This research was supported by the National Natural Science Foundation of China (31900767), the Research Project of Shanghai Science and Technology Commission (20dz2260300), and The Fundamental Research Funds for the Central Universities.

fNIRS Hyperscanning Protocol to Analyze Inter-Brain Synchrony in Counseling

The authors have nothing to disclose.

In the present protocol, the specific steps of how to conduct an fNIRS hyperscanning experiment in the natural setting of psychological counseling and how to calculate the IBS between counselor and client, as well as how to determine the lead-lag patterns of IBS across the counseling are described. The detailed operation can help researchers repeat an fNIRS hyperscanning experiment and further research in open science. Some critical issues about the experiment design, experiment conducting, and data analysis are discusse...

In recent years, using hyperscanning techniques to explore shared brain activities during dyadic or group interactions has become a popular research direction. Researchers often employ electroencephalogram (EEG)1, functional magnetic resonance imaging (fMRI)2, or functional near-infrared spectroscopy (fNIRS)3 to monitor the neural and brain activities of multiple subjects simultaneously. The neuroscience metric of inter-brain synchrony (IBS)4 is thus introduced to quantify the degree of brain activity coupling between people with meticulous analysis of the phase and amplitude alignment of neural or hemodynamic signals across time5. IBS refers to the phenomenon where the brain activities of two or more individuals become aligned or synchronized during social interactions. This synchronization can occur in various forms, such as in the phase, frequency, or amplitude of brain oscillations6,7,8.
In the realm of naturalistic social interactions involving multiple participants, a vast array of research has illuminated the phenomenon of IBS, particularly in contexts as diverse as parent-child dynamics9, educator-student exchanges10, romantic partnerships11, and audience-performer engagements12. Notably, IBS exhibits heightened levels within intimate relationships such as parent-child and romantic partnerships, compared to interactions with strangers13,14, underscoring its sensitivity to the depth of emotional connection. Concurrently, this heightened IBS frequently coincides with enhanced collaboration efficiency and behavioral improvements, suggesting a functional role in facilitating positive social outcomes15.
In the context of counseling, the working alliance -- a pivotal construct closely tied to counseling efficacy16&#160;-- embodies a distinct interpersonal dynamic that gradually evolves between the counselor and client during the therapeutic process. At its essence, this alliance rests upon the fostering of profound emotional ties and the establishment of efficient collaborative frameworks17. Therefore, exploring IBS within counseling interactions provides a new perspective that enhances the understanding of the complexities and quality of these therapeutic relationships.
Counselor empathy, as perceived by the client, contributes to the development of a working alliance18. This indicates that the establishment of the working alliance may arise from the mutual understanding and corresponding neural activities between the counselor and the client. Empathy can be dissociated into two components: affective empathy and cognitive empathy. The inferior frontal gyrus (IFG) is implicated in affective empathy and is also associated with the neural processes underlying face-to-face communication19. The right temporal-parietal junction (rTPJ), an important part of the Theory of Mind network, is closely linked to cognitive empathy, particularly in understanding the mental states of others20. Consequently, early brain synchronization studies in counseling prioritized these two regions as regions of interest (ROIs) and identified IBS primarily in the rTPJ21. Subsequent research has thus focused predominantly on the rTPJ22. Studies have found that neural synchronization in the rTPJ between clients and therapists during counseling is significantly higher than in conversational contexts. There is a positive correlation between increased synchronous neural activity in the rTPJ and the strength of the therapeutic alliance21. The unique activity patterns in counseling may result from the in-depth exploration of emotional expressions and personal experiences. This suggests that IBS warrants further investigation within counseling. Additionally, the correlation between enhanced rTPJ activity and the strength of the working alliance indicates that IBS could serve as a neurobiological basis for assessing counseling relationships, offering a novel evaluation metric.
While these findings underscore the promising role of IBS in counselor-client dynamics, they also emphasize the need for further clarification regarding the direct causal link between brain synchronization, counseling effectiveness, and the working alliance. To advance this burgeoning field, developing standardized hyperscanning protocols and rigorous data analysis methodologies is paramount. By refining the methodological toolkit, it is possible to more precisely map the neural underpinnings of effective counseling, ultimately enhancing the quality of therapeutic interventions and their outcomes.
This article provides a protocol on how to conduct an fNIRS-based hyperscanning study and how to observe and analyze the IBS between the counselor-client dyads. fNIRS is a non-invasive imaging technique used to measure brain activity. It works by detecting changes in blood oxygenation and blood volume within the brain, which are indirect markers of neural activity. This is achieved by emitting near-infrared light into the brain and measuring the amount of light absorbed or scattered by the blood cells23. Thus, the hemodynamic/oxygenation activity is measured. Comparatively, fNIRS offers higher temporal resolution than fMRI, and it is less vulnerable to motion artifacts than EEG, rendering it well suited for studying social interactions in natural settings such as psychological counseling8.
This article also presents the specific steps of computing IBS via the method of wavelet transform coherence (WTC)24. WTC is an analytical technique that measures the relationship between two signals across different frequencies over time. It is beneficial for identifying areas of synchrony between brain regions or between participants in a study. It calculates the coherence between two time series by analyzing their cross-spectrum using wavelet transforms. To contextualize the importance of WTC, it is essential to first understand the foundational concepts of Wavelet Transform (WT)25, Coherence26, and how they converge in the framework of WTC27.
Wavelet Transform, a mathematical tool, excels at decomposing complex signals into their constituent time-frequency components, enabling the analysis of both localized changes in frequency over time and the overall frequency content of a signal27. This characteristic is particularly advantageous when studying neural activity, which is inherently non-stationary and exhibits dynamic changes across different frequencies. Coherence, on the other hand, quantifies the degree to which two signals share similar frequency components and phase relationships, serving as a metric of synchronization between them26. By combining these two concepts, WTC provides a powerful means to assess IBS, capturing both the temporal evolution and frequency specificity of neural coupling between individuals and providing insights into how different parts of the brain or brains interact dynamically throughout a task or stimulus24.
While the traditional WTC framework merely tests the correlation between the brain signals of different individuals, a method considering the directionality of the interaction between the counselor and client is presented here. There are different lead-lag patterns,where one signal consistently precedes variations in the other by a specific time interval, indicating a temporal relationship in IBS according to previous studies28,29. The IBS may not occur simultaneously between the counselor and the client during counseling. Thus, a comprehensive method is needed to explore the directionality of IBS. The method clarifies the role that counselors play throughout various phases of counseling (leading the IBS, in-phase IBS with the client, or led by the client).
This study proposes a detailed and implementable protocol based on the research question of whether IBS scores between counselors and clients can serve as potential biomarkers for assessing alliance quality or outcomes among clients with different adult attachment styles. The protocol outlines the utilization of fNIRS hyperscanning technology to investigate IBS between psychological counselors and clients within a counseling context. It provides comprehensive descriptions of the experimental procedures, precautions for each step, and subsequent data processing methods. It is anticipated that this protocol will offer valuable insights and guidance to future scholars interested in exploring IBS within the realm of psychological counseling.The specific protocol for data collection and processing is presented as follows.

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functional near-infrared spectroscopy hyperscanning study in psychological counseling

Functional Near-Infrared Spectroscopy (fNIRS) hyperscanning is an innovative technique that enables real-time monitoring of brain activity among multiple individuals engaged in social interactions. Researchers in this field quantify concurrent brain activities through the index of inter-brain synchrony (IBS). In psychological counseling research, the use of fNIRS to measure IBS has garnered attention for its potential to illuminate the dynamics of counselor-client interactions. Nevertheless, the field currently lacks a standardized protocol for precisely measuring IBS between counselors and clients, which would facilitate the revelation of real-time interaction patterns during counseling sessions. To address this need, this paper proposes a detailed standardized protocol, outlining the procedural steps for conducting fNIRS hyperscanning in psychological counseling settings, focusing on the acquisition of brain signals, calculation of IBS between counselors and clients, and analysis of lead-lag patterns of IBS throughout counseling sessions. Implementing this standardized fNIRS hyperscanning pipeline not only enhances the reproducibility and reliability of IBS measurements in psychological counseling research but also facilitates deeper insights into the neural mechanisms underlying working alliance. By integrating fNIRS hyperscanning into naturalistic counseling environments, researchers can advance understanding of how IBS correlates with counseling outcomes, potentially informing personalized approaches to mental health treatment.

The results showed that there was a marginally significant effect that the secure group had higher task-related WTC increments than the dismissing group (t = 2.50, adjusted p = 0.07) at channel 19 in the angular gyrus (ANG; see Figure 2). The WTC values at CH19 were selected for further analysis of IBS. As to the time lag effect in IBS, significantly higher late-stage counselor-led IBS was observed in the dismissing group (M = 0.04, SD = 0.07) compared to ...

Functional Near-Infrared Spectroscopy Hyperscanning Study in Psychological Counseling

The protocol aims to provide a detailed description of the methodology for conducting an fNIRS hyperscanning study in psychological counseling. This includes the preparations for the experiment, the procedure for collecting data, and the subsequent data analysis process.

Functional Near-Infrared Spectroscopy (fNIRS) hyperscanning is an innovative technique that enables real-time monitoring of brain activity among multiple ...

functional-near-infrared-spectroscopy-hyperscanning-study

Research

JoVE Journal

Neuroscience

270 Views.  East China Normal University. The protocol aims to provide a detailed description of the methodology for conducting an fNIRS hyperscanning study in psychological counseling. This includes the preparations for the experiment, the procedure for collecting data, and the subsequent data analysis process.

Video: Functional Near-Infrared Spectroscopy Hyperscanning Study in Psychological Counseling

Using MazeSuite and Functional Near Infrared Spectroscopy to Study Learning in Spatial Navigation

MazeSuite is a complete toolset to prepare, present and analyze navigational and spatial experiments1. MazeSuite can be used to design and edit adapted virtual 3D environments, track a participants' behavioral performance within the virtual environment and synchronize with external devices for physiological and neuroimaging measures, including electroencephalogram and eye tracking.
Functional near-infrared spectroscopy (fNIR) is an optical brain imaging technique that enables continuous, noninvasive, and portable monitoring of changes in cerebral blood oxygenation related to human brain functions2-7. Over the last decade fNIR is used to effectively monitor cognitive tasks such as attention, working memory and problem solving7-11. fNIR can be implemented in the form of a wearable and minimally intrusive device; it has the capacity to monitor brain activity in ecologically valid environments. 
Cognitive functions assessed through task performance involve patterns of brain activation of the prefrontal cortex (PFC) that vary from the initial novel task performance, after practice and during retention12. Using positron emission tomography (PET), Van Horn and colleagues found that regional cerebral blood flow was activated in the right frontal lobe during the encoding (i.e., initial na&iuml;ve performance) of spatial navigation of virtual mazes while there was little to no activation of the frontal regions after practice and during retention tests. Furthermore, the effects of contextual interference, a learning phenomenon related to organization of practice, are evident when individuals acquire multiple tasks under different practice schedules13,14. High contextual interference (random practice schedule) is created when the tasks to be learned are presented in a non-sequential, unpredictable order. Low contextual interference (blocked practice schedule) is created when the tasks to be learned are presented in a predictable order.
Our goal here is twofold: first to illustrate the experimental protocol design process and the use of MazeSuite, and second, to demonstrate the setup and deployment of the fNIR brain activity monitoring system using Cognitive Optical Brain Imaging (COBI) Studio software15. To illustrate our goals, a subsample from a study is reported to show the use of both MazeSuite and COBI Studio in a single experiment. The study involves the assessment of cognitive activity of the PFC during the acquisition and learning of computer maze tasks for blocked and random orders. Two right-handed adults (one male, one female) performed 315 acquisition, 30 retention and 20 transfer trials across four days. Design, implementation, data acquisition and analysis phases of the study were explained with the intention to provide a guideline for future studies.

MazeSuite is a complete toolset to prepare, present and analyze navigational and spatial experiments. Functional near-infrared spectroscopy (fNIR) is an optical brain imaging technique that enables noninvasive and portable monitoring of cerebral blood oxygenation changes. This paper summarizes collective use of MazeSuite and fNIR within a cognitive processing learning paradigm.

MazeSuite is a complete toolset to prepare, present and analyze navigational and spatial experiments1. MazeSuite can be used to design and edit adapted ...

Using Fluorescence Activated Cell Sorting to Examine Cell-Type-Specific Gene Expression in Rat Brain Tissue

The brain is comprised of four primary cell types including neurons, astrocytes, microglia and oligodendrocytes. Though they are not the most abundant cell type in the brain, neurons are the most widely studied of these cell types given their direct role in impacting behaviors. Other cell types in the brain also impact neuronal function and behavior via the signaling molecules they produce. Neuroscientists must understand the interactions between the cell types in the brain to better understand how these interactions impact neural function and disease. To date, the most common method of analyzing protein or gene expression utilizes the homogenization of whole tissue samples, usually with blood, and without regard for cell type. This approach is an informative approach for examining general changes in gene or protein expression that may influence neural function and behavior; however, this method of analysis does not lend itself to a greater understanding of cell-type-specific gene expression and the effect of cell-to-cell communication on neural function. Analysis of behavioral epigenetics has been an area of growing focus which examines how modifications of the deoxyribonucleic acid (DNA) structure impact long-term gene expression and behavior; however, this information may only be relevant if analyzed in a cell-type-specific manner given the differential lineage and thus epigenetic markers that may be present on certain genes of individual neural cell types. The Fluorescence Activated Cell Sorting (FACS) technique described below provides a simple and effective way to isolate individual neural cells for the subsequent analysis of gene expression, protein expression, or epigenetic modifications of DNA. This technique can also be modified to isolate more specific neural cell types in the brain for subsequent cell-type-specific analysis.

The goal of this protocol is to use the fluorescence activated cell sorting (FACS) technique to sort specific types of neural cells for subsequent analysis of cell-type-specific gene expression, epigenetic markers, and or protein expression.

The brain is comprised of four primary cell types including neurons, astrocytes, microglia and oligodendrocytes. Though they are not the most abundant ...

Simultaneous ex vivo Functional Testing of Two Retinas by in vivo Electroretinogram System

An In vivo electroretinogram (ERG) signal is composed of several overlapping components originating from different retinal cell types, as well as noise from extra-retinal sources. Ex vivo ERG provides an efficient method to dissect the function of retinal cells directly from an intact isolated retina of animals or donor eyes. In addition, ex vivo ERG can be used to test the efficacy and safety of potential therapeutic agents on retina tissue from animals or humans. We show here how commercially available in vivo ERG systems can be used to conduct ex vivo ERG recordings from isolated mouse retinas. We combine the light stimulation, electronic and heating units of a standard in vivo system with custom-designed specimen holder, gravity-controlled perfusion system and electromagnetic noise shielding to record low-noise ex vivo ERG signals simultaneously from two retinas with the acquisition software included in commercial in vivo systems. Further, we demonstrate how to use this method in combination with pharmacological treatments that remove specific ERG components in order to dissect the function of certain retinal cell types.

Simultaneous ex vivo Functional Testing of Two Retinas by in vivo Electroretinogram System

Ex vivo ERG can be used to record electrical activity of retinal cells directly from isolated intact retinas of animals or humans. We demonstrate here how common in vivo ERG systems can be adapted for ex vivo ERG recordings in order to dissect the electrical activity of retinal cells.

An In vivo electroretinogram (ERG) signal is composed of several overlapping components originating from different retinal cell types, as well as noise ...

Neuroimaging Field Methods Using Functional Near Infrared Spectroscopy (NIRS) Neuroimaging to Study Global Child Development: Rural Sub-Saharan Africa

Portable neuroimaging approaches provide new advances to the study of brain function and brain development with previously inaccessible populations and in remote locations. This paper shows the development of field functional Near Infrared Spectroscopy (fNIRS) imaging to the study of child language, reading, and cognitive development in a rural village setting of C&#244;te d&#39;Ivoire. Innovation in methods and the development of culturally appropriate neuroimaging protocols allow&#160;a first-time look into the brain&#39;s development and children&#39;s learning outcomes in understudied environments. This paper demonstrates protocols for transporting and setting up a mobile laboratory, discusses considerations for field versus laboratory neuroimaging, and presents a guide for developing neuroimaging consent procedures and building meaningful long-term collaborations with local government and science partners. Portable neuroimaging methods can be used to study complex child development contexts, including the impact of significant poverty and adversity on brain development. The protocol presented here has been developed for use in C&#244;te d&#39;Ivoire, the world&#39;s primary source of cocoa, and where reports of child labor in the cocoa sector are common. Yet, little is known about the impact of child labor on brain development and learning. Field neuroimaging methods have the potential to yield new insights into such urgent issues, and the development of children globally.

Neuroimaging Field Methods Using Functional Near Infrared Spectroscopy NIRS Neuroimaging to Study Global Child Development: Rural Sub-Saharan Africa

Portable neuroimaging approaches (functional Near Infrared Spectroscopy) provide advances to the study of the brain in previously inaccessible regions; here, rural C&#244;te d&#39;Ivoire. Innovation in methods and development of culturally-appropriate neuroimaging protocols permits novel study of the brain&#39;s development and children&#39;s learning outcomes in environments with significant poverty and adversity.

Portable neuroimaging approaches provide new advances to the study of brain function and brain development with previously inaccessible populations and in ...

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.

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 ...

Functional Magnetic Resonance Spectroscopy at 7 T in the Rat Barrel Cortex During Whisker Activation

Nuclear magnetic resonance (NMR) spectroscopy offers the opportunity to measure cerebral metabolite contents in vivo and noninvasively. Thanks to technological developments over the last decade and the increase in magnetic field strength, it is now possible to obtain good resolution spectra in vivo in the rat brain. Neuroenergetics (i.e., the study of brain metabolism) and, especially, metabolic interactions between the different cell types have attracted more and more interest in recent years. Among these metabolic interactions, the existence of a lactate shuttle between neurons and astrocytes is still debated. It is, thus, of great interest to perform functional proton magnetic resonance spectroscopy (1H-MRS) in a rat model of brain activation and monitor lactate. However, the methyl lactate peak overlaps lipid resonance peaks and is difficult to quantify. The protocol described below allows metabolic and lactate fluctuations to be monitored in an activated brain area. Cerebral activation is obtained by whisker stimulation and 1H-MRS is performed in the corresponding activated barrel cortex, whose area is detected using blood-oxygen-level-dependent functional magnetic resonance imaging (BOLD fMRI). All steps are fully described: the choice of anesthetics, coils, and sequences, achieving efficient whisker stimulation directly in the magnet, and data processing.

After checking by blood-oxygen-level-dependent functional magnetic resonance imaging (BOLD fMRI) that the corresponding somatosensory barrel field cortex area (called S1BF) is correctly activated, the main goal of this study is to quantify lactate content fluctuations in the activated rat brains by localized proton magnetic resonance spectroscopy (1H-MRS) at 7 T.

Nuclear magnetic resonance (NMR) spectroscopy offers the opportunity to measure cerebral metabolite contents in vivo and noninvasively. Thanks to ...

Using Near-infrared Fluorescence and High-resolution Scanning to Measure Protein Expression in the Rodent Brain

Neuroscience is the study of how cells in the brain mediate various functions. Measuring protein expression in neurons and glia is critical for the study of neuroscience as cellular function is determined by the composition and activity of cellular proteins. In this article, we describe how immunocytochemistry can be combined with near-infrared high-resolution scanning to provide a semi-quantitative measure of protein expression in distinct brain regions. This technique can be used for single or double protein expression in the same brain region. Measuring proteins in this fashion can be used to obtain a relative change in protein expression with an experimental manipulation, molecular signature of learning and memory, activity in molecular pathways, and neural activity in multiple brain regions. Using the correct proteins and statistical analysis, functional connectivity among brain regions can be determined as well. Given the ease of implementing immunocytochemistry in a laboratory, using immunocytochemistry with near-infrared high-resolution scanning can expand the ability of the neuroscientist to examine neurobiological processes at a systems level.

Here, we present a protocol that uses near-infrared dyes in conjunction with immunohistochemistry and high-resolution scanning to assay proteins in brain regions.

Neuroscience is the study of how cells in the brain mediate various functions. Measuring protein expression in neurons and glia is critical for the study ...

Conducting Concurrent Electroencephalography and Functional Near-Infrared Spectroscopy Recordings with a Flanker Task

Concurrent EEG and fNIRS recordings offer an excellent opportunity to gain a full understanding of the neural mechanism of cognitive processing by inspecting the relationship between the neural and hemodynamic signals. EEG is an electrophysiological technology that can measure the rapid neuronal activity of the cortex, whereas fNIRS relies on the hemodynamic responses to infer brain activation. The combination of EEG and fNIRS neuroimaging techniques can identify more features and reveal more information associated with the functioning of the brain. In this protocol, fused EEG-fNIRS measurements were performed for concurrent recordings of evoked-electrical potentials and hemodynamic responses during a Flanker task. In addition, the critical steps for setting up the hardware and software system as well as the procedures for data acquisition and analysis were provided and discussed in detail. It is expected that the present protocol can pave a new avenue for improving the understanding of the neural mechanisms underlying various cognitive processes by using the EEG and fNIRS signals.

The present protocol describes how to perform concurrent EEG and fNIRS recordings and how to inspect the relationship between the EEG and fNIRS data.&#160;

Concurrent EEG and fNIRS recordings offer an excellent opportunity to gain a full understanding of the neural mechanism of cognitive processing by ...

Qualitative and Comparative Cortical Activity Data Analyses from a Functional Near-Infrared Spectroscopy Experiment Applying Block Design

Neuroimaging studies play a pivotal role in the evaluation of pre- vs. post-interventional neurological conditions such as in rehabilitation and surgical treatment. Among the many neuroimaging technologies used to measure brain activity, functional near-infrared spectroscopy (fNIRS) enables the evaluation of dynamic cortical activities by measuring the local hemoglobin levels similar to functional magnetic resonance imaging (fMRI). Also, due to lesser physical restriction in fNIRS, multiple variants of sensorimotor tasks can be evaluated. Many laboratories have developed several methods for fNIRS data analysis; however, despite the fact that the general principles are the same, there is no universally standardized method. Here, we present the qualitative and comparative analytic methods of data obtained from a multi-channel fNIRS experiment using a block design. For qualitative analysis, we used a software for NIRS as a mass-univariate approach based on the generalized linear model. The NIRS-SPM analysis shows qualitative results for each session by visualizing the activated area during the task. In addition, the non-invasive three-dimensional digitizer can be used to estimate the fNIRS channel locations relative to the brain. To corroborate the NIRS-SPM findings, the amplitude of the changes in hemoglobin levels induced by the sensorimotor task can be statistically analyzed by comparing the data obtained from two different sessions (before and after intervention) of the same study subject using a multi-channel hierarchical mixed model. Our methods can be used to measure the pre- vs. post-intervention analysis in a variety of neurological disorders such as movement disorders, cerebrovascular diseases, and neuropsychiatric disorders.

We describe the analysis of continuous-wave functional near-infrared spectroscopy experiment using a block design with a sensorimotor task. To increase the reliability of the data analysis, we used the qualitative general linear model-based statistical parametric mapping and the comparative hierarchical mixed models for multi-channels.

Neuroimaging studies play a pivotal role in the evaluation of pre- vs. post-interventional neurological conditions such as in rehabilitation and surgical ...

Inter-Brain Synchrony in Open-Ended Collaborative Learning: An fNIRS-Hyperscanning Study

fNIRS hyperscanning is widely used to detect the neurobiological underpinnings of social interaction. With this technique, researchers qualify the concurrent brain activity of two or more interactive individuals with a novel index called inter-brain synchrony (IBS) (i.e., phase and/or amplitude alignment of the neuronal or hemodynamic signals across time). A protocol for conducting fNIRS hyperscanning experiments on collaborative learning dyads in a naturalistic learning environment is presented here. Further, a pipeline of analyzing IBS of oxygenated hemoglobin (Oxy-Hb) signal is explained. Specifically, the experimental design, the process of NIRS data recording, data analysis methods, and future directions are all discussed. Overall, implementing a standardized fNIRS hyperscanning pipeline is a fundamental part of second-person neuroscience. Also, this is in line with the call for open-science to aid the reproducibility of research.

The protocol for conducting fNIRS hyperscanning experiments on collaborative learning dyads in a naturalistic learning environment is outlined. Further, a pipeline to analyze the Inter-Brain Synchrony (IBS) of oxygenated hemoglobin (Oxy-Hb) signals is presented.