This work was supported by Academy of Finland, grant numbers [259952, 276643]. We would like to thank Mikko Sams for the supervision and Enrico Glerean, Jussi Alho, Anna &#196;im&#228;l&#228; for helping with the data, Johan West&#246; for helping with the visualization as well as Marita Kattelus and Toni Auranen from Advanced Magnetic Imaging (AMI) Centre, Aalto NeuroImaging, Aalto University, Espoo, Finland for their help and support.

The protocol was approved by the Aalto University Research Ethics Committee.
1. Participant screening and preparation
<ol>
	<li>Match the different groups of recruited participants (here, heterosexual and homosexual groups) according to age, handedness, and level of education. Ensure that all participants can fully comprehend the language of the movie and feel comfortable watching it without subtitles. Alternatively, consider screening a movie in the native tongue of participants.
	<ol>
		<l...

<ol>
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Investigation of the accuracy of intuitive judgments of the trustworthiness of human faces.</a> Canadian Journal of Behavioural Science. 40 (3), 171-177 (2008).</li><li>Borkenau, P., Brecke, S., M&#246;ttig, C., Paelecke, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extraversion+is+accurately+perceived+after+a+50-ms+exposure+to+a+face.">Extraversion is accurately perceived after a 50-ms exposure to a face.</a> Journal of Research in Personality. 43 (4), 703-706 (2009).</li><li>Rule, N. 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This paper outlines the novel method for investigating the implicit brain using a post-movie subliminal measurement referred to as PMSM. In a recently published study, this method has shown that 1) implicit brain response is dynamic and 2) there is continuous learning from the social environment as well as formulation of judgment based on contextual knowledge (and not solely based on facial characteristics). Therefore, the proposed PMSM method can provide an alternative to the classical method when investigating implicit...

Recent studies show that initial social judgment becomes formulated within the first 32&#8211;100 ms of meeting another person1,2,3,4,5,6,7. Subliminal presentation of faces has been used extensively to investigate implicit biases towards different ethnic and racial groups (e.g., by presenting Caucasian American and African American faces that differ in skin color to subjects from both groups)8,9,10,11,12,13,14. However, social groups are also characterized by factors other than physical facial characteristics15.
Facial perception has been shown to be highly sensitive to contextual cues (i.e., body posture16, eye-gaze direction of the face17, a priori knowledge about the face18, visual background of the presented face19, presentation of the face separately or with other faces20). These factors can all affect facial perception. Weiser and Brosch21, in their extensive review, suggested to investigate facial perception in more naturalistic settings by ensuring that laboratory experiment is similar to real-life environments. Indeed, even simple tasks, such as recognizing people, have been shown to be more accurate when presented with video footage closer to real-life perception than when using static images22.
During the last several decades, brain imaging studies have proven that video clips can be successfully used to study realistic social perception23,24,25,26,27,28,29. The presented method is based on results from these studies and additional findings demonstrating that movie narratives can temporarily transport viewers to the world of a protagonist30. The protocol combines movie viewing with subliminal stimulus presentation as an alternative method to investigate implicit social bias formation under naturalistic conditions.
The protocol for this novel approach, post-movie subliminal measurement (PMSM), is presented here. When watching a socially engaging movie, the spectator gains knowledge about the protagonist and becomes familiar with his/her identity and world views. In contrast with other narrative art forms, movies are unique in that they present a compelling, rich, and complex story over a short time period. Furthermore, audiovisual and cinematic properties of movies synchronize brain activity across spectators23,25,29,31. Thus, it is helpful to ensure that subjects are presented with the information in a considerably similar way.
The PMSM method shows that when the face of a protagonist is presented subliminally after the movie (vs. before), implicit neural responses are successfully evoked. These responses depend on knowledge that the viewer gains about the protagonist&#39;s character with respect to his/her implicit social views. As there is a vast number of movies available that depict a variety of social characters, the PMSM method enables investigation of the brain&#39;s complex implicit views in a manner that is close to real-life social perceptions.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>3T Siemens MAGNETOM Skyra</td>
			<td>Siemens Healthcare, Erlangen, Germany</td>
			<td></td>
			<td>MRI device, using a standard 20-channel receiving head-neck coil</td>
		</tr>
		<tr>
			<td>Avid Media Composer</td>
			<td><a href="https://www.avid.com/media-composer" target="_blank">https://www.avid.com/media-composer</a></td>
			<td></td>
			<td>Video editing software used to create the stimuli.</td>
		</tr>
		<tr>
			<td>EAR-tip</td>
			<td>Etymotic Research, ER3, IL, USA</td>
			<td></td>
			<td>Earplugs compatible for MRI</td>
		</tr>
		<tr>
			<td>FSL software</td>
			<td><a href="https://www.win.ox.ac.uk/" target="_blank">https://www.win.ox.ac.uk/</a>, version 5.0.9</td>
			<td></td>
			<td>Software used to analyse the data.</td>
		</tr>
		<tr>
			<td>Panasonic PT-DZ110X projector</td>
			<td>Panasonic Corporation, Osaka, Japan</td>
			<td></td>
			<td>The stimuli were back-projected on a semitransparent screen</td>
		</tr>
		<tr>
			<td>Presentation software</td>
			<td>Neurobehavioral Systems Inc., Albany, California, USA</td>
			<td></td>
			<td>Software used to present stimuli during the fMRI scan</td>
		</tr>
		<tr>
			<td>Sensimetrics S14 insert earphones</td>
			<td>Sensimetrics Corporation, Malden, Massachusetts, USA</td>
			<td></td>
			<td>Auditory stimulation was delivered through Sensimetrics S14 insert earphones</td>
		</tr>
	</tbody>
</table>

post-movie subliminal measurement (pmsm), for investigating implicit social bias

New knowledge is continuously gained from a social environment that can influence how people respond to each other. Such responses often occur implicitly, at a subliminal perceptual level, and related brain mechanisms can be experimentally isolated by presenting the stimuli quickly. Subliminal presentation of faces that belong to different ethnicity groups, races, or gender has been shown to be successful in investigating social implicit responses. However, many implicit responses are based on knowledge previously gained about the faces (e.g., sexual orientation, political views, and socioeconomic status) and not solely on physical appearance. Here, a novel method called post-movie subliminal measurement (PMSM) is presented. When watching a socially engaging movie, a spectator gains knowledge about the protagonist and becomes familiar with his/her identity and world views. When the face of the protagonist is presented subliminally after the movie, it evokes an implicit neural response depending on what is learned about the protagonist. With a vast number of movies available, each depicting a variety of people with different identities, the PMSM method enables investigation of the brain&#39;s complex implicit biases in a manner that resembles real-life social perceptions.

Presented here are some results using PMSM from the published article by Afdile et al.34. Here, implicit bias was investigated among homosexual and heterosexual subjects (15 heterosexuals, 14 homosexuals) toward the protagonist after realizing he is homosexual in the movie, making him an &#34;ingroup&#34; to the homosexual participants and &#34;outgroup&#34; to the heterosexual participants. In line with our IAT results, this factor was shown to be significant in both groups, in which the heterose...

Watch this Scientific Journal Video about Post-Movie Subliminal Measurement PMSM, for Investigating Implicit Social Bias at JoVE.com

Post-Movie Subliminal Measurement PMSM, for Investigating Implicit Social Bias

This protocol describes the use of movies to investigate brain mechanisms underlying implicit social biases during functional magnetic resonance imaging. When the face of a protagonist is presented after a movie subliminally, it evokes an implicit response based on knowledge of the protagonist gained during the movie.

Post-Movie Subliminal Measurement (PMSM), for Investigating Implicit Social Bias

New knowledge is continuously gained from a social environment that can influence how people respond to each other. Such responses often occur implicitly, ...

Post-movie subliminal measurement can help answer key questions in understanding the neuro-processing underlying social perception, such as how contextual knowledge bias our implicit response toward others. Implicit bias is a very challenging phenomenon to investigate. By presenting stimuli for very brief periods of time, one can measure the implicit response of the brain that perceives conscious processing.In our new paradigm, we are presenting movies to experimental subjects to shape their implicit bias. This can be relevant to researchers who are interested in investigating social perception and implicit bias amongst social groups that cannot be differentiated based on their appearances, such as political groups, religious groups, groups with different sexual orientation, and more. The following protocol will be demonstrated by Marita Kattelus and Johan Westo.Upon arrival of the participants, they are briefed with the stages of the experiment, risks and usage of their data. The briefing is concluded with asking if they have any questions regarding the experiment, and that further briefing will be delivered after the experiment is done. The participant reads the briefing of the experiment and sign the consent form.The participant is asked to remove all metal objects from their clothing, or preferably change to a metal-free lab cloth. The body of the participants should be scanned using metal detection to assure no metal was forgotten to be taken off. Standard contraindications to MRIs should be respected.The participant is asked to enter the fMRI laboratory for a configuration of the video projector and audio system. Assure that the headphones are presenting the sound correctly. The participant lies down on an fMRI bed while a sample video is played to see if the picture is easily visible, and the audio level is comfortable and clear.Should any complaints arise from the participant, adjustments should be made accordingly. Connect the fMRI-compatible eye tracking system. The tracking is used to assure that the subject is attentive during the experiment, and does not fall asleep or daydream during the fMRI scan.Once the eye tracking is secured, start the calibration process to begin recording the eye movements. Inform the subject that the experiment is about to begin, and instruct the participants to relax as if they were to watch a TV program in his or her home. Remind the participant to follow the instruction slides that will guide them through the scan of the different steps.Now, go ahead and start the fMRI scan. Move the participant to a different room where additional behavior data can be collected. Once all the data is collected, debrief the participant of the actual aim of the experiment, and answer any additional questions that might arise.During the fMRI scan, the participant will be presented with a 30-minute audiovisual content, which includes instruction slides, four minutes baseline measurement, a 20-minute movie, four-minute post-movie subliminal measurement, and concluding slides. Note that the movie duration can be longer if needed. The four minutes of subliminal measurements, baseline and post-movie, are identical.The participant should not be informed of the nature or purpose at this stage, and should be presented with the following instruction. Soon, you will see a calibration clip. This clip is meant for calibrating the MRI scanner for your responses.The clip is only four minutes long, and will look like white noise on a TV screen. Please keep your eyes fixated at the mark in the center of the screen until notified otherwise. Following the instruction slide, a fixation cross will appear in the middle of the screen for two seconds, followed by the four-minute stimuli for the baseline subliminal measurement.The four-minute subliminal stimuli contain white noise virtually divided into 16 blocks of 15 seconds each. The 16 blocks of white noise contain two types of blocks, a rest block, followed by a condition block. To create the four-minute subliminal stimuli, start with a 15-second white noise clip.The white noise also serves as a masking stimulus for the face that is being flashed. Therefore, use a dynamic white noise clip that has big distortion and movements. Once you have the 15-second dynamic white noise clip, go ahead and create the 15-second condition block by inserting the 10 flashes of the protagonist's face in it.The subliminal flashes should be at a 40-millisecond duration, starting at the onset of the condition block inserted every 1, 500 millisecond. For an image of the protagonist's face, you can search for an image of the actor on the internet, or you can take a frame from the movie. When using the internet, make sure the actor looks similar to his appearance in the movie.Face images taken from the internet or the movie should have a neutral face expression, big, clear, and well-lit, as well as facing the camera. Adjust the image by centering it in the middle of the screen. Make sure there are no salient things in the background of the face.If so, go ahead and cut them out or mask them to create a neutral image. Finally, flip the image from color to black and white before inserting it. Once you have the rest block and the condition block ready, duplicate them to create the four-minute subliminal stimuli.By the end of the process, you should have a four minutes containing 16 blocks each of 15 seconds. Representative results. In the following section, we will share some of the strong and weak results using post-movie subliminal measurement from our recent study.Here we investigated implicit bias among 14 homosexual and 15 heterosexual subjects toward the protagonist of the movie Priest. The 20 minutes edited version tells the story of a Catholic priest in a struggle between his devotion to his faith, and his desire to be loved by another man. To analyze the differences between the homosexual and heterosexual group, we ran a two-way mixed effect ANOVA, which compares the differences in bold responses between the face after watching the movie, versus the baseline.In line with the hypothesis, the results revealed significantly larger bold signals in the homosexual versus heterosexual subjects in response to the face of the homosexual protagonist. Although the heterosexual group also showed significance in multiple regions including right amygdala, left insula, and bilateral central sulcus, the results were not strong enough to survive correction. This implies that in-group and out-group bias might differ in their strength, and therefore, it is advised to avoid using small sample groups when conducting group comparisons.This video shows you how our method can be set up in an fMRI study. The post-movie subliminal measurement can also be used with other newer imaging modalities. When using this method, it's important to choose a movie that is engaging, professionally executed, and relevant to your topic of investigation.Luckily, a quick search on the internet can help you find any movie depicting any social group in almost any language. This what makes movie characters a great candidate as a stimulus to investigate social perception and implicit bias.

post-movie-subliminal-measurement-pmsm-for-investigating-implicit

Research

JoVE Journal

Neuroscience

5.5K Views.  Aalto University. Post-movie subliminal measurement can help answer key questions in understanding the neuro-processing underlying social perception, such as how contextual knowledge bias our implicit response toward others. Implicit bias is a very challenging phenomenon to investigate. By presenting stimuli for very brief periods of time, one can measure the implicit response of the brain that perceives conscious processing.In our new paradigm, we are presenting movies to experimental subjects to shape...

Video: Post-Movie Subliminal Measurement PMSM, for Investigating Implicit Social Bias

This protocol describes the use of movies to investigate brain mechanisms underlying implicit social biases during functional magnetic resonance imaging. When the face of a protagonist is presented after a movie subliminally, it evokes an implicit response based on knowledge of the protagonist gained during the...

assessment of post-chronic social defeat treatment in mice: social threat-safety test, scoring, and analysis

Assessment of Post-Chronic Social Defeat Treatment in Mice: Social Threat-Safety Test, Scoring, and Analysis  

investigating social cognition in infants and adults using dense array electroencephalography (deeg)

Investigating Social Cognition in Infants and Adults Using Dense Array Electroencephalography dEEG

Dense array electroencephalography is being used increasingly to study social cognitive functions in infants and adults. Here we present an established methodology that represents a significant improvement on conventional methodologies for studying EEG in infants and...

Investigating Social Cognition in Infants and Adults Using Dense Array Electroencephalography (dEEG)

social threat-safety test uncovers psychosocial stress-related phenotypes

Author Spotlight: Unveiling Mechanisms of Stress Resilience - Significant Findings, Advancements, and Future Research

The social threat-safety test allows a simultaneous assessment of social avoidance development as a measurement of aversive conditioned learning and social threat-safety discrimination ability, both utilized to identify stress-susceptible and stress-resilient individuals within a single group of chronically socially defeated male...

Threat-Safety Test for Stress Susceptibility or Resilience in Mice Models

assessment of mouse judgment bias through an olfactory digging task

Assessment of Mouse Judgment Bias through an Olfactory Digging Task

This article provides a detailed description of a novel mouse judgment bias protocol. Evidence of this olfactory digging task's sensitivity to affective state is also demonstrated and its utility across diverse research fields is...

the social dimension of stress: experimental manipulations of social support and social identity in the trier social stress test

The Social Dimension of Stress: Experimental Manipulations of Social Support and Social Identity in the Trier Social Stress Test

Previous research on the social dimension of stress has focused on two important variables: social identity and social support. This protocol introduces an effective experimental manipulation of these two social variables and describes their implementation in a standard stress induction paradigm (Trier Social Stress Test).

aav systems and mouse models for investigating ectopic expression of neurod1 in transduced cells at subacute and chronic times post-ischemic stroke

Author Spotlight: Targeted AAV-Mediated Neurod1 Expression for Post-Stroke Neurogenesis and Functional Recovery

This protocol describes the viral-mediated ectopic expression of Neurod1 following cortical ischemic stroke. Neurod1 is delivered (1) using the Cre-Flex AAV system in wild-type mice during the subacute phase post-stroke (7 days) and (2) using a single AAV vector in conditional reporter mice during the chronic phase post-stroke (21...

AAV Systems and Mouse Models for Investigating Ectopic Expression of Neurod1 in Transduced Cells at Subacute and Chronic Times Post-Ischemic Stroke

home-based eeg hyperscanning for infant-caregiver social interactions

Author Spotlight: Capturing Infant-Caregiver Interactions Through Synchronized Multimodal Data Collection

This protocol describes how synchronized electroencephalography, electrocardiography, and behavioral recordings were captured from infant-caregiver dyads in a home...

Home-Based EEG Hyperscanning for Infant-Caregiver Social Interactions

the trier social stress test protocol for inducing psychological stress

The Trier Social Stress Test Protocol for Inducing Psychological Stress

This article describes a protocol for inducing psychological stress in participants, which enables researchers to measure psychological, physiological and neuroendocrine responses to stress within single participants or between...

the "motor" in implicit motor sequence learning: a foot-stepping serial reaction time task

The &quot;Motor&quot; in Implicit Motor Sequence Learning: A Foot-stepping Serial Reaction Time Task

We introduce the foot-stepping serial reaction time (SRT) task. This modified SRT task, complementing the classic SRT task that involves only finger-pressing movement, better approximates daily sequenced activities and allows researchers to study the dynamic processes underlying discrete response measures and disentangle the explicit process operating in implicit sequence...

The "Motor" in Implicit Motor Sequence Learning: A Foot-stepping Serial Reaction Time Task

assessment of social interaction behaviors

Assessment of Social Interaction Behaviors

Here we describe a detailed protocol for examination of sociability in mice by using Crawley's sociability and preference for social novelty test. We describe the advantages and possible applications for this procedure, including critical details important for correct interpretation of the...

Social Threat-Safety Test Uncovers Psychosocial Stress-Related Phenotypes

acquisition of  pre&#45;and post&#45;bolus 3d angiograms and intravascular pressure from a catheterized sheep for measurement of the vessel circumference

Acquisition of  Pre&#45;and Post&#45;Bolus 3D Angiograms and Intravascular Pressure from a Catheterized Sheep for Measurement of the Vessel Circumference

straightforward assay for quantification of social avoidance in drosophila melanogaster

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

Here, we present a protocol to quantify the avoidance of stressed individuals. This paradigm is powerful yet user-friendly and can be used to assess the influence of genes and environment on one kind of social interaction in Drosophila...

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

a modified trier social stress test for vulnerable mexican american adolescents

A Modified Trier Social Stress Test for Vulnerable Mexican American Adolescents

Here, we present a protocol that provoked cortisol reactivity in a vulnerable adolescent Mexican American sample utilizing a modified version of the Trier Social Stress Test (TSST). Saliva samples were collected at baseline, 15, 30, and 45 min post-TSST onset. Future research could utilize this modified TSST with vulnerable...

measuring engagement of spectators of social digital games

Measuring Engagement of Spectators of Social Digital Games

We propose a methodology that enables measuring engagement of spectators in a social digital game combining physiological and self-reported data. As this digital game involves a group of freely moving people, the experience is filmed using a synchronizing technique that links physiological data with events in the...

collection of post-mating semen from the female reproductive tract and measurement of semen liquefaction in mice

Collection of Post-mating Semen from the Female Reproductive Tract and Measurement of Semen Liquefaction in Mice

Semen liquefaction is required for sperm to be liberated from the seminal gel. This study provides the procedures for collecting semen from the female reproductive tract post-coitus, and measuring semen liquefaction...

glioblastoma relapse post-resection model for therapeutic hydrogel investigations

Glioblastoma Relapse Post-Resection Model for Therapeutic Hydrogel Investigations

The present protocol establishes a glioblastoma (GBM) relapse post-resection model using microscopy to investigate the therapeutic effect of an injectable, bioresponsive hydrogel in...

a system for tracking the dynamics of social preference behavior in small rodents

A System for Tracking the Dynamics of Social Preference Behavior in Small Rodents

Described here is a novel automated experimental system that offers an alternative to the three-chamber test and also solves several caveats. This system supplies multiple behavioral parameters that enable rigorous analysis of small rodent behavioral dynamics during the social preference and social novelty preference...

the sol braiding method for handling thick hair during transcranial magnetic stimulation: an address for potential bias in brain stimulation

Author Spotlight: Increasing TMS Accessibility Through Braiding Techniques for Diverse Hair Types

Hair type commonly seen in historically underrepresented minorities appears to interfere with transcranial magnetic stimulation (TMS). Here we describe a hair braiding method (The Sol Braiding Technique) that improves...

The Sol Braiding Method for Handling Thick Hair During Transcranial Magnetic Stimulation: An Address for Potential Bias in Brain Stimulation

Combining Behavioral Endocrinology and Experimental Economics: Testosterone and Social Decision Making

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in rodents have shown a direct link between testosterone and aggressive behavior1 and folk wisdom adapts these findings to humans, suggesting that testosterone induces antisocial, egoistic or even aggressive behavior2. However, many researchers doubt a direct testosterone-aggression link in humans, arguing instead that testosterone is primarily involved in status-related behavior3,4. As a high status can also be achieved by aggressive and antisocial means it can be difficult to distinguish between anti-social and status seeking behavior.
We therefore set up an experimental environment, in which status can only be achieved by prosocial means. In a double-blind and placebo-controlled experiment, we administered a single sublingual dose of 0.5 mg of testosterone (with a hydroxypropyl-&beta;-cyclodextrin carrier) to 121 women and investigated their social interaction behavior in an economic bargaining paradigm. Real monetary incentives are at stake in this paradigm; every player A receives a certain amount of money and has to make an offer to another player B on how to share the money. If B accepts, she gets what was offered and player A keeps the rest. If B refuses the offer, nobody gets anything. A status seeking player A is expected to avoid being rejected by behaving in a prosocial way, i.e. by making higher offers.
The results show that if expectations about the hormone are controlled for, testosterone administration leads to a significant increase in fair bargaining offers compared to placebo. The role of expectations is reflected in the fact that subjects who report that they believe to have received testosterone make lower offers than those who say they believe that they were treated with a placebo. These findings suggest that the experimental economics approach is sensitive for detecting neurobiological effects as subtle as those achieved by administration of hormones. Moreover, the findings point towards the importance of both psychosocial as well as neuroendocrine factors in determining the influence of testosterone on human social behavior.

The procedure described in this protocol shows that testosterone administration and folk beliefs about testosterone may be associated with directly opposed social behaviors.

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in ...

Combining Computer Game-Based Behavioural Experiments With High-Density EEG and Infrared Gaze Tracking

Experimental paradigms are valuable insofar as the timing and other parameters of their stimuli are well specified and controlled, and insofar as they yield data relevant to the cognitive processing that occurs under ecologically valid conditions. These two goals often are at odds, since well controlled stimuli often are too repetitive to sustain subjects' motivation. Studies employing electroencephalography (EEG) are often especially sensitive to this dilemma between ecological validity and experimental control: attaining sufficient signal-to-noise in physiological averages demands large numbers of repeated trials within lengthy recording sessions, limiting the subject pool to individuals with the ability and patience to perform a set task over and over again. This constraint severely limits researchers' ability to investigate younger populations as well as clinical populations associated with heightened anxiety or attentional abnormalities. Even adult, non-clinical subjects may not be able to achieve their typical levels of performance or cognitive engagement: an unmotivated subject for whom an experimental task is little more than a chore is not the same, behaviourally, cognitively, or neurally, as a subject who is intrinsically motivated and engaged with the task. A growing body of literature demonstrates that embedding experiments within video games may provide a way between the horns of this dilemma between experimental control and ecological validity. The narrative of a game provides a more realistic context in which tasks occur, enhancing their ecological validity (Chaytor &amp; Schmitter-Edgecombe, 2003). Moreover, this context provides motivation to complete tasks. In our game, subjects perform various missions to collect resources, fend off pirates, intercept communications or facilitate diplomatic relations. In so doing, they also perform an array of cognitive tasks, including a Posner attention-shifting paradigm (Posner, 1980), a go/no-go test of motor inhibition, a psychophysical motion coherence threshold task, the Embedded Figures Test (Witkin, 1950, 1954) and a theory-of-mind (Wimmer &amp; Perner, 1983) task. The game software automatically registers game stimuli and subjects' actions and responses in a log file, and sends event codes to synchronise with physiological data recorders. Thus the game can be combined with physiological measures such as EEG or fMRI, and with moment-to-moment tracking of gaze. Gaze tracking can verify subjects' compliance with behavioural tasks (e.g. fixation) and overt attention to experimental stimuli, and also physiological arousal as reflected in pupil dilation (Bradley et al., 2008). At great enough sampling frequencies, gaze tracking may also help assess covert attention as reflected in microsaccades - eye movements that are too small to foveate a new object, but are as rapid in onset and have the same relationship between angular distance and peak velocity as do saccades that traverse greater distances. The distribution of directions of microsaccades correlates with the (otherwise) covert direction of attention (Hafed &amp; Clark, 2002).

Procedures for recording high-density EEG and gaze data during computer game-based cognitive tasks are described. Using a video game to present cognitive tasks enhances ecological validity without sacrificing experimental control.

Experimental paradigms are valuable insofar as the timing and other parameters of their stimuli are well specified and controlled, and insofar as they ...

Brain Imaging Investigation of the Neural Correlates of Observing Virtual Social Interactions

The ability to gauge social interactions is crucial in the assessment of others&#x2019; intentions. Factors such as facial expressions and body language affect our decisions in personal and professional life alike 1. These "friend or foe" judgements are often based on first impressions, which in turn may affect our decisions to "approach or avoid". Previous studies investigating the neural correlates of social cognition tended to use static facial stimuli 2. Here, we illustrate an experimental design in which whole-body animated characters were used in conjunction with functional magnetic resonance imaging (fMRI) recordings. Fifteen participants were presented with short movie-clips of guest-host interactions in a business setting, while fMRI data were recorded; at the end of each movie, participants also provided ratings of the host behaviour. This design mimics more closely real-life situations, and hence may contribute to better understanding of the neural mechanisms of social interactions in healthy behaviour, and to gaining insight into possible causes of deficits in social behaviour in such clinical conditions as social anxiety and autism 3.

This article demonstrates an experimental design in which whole-body animated characters are used in conjunction with functional magnetic resonance imaging (fMRI) to investigate the neural correlates of observing virtual social interactions.

The ability to gauge social interactions is crucial in the assessment of others&#x2019; intentions. Factors such as facial expressions and body language ...

Social interactions are a fundamental and adaptive component of the biology of numerous species. Social recognition is critical for the structure and stability of the networks and relationships that define societies. For animals, such as mice, recognition of conspecifics may be important for maintaining social hierarchy and for mate choice 1.
A variety of neuropsychiatric disorders are characterized by disruptions in social behavior and social recognition, including depression, autism spectrum disorders, bipolar disorders, obsessive-compulsive disorders, and schizophrenia. Studies of humans as well as animal models (e.g., Drosophila melanogaster, Caenorhabditis elegans, Mus musculus, Rattus norvegicus) have identified genes involved in the regulation of social behavior 2. To assess sociability in animal models, several behavioral tests have been developed (reviewed in 3). Integrative research using animal models and appropriate tests for social behavior may lead to the development of improved treatments for social psychopathologies.
The three-chamber paradigm test known as Crawley's sociability and preference for social novelty protocol has been successfully employed to study social affiliation and social memory in several inbred and mutant mouse lines (e.g. 4-7). The main principle of this test is based on the free choice by a subject mouse to spend time in any of three box's compartments during two experimental sessions, including indirect contact with one or two mice with which it is unfamiliar. To quantitate social tendencies of the experimental mouse, the main tasks are to measure a) the time spent with a novel conspecific and b) preference for a novel vs. a familiar conspecific. Thus, the experimental design of this test allows evaluation of two critical but distinguishable aspects of social behavior, such as social affiliation/motivation, as well as social memory and novelty. "Sociability" in this case is defined as propensity to spend time with another mouse, as compared to time spent alone in an identical but empty chamber 7. "Preference for social novelty" is defined as propensity to spend time with a previously unencountered mouse rather than with a familiar mouse 7. This test provides robust results, which then must be carefully analyzed, interpreted and supported/confirmed by alternative sociability tests. In addition to specific applications, Crawley's sociability test can be included as an important component of general behavioral screen of mutant mice.

Here we describe a detailed protocol for examination of sociability in mice by using Crawley's sociability and preference for social novelty test. We describe the advantages and possible applications for this procedure, including critical details important for correct interpretation of the results.

Social interactions are a fundamental and adaptive component of the biology of numerous species. Social recognition is critical for the structure and ...

Dense array electroencephalography (dEEG), which provides a non-invasive window for measuring brain activity and a temporal resolution unsurpassed by any other current brain imaging technology1,2, is being used increasingly in the study of social cognitive functioning in infants and adults. While dEEG is enabling researchers to examine brain activity patterns with unprecedented levels of sensitivity, conventional EEG recording systems continue to face certain limitations, including 1) poor spatial resolution and source localization3,4,2) the physical discomfort for test subjects of enduring the individual application of numerous electrodes to the surface of the scalp, and 3) the complexity for researchers of learning to use multiple software packages to collect and process data. Here we present an overview of an established methodology that represents a significant improvement on conventional methodologies for studying EEG in infants and adults. Although several analytical software techniques can be used to establish indirect indices of source localization to improve the spatial resolution of dEEG, the HydroCel Geodesic Sensor Net (HCGSN) by Electrical Geodesics, Inc. (EGI), a dense sensory array that maintains equal distances among adjacent recording electrodes on all surfaces of the scalp, further enhances spatial resolution4,5,6 compared to standard dEEG systems. The sponge-based HCGSN can be applied rapidly and without scalp abrasion, making it ideal for use with adults7,8, children9,10,11, and infants12, in both research and clinical4,5,6,13,14,15 settings. This feature allows for considerable cost and time savings by decreasing the average net application time compared to other dEEG systems. Moreover, the HCGSN includes unified, seamless software applications for all phases of data, greatly simplifying the collection, processing, and analysis of dEEG data.
The HCGSN features a low-profile electrode pedestal, which, when filled with electrolyte solution, creates a sealed microenvironment and an electrode-scalp interface. In all Geodesic dEEG systems, EEG sensors detect changes in voltage originating from the participant's scalp, along with a small amount of electrical noise originating from the room environment. Electrical signals from all sensors of the Geodesic sensor net are received simultaneously by the amplifier, where they are automatically processed, packaged, and sent to the data-acquisition computer (DAC). Once received by the DAC, scalp electrical activity can be isolated from artifacts for analysis using the filtering and artifact detection tools included in the EGI software. Typically, the HCGSN can be used continuously for only up to two hours because the electrolyte solution dries out over time, gradually decreasing the quality of the scalp-electrode interface.
In the Parent-Infant Research Lab at the University of Toronto, we are using dEEG to study social cognitive processes including memory, emotion, goals, intentionality, anticipation, and executive functioning in both adult and infant participants.

Dense array electroencephalography is being used increasingly to study social cognitive functions in infants and adults. Here we present an established methodology that represents a significant improvement on conventional methodologies for studying EEG in infants and adults.

Dense array electroencephalography (dEEG), which provides a non-invasive window for measuring brain activity and a temporal resolution unsurpassed by any ...

Drosophila melanogaster is an emerging model to study different aspects of social interactions. For example, flies avoid areas previously occupied by stressed conspecifics due to an odorant released during stress known as the Drosophila stress odorant (dSO). Through the use of the T-maze apparatus, one can quantify the avoidance of the dSO by responder flies in a very affordable and robust assay. Conditions necessary to obtain a strong performance are presented here. A stressful experience is necessary for the flies to emit dSO, as well as enough emitter flies to cause a robust avoidance response to the presence of dSO. Genetic background, but not their group size, strongly altered the avoidance of the dSO by the responder flies. Canton-S and Elwood display a higher performance in avoiding the dSO than Oregon and Samarkand strains. This behavioral assay will allow identification of mechanisms underlying this social behavior, and the assessment of the influence of genes and environmental conditions on both emission and avoidance of the dSO. Such an assay can be included in batteries of simple diagnostic tests used to identify social deficiencies of mutants or environmental conditions of interest.

Here, we present a protocol to quantify the avoidance of stressed individuals. This paradigm is powerful yet user-friendly and can be used to assess the influence of genes and environment on one kind of social interaction in Drosophila melanogaster.

Drosophila melanogaster is an emerging model to study different aspects of social interactions. For example, flies avoid areas previously occupied by ...

Conditions Affecting Social Space in Drosophila melanogaster

The social space assay described here can be used to quantify social interactions of Drosophila melanogaster &#8212; or other small insects &#8212; in a straightforward manner. As we previously demonstrated 1, in a two-dimensional chamber, we first force the flies to form a tight group, subsequently allowing them to take their preferred distance from each other. After the flies have settled, we measure the distance to the closest neighbor (or social space), processing a static picture with free online software (ImageJ). The analysis of the distance to the closest neighbor allows researchers to determine the effects of genetic and environmental factors on social interaction, while controlling for potential confounding factors. Diverse factors such as climbing ability, time of day, sex, and number of flies, can modify social spacing of flies. We thus propose a series of experimental controls to mitigate these confounding effects. This assay can be used for at least two purposes. First, researchers can determine how their favorite environmental shift (such as isolation, temperature, stress or toxins) will impact social spacing 1,2. Second, researchers can dissect the genetic and neural underpinnings of this basic form of social behavior 1,3. Specifically, we used it as a diagnostic tool to study the role of orthologous genes thought to be involved in social behavior in other organisms, such as candidate genes for autism in humans 4.

Conditions Affecting Social Space in Drosophila melanogaster

The effect of genes and environment on social space of Drosophila melanogaster can be quantified through a powerful but straightforward analytical paradigm. We show here different factors that can affect this social space, and thus need to be taken into consideration when designing experiments in this paradigm.

The social space assay described here can be used to quantify social interactions of Drosophila melanogaster &#8212; or other small insects &#8212; in a ...

Culture of Neurospheres Derived from the Neurogenic Niches in Adult Prairie Voles

Neurospheres are primary cell aggregates that comprise neural stem cells and progenitor cells. These 3D structures are an excellent tool to determine the differentiation and proliferation potential of neural stem cells, as well as to generate cell lines than can be assayed over time. Also, neurospheres can create a niche (in vitro) that allows the modeling of the dynamic changing environment, such as varying growth factors, hormones, neurotransmitters, among others. Microtus ochrogaster (prairie vole) is a unique model for understanding the neurobiological basis of socio-sexual behaviors and social cognition. However, the cellular mechanisms involved in these behaviors are not well known. The protocol aims to obtain neural progenitor cells from the neurogenic niches of the adult prairie vole, which are cultured under non-adherent conditions, to generate neurospheres. The size and number of neurospheres depend on the region (subventricular zone or dentate gyrus) and sex of the prairie vole. This method is a remarkable tool to study sex-dependent differences in neurogenic niches in vitro and the neuroplasticity changes associated with social behaviors such as pair bonding and biparental care. Also, cognitive conditions that entail deficits in social interactions (autism spectrum disorders and schizophrenia) could be examined.

We established the conditions to culture neural progenitor cells from the subventricular zone and dentate gyrus of the adult brain of prairie voles, as a complementary in vitro study, to analyze the sex-dependent differences between neurogenic niches that could be part of functional plastic changes associated with social behaviors.

Neurospheres are primary cell aggregates that comprise neural stem cells and progenitor cells. These 3D structures are an excellent tool to determine the ...

Adolescent Social Stress Paradigm in C57BL&amp;#47;6 Mice for Behavioral Research

Social Defeat Stress Model for Adolescent C57BL&#47;6 Male and Female Mice

Social adversity in adolescence is prevalent and can negatively impact mental health trajectories. Modeling social stress in adolescent male and female rodents is needed to understand its effects on ongoing brain development and behavioral outcomes. The chronic social defeat stress paradigm (CSDS) has been widely used to model social stress in adult C57BL/6 male mice by leveraging on the aggressive behavior displayed by an adult male rodent to an intruder invading its territory. An advantage of this paradigm is that it allows to categorize defeated mice into resilient and susceptible groups based on their individual differences in social behavior 24 h after the last defeat session. Implementing this model in adolescent C57BL/6 mice has been challenging because adult or adolescent mice do not typically attack early adolescent male or female mice and because adolescence is a short period of life, encompassing discreet temporal windows of vulnerability. This limitation was overcome by adapting an accelerated version of the CSDS to be used for adolescent male and female mice. This 4-day stress paradigm with 2 physical attack sessions per day uses a C57BL/6 male adult to prime the CD-1 mouse for aggressiveness such that it readily attacks the male or female adolescent mouse. This model was termed accelerated social defeat stress (AcSD) for adolescent mice. Adolescent exposure to AcSD induces social avoidance 24 h later in both males and females, but only in a subset of defeated mice. This vulnerability occurs despite the number of attacks being consistent across sessions between resilient and susceptible groups. The AcSD model is short enough to allow exposure during discrete periods within adolescence, allows the segregation of mice according to the presence or absence of social avoidance behavior, and is the first model available to study social defeat stress in adolescent C57BL/6 female mice.

Author Spotlight: Understanding Adolescent Social Adversity Effects on Neurodevelopment in Mice

We developed an accelerated social defeat stress model for adolescent C57BL/6 mice, which works in both males and females and allows exposure during discrete adolescent periods. Exposure to this model induces social avoidance, but only in a subset of defeated male and female mice.

Social adversity in adolescence is prevalent and can negatively impact mental health trajectories. Modeling social stress in adolescent male and female ...

Real-Life EEG and ECG Synchronization in Infant-Caregiver Interactions

Prior hyperscanning studies that record the brain activities of caregivers and children concurrently have primarily been conducted within the confines of the laboratory, thus limiting the generalizability of results to real-life settings. Here, a comprehensive protocol for capturing synchronized electroencephalography (EEG), electrocardiography (ECG), and behavioral recordings from infant-caregiver dyads during various interactive tasks at home is proposed. This protocol demonstrates how to synchronize the different data streams and report EEG data retention rates and quality checks. Additionally, critical issues and possible solutions with respect to the experimental setup, tasks, and data collection in home settings are discussed. The protocol is not limited to infant-caregiver dyads but can be applied to various dyadic constellations. Overall, we demonstrate the flexibility of EEG hyperscanning setups, which allow experiments to be conducted outside of the laboratory to capture participants' brain activities in more ecologically valid environmental settings. Yet, movement and other types of artifacts still constrain the experimental tasks that can be performed in the home setting.

This protocol describes how synchronized electroencephalography, electrocardiography, and behavioral recordings were captured from infant-caregiver dyads in a home setting.

Prior hyperscanning studies that record the brain activities of caregivers and children concurrently have primarily been conducted within the confines of ...