		</tr>
	</tbody>
</table>

conducting hyperscanning experiments with functional near-infrared spectroscopy

Concurrent brain recordings of two or more interacting persons, an approach termed hyperscanning, are gaining increasing importance for our understanding of the neurobiological underpinnings of social interactions, and possibly interpersonal relationships. Functional near-infrared spectroscopy (fNIRS) is well suited for conducting hyperscanning experiments because it measures local hemodynamic effects with a high sampling rate and, importantly, it can be applied in natural settings, not requiring strict motion restrictions. In this article, we present a protocol for conducting fNIRS hyperscanning experiments with parent-child dyads and for analyzing brain-to-brain synchrony. Furthermore, we discuss critical issues and future directions, regarding the experimental design, spatial registration of the fNIRS channels, physiological influences and data analysis methods. The described protocol is not specific to parent-child dyads, but can be applied to a variety of different dyadic constellations, such as adult strangers, romantic partners or siblings. To conclude, fNIRS hyperscanning has the potential to yield new insights into the dynamics of the ongoing social interaction, which possibly go beyond what can be studied by examining the activities of individual brains.

Representative data of one parent-child dyad during the cooperative condition are shown in Figure 1. The cooperative task consists of three 30 s rest blocks and two task blocks, with 20 trials each, presented in alternating order. In each trial, participants have to react as simultaneously as possible to a signal to earn a point11.
<img alt="Figure 1" class="xfigimg" src="/fi...

Watch this Scientific Journal Video about Conducting Hyperscanning Experiments with Functional Near-Infrared Spectroscopy at JoVE.com

Conducting Hyperscanning Experiments with Functional Near-Infrared Spectroscopy

The present protocol describes how to conduct fNIRS hyperscanning experiments and analyze brain-to-brain synchrony. Further, we discuss challenges and possible solutions.

Concurrent brain recordings of two or more interacting persons, an approach termed hyperscanning, are gaining increasing importance for our understanding ...

By measuring brain activities of two or more people simultaneously, this method can shed light on the neurobiological mechanisms underlying social interactions. Functional near-infrared spectroscopy, or fNIRS, is well suited for conducting such hyperscanning experiments because it measures brain hemodynamic response with a high sampling rate and can be applied in relatively natural settings. While this protocol shows how to conduct simultaneous fNIRS recordings of adult-child dyads, it can be adapted to different dyadic constellations such as romantic partners and to different experimental tasks.Begin by choosing a cap size that is the same size or slightly larger than the participant's head circumference. Cut 15 holes with a diameter of approximately 15 millimeters each, arranged in a horizontal three-by-five grid into the forehead area of each of two raw EEG caps. Ensure that the holes are spaced 30 millimeters from each other in any direction, that the middle column of holes is located in the center of the forehead.In order to make the caps more comfortable and minimize pressure marks, attach soft foam material at the inner side of the holder grid between the probe sockets and at the edges. Next, mount an empty three-by-five probe holder grid to each of the modified EEG caps such that the holder grid itself is placed on the inside of the cap and the holder sockets stick in the holes. Then, gently insert the probes into the appropriate holder sockets on the grid such that only the first ridge of each probe is mounted in the socket which results in one clicking sound.Open the probe set monitor window at the nearest measurement system and select two probe sets arranged in a three-by-five grid each, one for the participating child and one for the adult. Ensure that the probe arrangements of the two caps corresponds to the arrangements in the probe set window. Next, switch on the laser diodes of the nearest measurement system 30 minutes before measuring so that the system reaches a stable operating temperature.Set all the necessary options at the nearest measurement system. Make sure that the device is set to event-related measurement and that the RS232 serial input necessary for receiving triggers from the experimental paradigm is active. Then, prepare the experimental paradigm by starting the technical computing software that serves as base for the Psychophysics Toolbox extensions and setting the current directory to the folder that the paradigm is saved in.Finally, place two chin rests in front of the computer screen to prevent head movements during the experiment. Begin by escorting the two participants into the testing room and explaining the experimental set-up. Then, seat the participants next to each other in front of the computer screen.Adjust the height of the chin rests such that both participants sit comfortably. Provide instructions and administer practice trials of both the cooperative and the competitive game. Next, measure and mark the Fpz point according to the 10-20 system, which is 10%of the distance between nasion and inion on each participant's head.Then turn off the laser, and place the caps with the probes carefully on the participants'heads. Ensure that the middle probe of the bottom row is placed on Fpz and the middle probe column is aligned along the sagittal reference curve. Place the fiber strings on the holder arm attached to the nearest measurement system so that they hang loosely without contact with the participant or chair and that they do not pull on the caps.Next, push each probe further into its socket until the small white nose in the center of the top of the probe casing is visible. Then, turn the laser on again, and test the signal quality by clicking on the Auto Gain button in the probe set monitor window of the nearest measurement system. If a channel is marked in yellow, meaning insufficient signal, gently put the hair underneath the surrounding probe tip aside.Make probe adjustments, and click the Auto Gain button again to see if signal quality has improved or is marked in green. Once the signal quality is satisfactory, place a towel over the participants'hands so that they cannot see the hand movements of their respective game partner. Then, start the experiment.Then, after the experiment is complete, save the data and export the raw light intensity data as a text file by clicking on the Text File Out button. Finally, clean the probes, probe holders, and chin rests with ethnol, and wash the caps with mild detergent. As a measure of brain-to-brain synchrony, the wavelet coherence is calculated between the fNIRS signals of interacting subjects.Wavelet coherence coefficients are visualized using a color map ranging from blue, little or no coherence, to red, meaning strong or maximum coherence. Results show a strong coherence throughout the experiment in a high-frequency band until a period length of about one second, which likely results from the cardiac rhythms of the subjects. Additionally, a strong coherence appears in a lower frequency band between the two-and eight-second period length.Coherence in this low frequency range likely reflects a synchronization of brain activities of both subjects during the task. While attempting this procedure, it's important to double-check that the caps are placed correctly on the participants'heads and to ensure a good signal quality. After its development, this technique paved the way for researchers in the field of social neuroscience to explore brain-to-brain synchrony in a variety of different tasks, examining behavior correlates and associations to characteristics of the individuals and their relationship.

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9.6K visualizações.  RWTH Aachen University. Medindo as atividades cerebrais de duas ou mais pessoas simultaneamente, este método pode lançar luz sobre os mecanismos neurobiológicos subjacentes às interações sociais. Espectroscopia funcional quase infravermelha, ou fNIRS, é adequada para a realização de tais experimentos de hiperscanagem porque mede a resposta hemodinâmica cerebral com uma alta taxa de amostragem e pode ser aplicada em ambientes relativamente naturais. Embora este protocolo mostre como realizar gravações sim...