Hormonal responses to brief social interactions: The role of psychosocial stress and relationship status

Nora Nickels McLean; Dario Maestripieri

doi:10.1371/journal.pone.0287153

. 2023 Jun 23;18(6):e0287153. doi: 10.1371/journal.pone.0287153

Hormonal responses to brief social interactions: The role of psychosocial stress and relationship status

Nora Nickels McLean ^1,^*, Dario Maestripieri ²

Editor: Alexander N Sokolov³

PMCID: PMC10289427 PMID: 37352264

Abstract

This study investigated the effects of psychosocial stress on hormonal responses to a social interaction with an opposite-sex individual to test the hypothesis that stress may interfere with or suppress adaptive neuroendocrine responses to courtship opportunities. Heterosexual men and women were randomly assigned prior to arrival to either a control or psychosocial stress condition (Trier Social Stress Test) and subsequently went through a social interaction test with an opposite-sex individual. Expected increases of testosterone for control participants who interacted with opposite-sex individuals were not observed, and changes in testosterone were not observed for those in the psychosocial stress condition either. However, exploratory analyses in control participants showed main and interaction effects of relationship status were significant for both cortisol and testosterone. Specifically, single individuals showed higher levels of cortisol compared to those in a relationship, and single individuals showed significantly higher concentrations of cortisol after a social interaction when compared to individuals who were in a relationship. For testosterone, only individuals in a relationship decreased in testosterone following the social interaction. This study suggests that relationship status and psychosocial stress may be important variables moderating the relationship between an ecological cue of a potential courtship opportunity and subsequent adaptive physiological responses.

Introduction

Across many vertebrate species, testosterone regulates energy distribution and promotes investment into mating efforts in a variety of ways [1–4]. The “challenge hypothesis” [5], which has been used a conceptual framework to study the relationship between rises in androgen levels and male socio-sexual and aggressive behaviors in mating contexts, has been supported through evidence in multiple nonhuman primate and other vertebrate species [e.g., 6–8]. In humans, the importance of testosterone for male mating effort has been documented by many studies reporting on both baseline testosterone levels and testosterone responses to social stimuli [2,9–11]. The role of testosterone as a signal to coordinate behavioral investment in courtship and mate pursuit has more recently been explored in the context of the initiation of human romantic relationships [9]. Specifically, numerous studies have examined the reactive testosterone responses of heterosexual male individuals to naturalistic social interactions with female individuals [12–20].

Roney and colleagues [17] were some of the first to test for hormonal reactions of human men to brief social encounters with opposite-sex individuals, considered to be potential mating partners. They found that salivary testosterone of young men increased significantly over baseline levels after engaging in a short conversation with a young woman (this effect was only evident, or was most pronounced, in single men than in men in stable romantic relationships; see below) but did not increase significantly over baseline levels after engaging in short conversation with another young man [17,18]. Consistent with the notion that dominance may also play a role in fluctuations of testosterone during brief encounters with a potential mate, van der Meij et al. [20] found that salivary testosterone increases after brief interactions with women were most evident in men with aggressive dominant personalities, men who were not involved in committed, romantic relationships, and men who had been sexually inactive for over a month. Therefore, individual differences in personality, relationship status, and sexual motivation may affect testosterone reactivity that occur in response to courtship opportunities.

Lopez et al. [21] replicated in women the findings of Roney and colleagues [17,18]. Female participants showed significant reactive increases in testosterone after viewing a movie clip of a courtship situation with an attractive man and a young woman [21]. It is possible that female-female competition for potential courtship opportunities may induce a similar testosterone increase in women to aid in self-confidence, to preserve dominance and status, and to emphasize motivation to affiliate with potential mates. Although most previous studies of testosterone and competition have been conducted with men, there is some evidence that similar effects can be observed in women as well [22,23]. Researchers are currently calling for further research on the endocrinology of competition and courtship in women and are emphasizing the importance of including female participants in study designs [24,25]. The purpose of the current study includes observing potential short-term androgen increases in both men and women.

All social evaluative interactions involve stressful components, and courtship in particular may cause significant apprehension during courtship interactions [26–28]. We know that human courtship behavior did not evolve in a stress-free environment, and that any kind of social evaluation places pressures upon and threatens one’s own self perceptions, through dampening self-motivation, self-confidence, or perception of social status. Further, the “dual hormone hypothesis” postulates that testosterone’s association with status-relevant and socio-sexual behavior depends on cortisol, a hormone released during physical and psychological stress [29–32]. Therefore, it is important that we consider how stress may affect courtship behavior in either adaptive or maladaptive ways. The purpose of the current study also includes observing the effects of stress on androgen reactivity to potential mating scenarios.

Cortisol is a glucocorticoid hormone released by the hypothalamic-pituitary-adrenal (HPA) axis in response to stress as part of its broader role as a physiological regulator of blood pressure, metabolism, and energy mobilization [33,34]. Much research places the focus of cortisol increase as a coping mechanism to aversive physical or psychological conditions and as an adaptive function to divert energy and suppress unessential processes [34]. Importantly, cortisol responses are also elicited when one is faced with a psychological threat to one’s self esteem and when psychological well-being is threatened [35]. We could potentially expect courtship apprehension to trigger a cortisol increase in addition to a testosterone increase that may prove to alter courtship behaviors.

There is already some indirect evidence for the role of the HPA axis in human courtship, as displayed by cortisol increases that could reflect anxiety or apprehension before or during opportunities for courtship [18,19,36,37]. In multiple experiments, Roney et al. [18] found that changes in cortisol from baseline were significantly greater among male participants who interacted with female individuals relative to male participants in control conditions. Lower baseline cortisol concentrations have also been shown to predict larger testosterone responses to interactions with potential female mates [15], consistent with the dual hormone hypothesis. Similarly, van der Meij and colleagues [36] found that cortisol levels of men increased when they interacted with a female individual whom they reported as attractive. Beyond a one-on-one social interaction, cortisol has been found to be sensitive to the sex composition of the environment [38] and to anticipatory social exchanges where participants perceive potential interactions as occurring with a desirable dating partner [39].

It remains unclear overall how stress during courtship and subsequent increases in cortisol may interfere with or trigger other physiological and psychological responses to courtship [see 40, for different effects of psychosocial stress on cooperative behavior in male and female participants]. For example, an increase in cortisol may subdue or promote appropriate courtship behaviors based on the profile of testosterone reactivity, as predicted by the dual hormone hypothesis. If testosterone shows acute increases in response to a courtship opportunity, despite a simultaneous increase in cortisol, courtship behaviors may not be hindered in a maladaptive way. However, if a cortisol increase is accompanied by a testosterone decrease, stress may prove to interfere with subsequent motivation, self-confidence, and affiliative behaviors.

It is currently uncertain whether cortisol responses are generally associated with specific courtship behavior patterns, or whether the association is an effect that is strictly mediated by motivation, cognition, or individual differences in personality or relationship status. Further, anxiety itself may drive the physiological responses of testosterone, in addition to its effects on cortisol. The effects of stress on testosterone may depend specifically on the type of social stress induced and may mediate the relationship between individual differences and behavior. For example, Crowley et al. [41] found that testosterone mediated the relationship between uncertainty and the amount of disclosure between romantic partners.

The goal of the present study was to investigate testosterone and cortisol responses to social interactions with opposite-sex individuals in both men and women, with or without a prior exposure to psychosocial stress. Primary a priori hypotheses involved the effects of social interactions with opposite-sex individuals on testosterone in men and women, and we explored these hypotheses using two different psychosocial tasks. The first was a non-stressful social interaction task with an opposite-sex individual, which all participants experienced. The second was a psychosocial stress task that half of the participants experienced prior to their non-stressful social interaction with an opposite-sex individual. First, we predicted that we would replicate findings of increases in testosterone for both male and female participants during brief interactions with opposite-sex individuals. This increase would be expected specifically for individuals who have not gone through a psychosocial stress manipulation, replicating past studies that have found testosterone increases for men and exploring the possibility that this testosterone increase may be seen in both men and women. Second, we predicted that a psychosocial stress manipulation could alter a testosterone response in either of the following ways: 1) if cortisol increases dramatically due to psychosocial stress, testosterone increases could be dampened, and 2) if cortisol reactivity to stress is low, this may be associated with higher increases in testosterone. In addition to these main research questions, we planned exploratory analyses on whether testosterone increases would be most apparent in individuals whose motivations are in line with potential courtship behaviors (e.g., single individuals), due to lack of direct evidence for such findings in humans. Cortisol responses to social interactions as a function of relationship status are also considered exploratory, given mixed findings in nonhuman literature and the lack of direct evidence in studies using both men and women.

Method

Participants

156 participants (62 men, age: M = 22.56, SD = 4.37; 94 women, age: M = 21.89, SD = 3.37) were recruited from the University of Chicago campus and surrounding area through fliers, UChicago Marketplace, and a human subject recruitment website. Participants signed an electronic copy of the consent form prior to arrival and were paid $20 after completion of the study.

Experimental procedure

In this study, the eligibility criteria for participant recruitment were age (between 18 and 35 years) and heterosexual orientation. All experimental procedures were approved by the Social Science Institutional Review Board at the University of Chicago. All experimental procedures took place between 11:30 AM and 5:30 PM. Participants always interacted with an experimenter, or “greeter”, of the same sex throughout the entire experimental session. Upon arrival to the research building, participants were taken by a “greeter” to the testing room, where they completed questionnaires for 20 minutes. An initial demographic survey asked information about participants’ age, ethnicity, sexual orientation, socioeconomic status, marital or relationship status (single or in a relationship), etc. At the end of this period, they provided a baseline saliva sample. They then either took part in the Trier Social Stress Test or sat in a room doing nothing for a similar period of time as a control condition. Another saliva sample was collected after the TSST or the control condition (post-treatment), approximately fifteen minutes after the start of the TSST or control condition. Approximately ten to fifteen minutes after the TSST or control condition had ended, participants went through a brief social interaction task with a research assistant of the opposite sex. A final saliva sample (post-social interaction) was collected after the social interaction task. Upon completion of all procedures, participants were fully debriefed and given compensation.

Trier social stress test

The Trier Social Stress Test (TSST) [42] is a broadly used, standardized task that is used to study hormonal responses to mild psychosocial stress in a laboratory setting. In the current study, the experimenter explained to each participant that he or she would be giving a 5-minute presentation about himself or herself for a mock job interview. Each presentation took place in front of a “selection committee” composed of two unfamiliar judges trained to maintain neutral facial expressions and provide no positive feedback to the participant. Each participant was informed that he or she must keep speaking for 5 minutes and that the presentation would be video-recorded for subsequent analyses of content and non-verbal behavior. If the participant ever stopped speaking before the 5 minutes were up, the judges waited in silence for the participant to resume or otherwise prompted him or her to continue. If the participant again stopped speaking, one of the judges asked one of several standardized questions (e.g., “What do you think about teamwork?”). Upon completing the 5-min speech, the judges asked each participant to perform a difficult arithmetic calculation (i.e., serially subtracting the number 17 from 2,023) out loud for another 5 minutes or until he or she reached zero. Anytime the participant made a mistake, he or she was notified and asked to restart from the beginning. After this task, the judges thanked the participant and left the room.

Although the “greeter” who interacted with the participant was always the same sex as the participant, the TSST judges were either of the same or the opposite sex as the participant, in alternation. Therefore, female participants were assigned to interact with either two same-sex TSST judges (a female “talking judge” and a female “timing judge”), or two opposite-sex TSST judges (a male “talking judge” and a male “timing judge”). Likewise, male participants were assigned to interact with either two same-sex TSST judges (a male “talking judge” and a male “timing judge”), or two opposite-sex judges (a female “talking judge” and a female “timing judge”). The semi-random assignment of the judges was based on availability of the research assistants but was also counter-balanced as much as possible. Over the fourteen months of data collection, judges and greeters were assigned based on availability from a team of eighteen different undergraduate research assistants (age range 18–27; 46.7% female).

Participants who were assigned to the control condition and who did not participate in the TSST simply sat by themselves for 10 minutes until their original experimental “greeter” returned to let them know they could continue moving forward in the study.

Social interaction task

Following the control/TSST condition, all participants partook in a social interaction task, where they interacted with an opposite-sex research assistant whom they had not encountered yet at that point in the session. The social interaction task that was used in this experiment was adapted from brief social interaction tasks that have been used in several research studies in which a social interaction involving a research assistant posed as either another participant or experimenter led to physiological and behavioral changes [17,18]. In our study, the experimental “greeter” let the participant know that they needed approximately five or ten minutes to pass before moving on to the next part of the study, and that the participant was free to relax until the experimenter returned. Several minutes after the departure of the “greeter”, an opposite-sex researcher entered the room and introduced himself or herself as a research assistant there to collect data off of a digital video camera (earlier in the session, this video camera was used to collect a digital photograph of every participant, as well as used to record the TSST session for participants assigned to the TSST condition). Chairs were arranged in the room such that the participants always sat directly across from the researcher with a small conference table positioned between them. Research assistants then attempted to engage in natural, friendly conversation, while simultaneously uploading data from the digital video camera onto a computer or hard drive. The research assistants were free to use whatever means of engaging in conversation seemed natural to them. Scripts or specific prompts were not used to avoid interactions seeming excessively artificial. Conversations lasted seven minutes, at which point the experimenter re-entered the room and interrupted the research assistant and participant to seemingly complete the rest of the study protocol. As with greeters and judges, conversation partners were assigned based on availability from a team of eighteen different undergraduate research assistants (age range 18–27; 46.7% female). Participants on average rated the opposite-sex research assistant slightly above (i.e., slightly above a rating of 4) or slightly below (i.e., slightly below a rating of 4) the midpoints of the seven-point scales that assessed perception of physical attractiveness (M = 4.33, SD = 1.60), desirability as a short-term romantic partner (M = 3.32, SD = 1.78), and desirability as a long-term romantic partner (M = 3.34, SD = 1.70). Such ratings suggest that the participants did not have particularly strong interest in the opposite-sex research assistant but did rate individuals as above average in terms of attractiveness.

Saliva sample collection and hormonal assays

All saliva samples were collected between 12:00 PM and 5:00 PM, as previous studies have shown that afternoon cortisol and testosterone levels, although lower than morning levels, are more stable and therefore better suited for studies of social endocrinology [35,43–46]. Strong circadian variation is observed for both cortisol and testosterone, and by conducting testing at just one time of day (i.e., in the afternoon), researchers can avoid some of the variance determined through circadian fluctuation [44]. In terms of sampling, intervals of 15 minutes are commonly used due to the time it takes for steroid hormones to transfer from blood to saliva after negative feedback loop release [44]. Saliva was collected by passive drool into plastic tubes. Saliva samples were stored in a refrigerator at -20°F. Samples were assayed for testosterone and cortisol concentrations using ELISA kits purchased from Salimetrics. For both cortisol and testosterone assays, samples competed with hormones conjugated to horseradish peroxidase, and assay sensitivity was <0.007 ug/dL and 1 pg/mL, respectively. Saliva sample concentrations were calculated based on kit standards using a 4-parameter nonlinear regression curve fit. For cortisol, the average inter-assay CV based on concentration was 4.85% (3.54% for high controls; 6.17% for low controls), and the intra-assay CV based on concentration was 7.15%. For testosterone, the inter-assay CV based on concentration was 6.63% (4.86% for high controls; 8.40% for low controls), and the intra-assay CV based on concentration was 5.38%. CVs were calculated from the concentrations rather than raw optical densities. Specifically, the inter-assay CV was calculated using the mean values for the quality high and low controls on each plate, and the intra-assay CV was an average value calculated from the individual CVs from all the plate duplicates. The cutoff for reanalysis used was 10%; any duplicate above this cutoff were removed and reanalyzed. In the overall sample, baseline cortisol and testosterone levels were in line with assay protocol salivary example PM range norms.

Data analyses

Four individuals did not complete the full protocol due to researcher error, participant withdrawal from the study, or computer error, and therefore were not included in data analyses. Two more individuals were excluded from all hormonal analyses due to having baseline or delta hormonal concentrations over three standard deviations away from the mean. One participant was excluded from all hormonal analyses due to saliva samples being heavily contaminated with blood. After excluding these seven individuals, data were analyzed for a total of 149 individuals, of whom 76 (46 women, 30 men) underwent the TSST stress manipulation and 73 (47 women, 26 men) were in the control condition. Of all participants, 81 indicated their relationship status as single, and 68 indicated their relationship status as in a relationship at the time of the study. Of all TSST participants, 40 were single and 36 were in a relationship; of all control participants; 41 were single and 32 were in a relationship.

All statistical analyses were carried out with jamovi software (Version 2.2.5.0) [47]. When hormonal data were not normally distributed, they were log transformed. When sphericity assumptions were violated, Greenhouse-Geisser corrected p-values were reported. Alpha was set at 0.05 and adjusted for multiple comparisons where necessary.

Results

Manipulation check

To confirm the effects of the TSST on participants’ stress levels, the effect of the TSST manipulation on physiological stress (cortisol) was tested with a 2 (treatment group: control vs. TSST) x 3 (timepoint: baseline, post-treatment condition, post-social interaction) mixed model, with timepoint as a repeated measure and cortisol concentration as the outcome variable. A Greenhouse-Geisser correction was applied since the assumption of sphericity was not met. An expected interaction effect between treatment group and timepoint was significant [F(1.71,251.13) = 14.58, p<0.001, η²_P = 0.09 (a moderate effect); Fig 1]. Specifically, individuals who underwent the TSST had higher cortisol concentrations post-treatment (t = -4.48, p<0.001) and post-social interaction (t = -3.73, p = 0.004).

Fig 1 — Salivary cortisol concentrations (measured at baseline, post-treatment, and post-social interaction) depicted by experimental condition. Values are mean ± SEM.

Testosterone responses to social interaction

The effects of the social interaction and TSST manipulation on testosterone was tested with a 2 (timepoint: pre-social interaction and post-social interaction) x 2 (treatment group: control vs. TSST) mixed model, with timepoint as a repeated measure and testosterone concentration as the outcome variable. Unlike what we hypothesized, there was no significant interaction effect between timepoint and treatment group [F(1,147) = 0.08, p = 0.77; Fig 2]. Specifically, there were no changes in testosterone concentrations for participants who were not exposed to psychosocial stress before a social interaction with an opposite-sex research assistant, and there were no changes in testosterone concentrations for participants who were exposed to psychosocial stress before a social interaction with an opposite-sex research assistant. No main effects for timepoint [F(1,147) = 0.58, p = 0.45] or treatment [F(1,147) = 0.003, p = 0.95] were found.

Fig 2 — Salivary testosterone concentrations (measured at pre-social interaction and post-social interaction) depicted by experimental condition. Values are mean ± SEM.

Exploratory analyses of relationship status

To explore how relationship status relates to hormone reactivity, subgroup analyses were performed separately in control and TSST participants. First, the effect of relationship status on testosterone responses in control participants was tested with a 2 (relationship status: single vs. in a relationship) x 2 (timepoint: pre-social interaction and post-social interaction) mixed model, with timepoint as a repeated measure and testosterone concentration as the outcome variable. No main effects of relationship status [F(1,71) = 1.25, p = 0.27] or timepoint [F(1,71) = 1.39, p = 0.25] were found. An interaction effect between relationship status and timepoint was found [F(1,71) = 11.45, p = 0.001, η²_P = 0.14 (a large effect); Fig 3], such that participants in a relationship had lower concentrations of testosterone after the social interaction when compared to prior to the social interaction (t = 3.04, p = 0.02). However, the hypothesized testosterone increase in single individuals after exposure to social interaction was not observed.

Fig 3 — Salivary testosterone concentrations (measured at pre-social interaction and post-social interaction) depicted by relationship status. Values are mean ± SEM.

The effect of relationship status on cortisol responses in control participants was tested with a 2 (relationship status: single vs. in a relationship) x 2 (timepoint: pre-social interaction and post-social interaction) mixed model, with saliva sample as a repeated measure and cortisol concentration as the outcome variable. An interaction effect between relationship status and timepoint was found [F(1,71) = 10.69, p = 0.001, η²_P = 0.13 (a medium effect); Fig 4]. Specifically, participants in a relationship had lower concentrations of cortisol after the social interaction when compared to prior to the social interaction (t = 3.13, p = 0.02), and participants in a relationship had lower concentrations of cortisol after the social interaction when compared to single participants after the social interaction (t = 3.33, p = 0.008). A main effect of relationship status was also found [F(1,71) = 5.67, p = 0.02, η²_P = 0.07 (a medium effect)], such that control participants in a relationship had lower levels of cortisol overall when compared to single participants (t = 2.38, p = 0.02). No main effect of timepoint was found [F(1,71) = 2.02, p = 0.16].

Fig 4 — Salivary testosterone concentrations (measured at pre-social interaction and post-social interaction) depicted by relationship status. Values are mean ± SEM.

Exploratory analyses of relationship status were also conducted for participants who went through the TSST stress condition. The effect of relationship status on testosterone responses in TSST participants was tested with a 2 (relationship status: single vs. in a relationship) x 2 (timepoint: pre-social interaction and post-social interaction) mixed model, with timepoint as a repeated measure and testosterone concentration as the outcome variable. No significant main effects of timepoint [F(1,74) = 0.063, p = 0.803] or relationship status [F(1,74) = 0.00, p = 0.99] were found. The interaction effect between timepoint and relationship status was also not significant [F(1,74) = 1.54, p = 0.22]. For cortisol, an expected significant main effect of timepoint was found in TSST participants [F(1,74) = 13.58, p<0.001, η²_P = 0.16 (a large effect)], such that participants post-social interaction had lower cortisol concentrations when compared to participants pre-social interaction (t = 3.69, p<0.001); this decrease was expected, as it was likely that participants’ cortisol concentrations were decreasing after the post-TSST increase. Similar to subgroup analyses with control participants, a significant main effect of relationship status was observed [F(1,74) = 5.17, p = 0.03, η²_P = 0.07 (a medium effect)], such that TSST participants in a relationship had lower levels of cortisol overall when compared to single participants (t = 2.27, p = 0.03).

Discussion

The present study investigated hormonal responses to brief social interactions following social evaluative stressors. We hypothesized that individuals would show increases in testosterone following brief social interactions and that exposure to prior social evaluative threat and relationship status would moderate these increases. However, among all control male and female participants, we did not find significant increases in testosterone after taking part in a brief conversation with a researcher of the opposite sex. Further, the psychosocial stress treatment did not moderate or cause changes in testosterone following the social interaction. Exploratory subgroup analyses did show a difference in testosterone at differing time points for individuals who were in a relationship for control participants. Specifically, individuals in a relationship had significantly lower testosterone concentrations after the social interaction compared to testosterone concentrations prior to the interaction. These results do not replicate those of previous studies showing that in single men, verbal interactions with women (of less than five or ten minutes) can causally induce increases in testosterone [17,20]. However, the results may be consistent with past findings that have shown singles to have higher testosterone concentrations [48] and with findings that testosterone concentrations are higher after a social interaction for those who are single when compared to those who are in a relationship [17,20]. Overall, these results only partly support the challenge hypothesis. General increases in testosterone were not observed in men and women during psychosocial encounters, yet it is possible that the significant findings via exploratory analyses focusing on relationship status provide evidence that motivational differences could lead to differences in testosterone reactivity that correspond with appropriate socio-sexual behavior.

Similar to testosterone, exploratory subgroup analyses showed differences in cortisol between single individuals and individuals in a relationship for control participants. Specifically, singles showed significantly higher cortisol concentrations when compared to individuals who were in a relationship overall, and single individuals had higher concentrations of cortisol after the social interaction when compared to individuals in a relationship. These results are partly consistent with those of previous studies in suggesting that cortisol increases could reflect anxiety or apprehension before or during opportunities for courtship [18,19,36]. For example, Roney et al. [18] found that changes in cortisol from baseline were significantly greater among male participants who interacted with women relative to men in control conditions. Specifically, Roney et al. [18] and van der Meij et al. [36] found that singles showed greater hormonal reactivity when compared to individuals in a relationship. These results are also consistent with past findings that have shown singles to have higher cortisol concentrations [49].

Further, these results are consistent with an article by Zilioli and Bird [50], who after reviewing the literature on testosterone reactivity (primarily in human men), concluded that both motivational factors and situational factors have the ability to influence the relationship between evolutionarily salient social contexts, such as an interaction with a potential mate, and physiological reactivity to these contexts [50]. Differences in relationship status could, at least in part, reflect differences in socio-sexual motivation. A situational factor, which the authors define as an external factor outside of the control of the individual, could influence the relationship between ecological cue and physiological response [50]. In our study, psychosocial stress may have interfered with the physiological response to social stimuli in control participants, which explains why the significant changes in testosterone in individuals in a relationship were no longer observed in TSST participants. This possible interference may also be evidence of the dual hormone hypothesis; an increase in cortisol may have subdued the testosterone reactivity observed in control participants in a relationship. However, our results did not support the expected evidence of the dual hormone hypothesis whereby changes in cortisol altered testosterone reactivity meant to promote appropriate courtship behavior, as expected increases in testosterone were not observed in control participants. Future research on the relation between stress and courtship is needed, especially in light of recent studies reporting anxiety-related reactions during first encounters with individual perceived as attractive [51].

Our interpretation of the results is tentative, and we acknowledge that our study has some methodological limitations. First, previous laboratory studies of human courtship have used highly attractive research assistants as social interaction partners, who engaged in friendlier, more deliberately flirtatious conversations [19], and some field studies often increase validity by measuring hormones before, during, and after dating events [36]. As this study chose to keep the social interaction more neutral, this may have resulted in fewer participants interpreting the encounter as a potential courtship opportunity, and we did not ask participants if they were given the impression to flirt from the research assistant. This difference may in part account for the discrepancy between these results and those of previous studies. Further, we did not use a control condition in which participants either engaged in conversation with a same-sex social interaction partner or simply did not participate in conversation at all. Second, although it remains crucially important for future research to focus on context-dependent acute hormonal changes in women, recent work has pointed out that it is challenging to measure variability in testosterone concentrations in women using standard enzyme immunoassays (EIAs), which may tend to inflate estimates of lower concentration estimates in women [52]. Despite the ease and cost-effectiveness of EIAs, we encourage further research using liquid chromatography mass spectrometry to measure hormone concentrations, as this methodology may be free from some of the limitations involved with EIAs [52]. Third, budgetary restrictions limited the number of hormone samples taken throughout the experimental procedure. Only one baseline hormonal assessment and one test assessment after the TSST / control were measured, and allowing time for hormone levels to return back to baseline after the experimental treatment could have simplified the interpretation of our results.

Overall, this study highlights the importance of individual differences in relationship status and situational variables as factors in moderating hormonal reactivity to a potential courtship interaction. It appears that psychosocial stress, while not affecting testosterone reactivity during a social interaction directly, may play a role in altering how men and women react to a social interaction interpreted as a potential courtship opportunity. While the exact mechanism remains unclear, it is possible that psychosocial stress may suppress the adaptive physiological response (increase in testosterone) in different ways dependent on relationship status for individuals who are engaged in a courtship. Future studies should continue to focus on the potential moderating factors, whether biological, sociological, or psychological in nature, as there is growing evidence that these factors have the potential to influence the relationship between evolutionarily salient social contexts (e.g., interaction with mates) and adaptive neuroendocrine responses.

Supporting information

S1 File. ZIP file containing data analysis script (.omt files) for three sets of analyses.

(ZIP)

Click here for additional data file.^{(356.2KB, zip)}

Acknowledgments

We thank Jim Roney for his helpful feedback on this manuscript. We thank Mint Poonpatanapricha, Coltan Scrivner Cameron Miller, Yanitza Roman, Kay Yang, Melanie Sykes, Alex Portee, Ian McCann, Jonah Lowenstein, Joseph Wiltzer, John Luo, Justin Pan, Emil Qiu, Antonia Theodosakis, and Kat Pervova for their assistance with data acquisition.

Data Availability

Data cannot be shared publicly because our study protocol, which was approved by the Social and Behavioral Sciences Institutional Review Board of the University of Chicago (IRB #12-1251), does not provide for sharing de-identified participant data outside the research team, we are not able to publicly share the data. This is because data contain potentially identifying information. Data are available from the Social and Behavioral Sciences Institutional Review Board Ethics Committee of the University of Chicago (contact via 773-702-2915, or sbs-irb@uchicago.edu) for researchers who meet the criteria for access to confidential data.

Funding Statement

This study was supported by the Rynerson Research Fund (NN) and Gianinno Graduate Research Fund (NN) from the University of Chicago. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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PLoS One. doi: 10.1371/journal.pone.0287153.r001

Decision Letter 0

Alexander N Sokolov

16 Mar 2022

PONE-D-21-20363Hormonal responses to brief social interactions: The role of psychosocial stress and relationship statusPLOS ONE

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Reviewer #2: Yes

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Reviewer #1: In this study, the authors investigate the effect of psychosocial stressors on hormonal reactivity to brief encounter with opposite sex individuals in heterosexual humans. The study appear relatively well designed and well suited to test the general question. However, it appears a bit unclear when reading the paper what the question really is. The authors mix up hypothesis and predictions and provide a list of examples of previous studies rather than clear testable hypothesis. Furthermore the statistical analyses are outdated (I strongly recommend the use of Generalized Linear Mixed Models) and some key variable appear uncontrolled for (see below).

Introduction

The introduction is relatively lengthy, not necessarily because it is too long but because it presents a suit of detailed findings from other studies with no clear direction of where the authors go. The main theories regarding hormonal responsiveness to social interactions are not named and the main theories published are not cited. The authors should revise the introduction to expand on theory and merge the exposition of previous findings into less lengthy phrasing.

In the introduction the interplay between Testosterone and cortisol in their hormonal regulation and the negative feedback effects that cortisol has on testosterone should be presented and discussed and incorporated in the hypothesis.

Line 47: please nominatively introduce the challenge hypothesis (Wingfield et al. 1990) and expand citation to other studies which directly cited it in non-human primates or other animals (e.g. Muller & Wrangham 2004; Hirschenhauser & Oliveira 2006; Girard-Buttoz et al. 2015). One key aspect of this hypothesis is differences in testosterone reactivity to social interactions depending on paternity status. It appears therefore crucial to control for this parameter in the analysis beyond simply assessing if individuals are single or not.

Line 75: Here the authors point out the important effect of anticipation. However this parameter was not tested in their experiment since the participants did not know that they would be placed in a (albeit neutral) courtship opportunity situation. Such anticipatory principle is therefore not relevant to the current study.

Line 93: Define ‘confederates’.

Line 121: present what cortisol is and highlight that it regulates the hormonal stress response in mammals including humans. In general, beyond providing a list of what testosterone and cortisol do and have been found to do it would be useful to provide clear cut presentation of what they regulate (e.g. muscle mass, aggressiveness for testosterone and energy mobilization for cortisol) and what they are expected to be sensitive too and why.

Line 123: What is presented in the following lines are not hypothesis but prediction and testable patterns… please formulate hypothesis first.

Methods:

The methods are overall clear and well detailed. As previously, mention the authors should use GLMM rather than anova to clarify methodology and include more control variables. The authors should also run first a full model with all the interactions considered and compared it to a null model using an ANOVA and then reduce this full model removing non-significant interactions to obtain a final reduced model where the interactions and single effects are interpretable. In fact, single effects in interactions cannot be interpreted on their own. As previously mentioned, the authors should control for some important factors such as the paternity status of the men and the maternity status of the woman as well as whether they failed or passed the mathematical test which may have played a role in their hormonal stress response (one can imagine that failing is more stressful and that individual who failed may have more struggle solving mathematical problems hence be exposed to a higher stress).

Line 135: Write men and woman rather than male and female, it should be clear right from the start that you test humans.

Line 147: Clarify what is SES

Line 217: Hormonal level is too general please indicate which level you mean… also the literature cited does not test and present clear result on circadian rhythm of cortisol and testosterone… please cite specific literature (e.g. for cortisol Doman et al. 1986).

Line 224: what concentration were the CV based on? Did you use high concentration and low concentration quality controls?

Line 238: Provide citation and version for R and Jamovi. Can you specify what you sued R for?

Results

Since the result section will be modified following the changes in statistical approach, I will not comment in details on this section. I would however recommend including a table of results to visualize which variables were included in the analyses, in particular in the final model. As presented it is hard to know if the approach is valid or not.

Discussion

Comments on the discussion will be provided on a revised version of the manuscript which revises the statistical approach.

Reviewer #2: The current study investigates whether inducing social stress and interacting with an opposite-sex individual affects participants hormone levels (testosterone and cortisol). It further investigates whether partnered and single participants respond differently to the induced stress or social situation.

My general impression is very positive. The manuscript is expertly written, the research questions are interesting, and the methods are appropriate. I really liked the limitations part in the discussion section (great that you discuss potential issues with salivary immunoassays).

This being said, I would like to share some thoughts that came to my mind while reading through the manuscript. The points raised do only address the results section and the rather complex analyses (that are sometimes hard to interpret). I really hope that my feedback is helpful and helps to improve the manuscript.

First, I am not sure whether I got this right, was the main analysis a 4-way interaction? Or did the authors compute several two-way interactions? This was not 100% clear to me, please specify what you did here in detail. For now, I assume an anova with a 4-way interaction (treatment group x sex x relationship status x saliva sample). If this is correct, I do have strong concerns regarding test power. Besides the fact that such higher order interactions are really complex and difficult to understand, three-way interactions require a much higher test power than two-way interactions (the problem is even more evident for four-way interactions). I would assume that these analyses are heavily underpowered and rather uninformative. Importantly, underpowered studies may not have the power to detect an expected (small) effect size, but, at the same time, underpowered studies that report significant effects might overestimate effect sizes which may lead to difficulties to replicate, due to a higher influence of random measurement error (Button et al., 2013).

Here is my suggestion how to deal with this issue:

I would recommend splitting the analyses, only investigating two-way interaction effects. Focus on the separate research questions you want to answer. To me, it seems as if the two-way interaction between treatment group and saliva sample is more of a method validation (does the TSST lead to higher cortisol/ testosterone levels?) and might be the first step. Then think about the second step and whether it is relevant to e.g. include a treatment variable in the analyses of the next research question. There were planned exploratory analyses regarding singles vs. partnered participants. I would recommend to do them separately from the main analyses, as a second step. Ask yourself if, for answering this specific research question, is it relevant to distinguish between participants who were in the TSST vs. control condition here? Is it relevant to distinguish between male and female participants here, or could this be a next step?

Second, please report (standardized) effect sizes and interpret your results accordingly. Reporting standardized effect sizes would make it easy for the reader to compare effect sizes across different analyses here and across different studies in general (and might also be helpful for future meta-analyses). It would also make it possible for the authors and for each reader to think about the reported effects in terms of effect sizes, not only based on p-values.

Minor:

- Page 17, line 252, something went wrong here (there are two p-values reported): “for control participants was not confirmed (t=1.24, p=1.0, p=0.002; Fig 1)”

- I do understand that you are not allowed to openly share your data, but I want to encourage you to think about sharing your analysis scripts (e.g. R script or SPSS syntax) and maybe, if possible, some material used in this study. This would make it easier for interested scholars to replicate this study in the future or to evaluate the analyses. Every little step in the direction of open and reproducible science helps.

Reference:

Button, K. S., Ioannidis, J. P., Mokrysz, C., Nosek, B. A., Flint, J., Robinson, E. S., & Munafò, M. R. (2013). Power failure: why small sample size undermines the reliability of neuroscience. Nature Reviews Neuroscience, 14, 365-376.

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Reviewer #1: Yes: Cedric Girard-Buttoz

Reviewer #2: No

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PLoS One. 2023 Jun 23;18(6):e0287153. doi: 10.1371/journal.pone.0287153.r002

Author response to Decision Letter 0

14 Sep 2022

Reference: PONE-D-21-20363

Title: Hormonal responses to brief social interactions: The role of psychosocial stress and relationship status

Journal: PLoS ONE

Dear Dr. Sokolov,

We would like to begin by thanking you and the reviewers for your time and thoughtful comments on our paper, which have contributed to an improved manuscript. We paid very close attention to the comments of the reviewers while preparing this revision and have documented the changes we have made below. In order to assist you in reviewing our manuscript’s revisions, we have included each of the reviewer’s comments in the decision letter below, and our response to a comment is italicized and underlined. We have also edited the manuscript to meet the journal requirements mentioned in the decision letter, including formatting for style requirements, editing the use of “male” and “female” as nouns, and clarifying data availability in our updated cover letter.

Thank you in advance for your review!

Sincerely,

Nora (Nickels) McLean

Reviewer #1:

1. The introduction is relatively lengthy, not necessarily because it is too long but because it presents a suit of detailed findings from other studies with no clear direction of where the authors go. The main theories regarding hormonal responsiveness to social interactions are not named and the main theories published are not cited. The authors should revise the introduction to expand on theory and merge the exposition of previous findings into less lengthy phrasing.

The authors have edited the introduction to expand on theory relevant to the research questions (e.g., the challenge hypothesis and the dual hormone hypothesis), as well as to help connect the expected fluctuations of these sex hormones to the research questions of interest. When possible, we have merged the exposition of previous findings to lead to less length phrasing. These recommendations have (very helpfully) led to a clearer flow between past findings and the current research questions.

2. Line 47: please nominatively introduce the challenge hypothesis (Wingfield et al. 1990) and expand citation to other studies which directly cited it in non-human primates or other animals (e.g. Muller & Wrangham 2004; Hirschenhauser & Oliveira 2006; Girard-Buttoz et al. 2015).

The introduction of testosterone in the manuscript has been edited to incorporate the conceptual framework of the challenge hypothesis and supporting nonhuman evidence.

3. One key aspect of this hypothesis is differences in testosterone reactivity to social interactions depending on paternity status. It appears therefore crucial to control for this parameter in the analysis beyond simply assessing if individuals are single or not.

In response to controlling for paternity and maternity status of men and women: only three participants reported having children at the time of the study; cortisol and testosterone levels did not significantly differ between those participants who had children and those who did not.

4. Line 75: Here the authors point out the important effect of anticipation. However this parameter was not tested in their experiment since the participants did not know that they would be placed in a (albeit neutral) courtship opportunity situation. Such anticipatory principle is therefore not relevant to the current study.

This is a helpful comment; participants were not aware of the social interaction that is going to happen. We have removed the anticipatory principle from this paragraph. A social interaction itself can involve stress during the experience, particularly in a potentially social evaluative scenario; therefore, the rest of this paragraph’s introduction to a stressful component to courtship remains in the manuscript.

5. Line 93: Define ‘confederates’.

The term “confederates” has been removed from description of this particular citation, and instead replaced with female “individuals”.

6. Line 121: present what cortisol is and highlight that it regulates the hormonal stress response in mammals including humans. In general, beyond providing a list of what testosterone and cortisol do and have been found to do it would be useful to provide clear cut presentation of what they regulate (e.g. muscle mass, aggressiveness for testosterone and energy mobilization for cortisol) and what they are expected to be sensitive too and why.

When introducing testosterone and cortisol, the authors have edited the manuscript to more thoroughly clarify the regulating functions of these hormones. This includes the clarification of important theories related to the two hormones, and better clarifying how their sensitivity may relate to our main and exploratory research questions.

7. Line 123: What is presented in the following lines are not hypothesis but prediction and testable patterns… please formulate hypothesis first.

The final paragraph of the introduction section has been rewritten to clarify hypotheses and predictions. Specifically, it refers to the main hypotheses and exploratory analyses that are explored in separate sections of the rewritten results section of the revised manuscript.

8. The methods are overall clear and well detailed. As previously, mention the authors should use GLMM rather than anova to clarify methodology and include more control variables. The authors should also run first a full model with all the interactions considered and compared it to a null model using an ANOVA and then reduce this full model removing non-significant interactions to obtain a final reduced model where the interactions and single effects are interpretable. In fact, single effects in interactions cannot be interpreted on their own. As previously mentioned, the authors should control for some important factors such as the paternity status of the men and the maternity status of the woman as well as whether they failed or passed the mathematical test which may have played a role in their hormonal stress response (one can imagine that failing is more stressful and that individual who failed may have more struggle solving mathematical problems hence be exposed to a higher stress).

Reviewers recommended two different statistical approaches for revisions for this manuscript. Based on concerns over power, the authors chose to split analyses and only investigate two-way interactions effects, focusing on specific research questions. Both the main research question and exploratory research questions were limited to two-way interaction effects to avoid concerns regarding test power, as recommended by Reviewer #2. The results section of the manuscript has been completely rewritten based on the new analysis.

Addressing paternity and maternity status of men and women: only three participants reported having children at the time of the study; cortisol and testosterone levels did not significantly differ between those participants who had children and those who did not.

Addressing performance on the mathematical test: the mathematical test that is completed as part of the Trier Social Stress Test is a serial subtraction task. No participant successfully completed the entire task by counting down to zero (with or without a mistake). Data was not collected on how many mistakes were made for each participant.

9. Write men and woman rather than male and female, it should be clear right from the start that you test humans.

Male and female are no longer used as nouns; they are replaced as adjective phrases or with men and women in the manuscript.

10. Line 147: Clarify what is SES

SES has been clarified as socioeconomic status in the manuscript.

11. Line 217: Hormonal level is too general please indicate which level you mean… also the literature cited does not test and present clear result on circadian rhythm of cortisol and testosterone… please cite specific literature (e.g. for cortisol Doman et al. 1986).

Cortisol and testosterone levels have been specified, and additional specific literature has been cited regarding diurnal rhythms and methodological procedures.

12. Line 224: what concentration were the CV based on? Did you use high concentration and low concentration quality controls?

Assay description in the method section of the manuscript has been edited to include specifics about both intra- and inter-assay CV calculations.

13. Line 238: Provide citation and version for R and Jamovi. Can you specify what you sued R for?

For the updated analyses, only jamovi was used. The citation and version have been added to the manuscript.

14. Since the result section will be modified following the changes in statistical approach, I will not comment in details on this section. I would however recommend including a table of results to visualize which variables were included in the analyses, in particular in the final model. As presented it is hard to know if the approach is valid or not.

Reviewers recommended two different statistical approaches for revisions for this manuscript. Based on concerns over power, the authors chose to split analyses and only investigate two-way interactions effects, focusing on specific research questions. Both main research questions and exploratory research questions were limited to two-way interaction effects to avoid concerns regarding test power, as recommended by Reviewer #2. The results section of the manuscript has been completely rewritten based on the new analysis.

Reviewer #2:

1. First, I am not sure whether I got this right, was the main analysis a 4-way interaction? Or did the authors compute several two-way interactions? This was not 100% clear to me, please specify what you did here in detail. For now, I assume an anova with a 4-way interaction (treatment group x sex x relationship status x saliva sample). If this is correct, I do have strong concerns regarding test power. Besides the fact that such higher order interactions are really complex and difficult to understand, three-way interactions require a much higher test power than two-way interactions (the problem is even more evident for four-way interactions). I would assume that these analyses are heavily underpowered and rather uninformative. Importantly, underpowered studies may not have the power to detect an expected (small) effect size, but, at the same time, underpowered studies that report significant effects might overestimate effect sizes which may lead to difficulties to replicate, due to a higher influence of random measurement error (Button et al., 2013). Here is my suggestion how to deal with this issue:

Reviewers recommended two different statistical approaches for revisions for this manuscript. Based on concerns over power, the authors chose to split analyses and only investigate two-way interactions effects, focusing on specific research questions. Both main research questions and exploratory research questions were limited to two-way interaction effects to avoid concerns regarding test power, as recommended by Reviewer #2. The results section of the manuscript has been completely rewritten based on the new analysis.

2. Second, please report (standardized) effect sizes and interpret your results accordingly. Reporting standardized effect sizes would make it easy for the reader to compare effect sizes across different analyses here and across different studies in general (and might also be helpful for future meta-analyses). It would also make it possible for the authors and for each reader to think about the reported effects in terms of effect sizes, not only based on p-values.

Standardized effect sizes and interpretations are now reported for all significant main and interaction effects reported in the manuscript.

3. Page 17, line 252, something went wrong here (there are two p-values reported): “for control participants was not confirmed (t=1.24, p=1.0, p=0.002; Fig 1)”

This error has been removed, due to new analyses replacing the prior results section of the original manuscript.

4. I do understand that you are not allowed to openly share your data, but I want to encourage you to think about sharing your analysis scripts (e.g. R script or SPSS syntax) and maybe, if possible, some material used in this study. This would make it easier for interested scholars to replicate this study in the future or to evaluate the analyses. Every little step in the direction of open and reproducible science helps.

.omt files (jamovi script files, written in R script) are being submitted with the revision. Separate files for main and exploratory analyses are prepared and have been submitted with the revision as a supporting information zip file.

Attachment

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PLoS One. doi: 10.1371/journal.pone.0287153.r003

Decision Letter 1

Alexander N Sokolov

29 Nov 2022

PONE-D-21-20363R1Hormonal responses to brief social interactions: The role of psychosocial stress and relationship statusPLOS ONE

Dear Dr. Nickels,

thank you for submitting your revised manuscript to PLOS ONE. After careful consideration, we feel that it has merit but as it currently stands, still has to be revised in order to fully meet PLOS ONE’s publication criteria. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised by Reviewer 1 listed below, especially concerning the technical aspects of your work.

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Alexander N. 'Sasha' Sokolov, Ph.D.

Academic Editor

PLOS ONE

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Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

Reviewer #1: Yes

Reviewer #2: No

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Reviewer #1: In this revised version, the authors have made a great work at reformatting the introduction and have now a nicely introduced paper. The paper has improved overall and is now in a nice shape. I have still a few comments and the authors should conduct an in depth detailed check of typos and bracket errors since I have seen a few and did not point each of them in detail. I am still advocating for different statistical approaches but what the author used, albeit outdated and not in line with current analytical standard, is not incorrect. The method lacks a few details regarding the hormonal measurements and the choice of the time frame for sample collection. The term confederates should either be explained clearly or removed throughout.

Introduction:

Line 105: “consistent” not “consisten”.

Methods

Line 191: by ‘confederates’ do you mean ‘judges’? I thought you removed the term confederate throughout the manuscript, maybe this is a typo or has been forgotten here.

Line 210: same here and throughout method description, change confederate to another term or define it somewhere.

Line 230-232: Can you clarify this sentence?

Line 236: here or before when describing the timing of saliva sample can you elaborate on the clearance delay of both cortisol and testosterone in saliva and give background citation to support the adequacy of your protocol?

Line 246: what does ‘base on concentration’ mean here. Explain here already that you used high and low QCs? Give CVs for each separately. Also why dii you calculate intra-assay CV as “an average value calculated from the individual CVs from all the plate duplicates”. You should use the average CV only from the duplicates of the QCs and not from your sample as you do not know if theyare contaminated or there were sampling error. Obviously the ducplicates with high CV should be removed and reanalyzed. Can you clarify if you did that and what was your cuttof for reanalysis?

Line 263: Please specify the distribution of single and ‘in relationship’ individuals across the TSST and the control conditions. #

Results

Line 273-276: This is methods and not results. As previously mention the authors should conduct GLMM analyses, ANOVA are highly outdated. With GLMM individual could be entered as random effect to control for the repeated sampling on each individual. I will not reiterate in details my recommendations from the first review but the authors should provide a test that their full model is significantly different from a null model comprising only the control predictors and not the test predictors.

Line 358: close the citation bracket

Discussion

The discussion as it stands is interesting and acknowledges nicely the limitations of the study but remains relatively shallow. The discussion should loop back to the theories introduced in the introduction (challenge hypothesis and dual hormone hypothesis) and specifically state whether the results here corroborate (even if only partly) or not each hypothesis.

Line 292: did you ask the participants specifically how they felt the interaction went and if they had the impression to flirt?

Reviewer #2: I think the authors did a good job and I am convinced by their detailed revisions. Indeed, I feel like they have taken almost all of my and the other reviewer’s comments into account or have at least explained their decisions in a reasonable manner. I appreciate their newly reported analyses and the work they put into rewriting their entire results part.

**********

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Reviewer #1: Yes: Cédric Girard-Buttoz

Reviewer #2: No

**********

PLoS One. 2023 Jun 23;18(6):e0287153. doi: 10.1371/journal.pone.0287153.r004

Author response to Decision Letter 1

14 Jan 2023

The response to specific reviewer and editor comments can be found in the response to reviewers document submitted in this revision. Thank you!

Attachment

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PLoS One. doi: 10.1371/journal.pone.0287153.r005

Decision Letter 2

Alexander N Sokolov

1 Jun 2023

Hormonal responses to brief social interactions: The role of psychosocial stress and relationship status

PONE-D-21-20363R2

Dear Dr. McLean,

thank you for your patience with processing your revised manuscript. We are pleased to inform you that the revision has been judged suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

As a minor comment please consider to re-formulate your Data Availability statement, exchanging the final phrase on possible data provision to qualified researchers upon request to the Social and Behavioral Sciences Institutional Review Board Ethics Committee of your University with the explanation why the data cannot be made available publicly.

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Alexander N. 'Sasha' Sokolov, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

Reviewer #1: No

Reviewer #2: No

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Reviewer #1: (No Response)

Reviewer #2: I didn't have any more requests in the previous round and I can only reiterate that I am satisfied with the author's revisions. In my opinion, this is an interesting manuscript that is worth publishing.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

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Reviewer #1: No

Reviewer #2: No

**********

PLoS One. doi: 10.1371/journal.pone.0287153.r006

Acceptance letter

Alexander N Sokolov

15 Jun 2023

PONE-D-21-20363R2

Hormonal responses to brief social interactions: The role of psychosocial stress and relationship status

Dear Dr. Nickels McLean:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

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on behalf of

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

S1 File. ZIP file containing data analysis script (.omt files) for three sets of analyses.

(ZIP)

Click here for additional data file.^{(356.2KB, zip)}

Attachment

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Data Availability Statement

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Hormonal responses to brief social interactions: The role of psychosocial stress and relationship status

Nora Nickels McLean

Dario Maestripieri

Roles

Abstract

Introduction

Method

Participants

Experimental procedure

Trier social stress test

Social interaction task

Saliva sample collection and hormonal assays

Data analyses

Results

Manipulation check

Fig 1. Changes in cortisol concentrations between control and TSST participants.

Testosterone responses to social interaction

Fig 2. Changes in testosterone concentrations following social interaction as a function of treatment.

Exploratory analyses of relationship status

Fig 3. Changes in testosterone concentrations following social interaction as a function of relationship status in control participants.

Fig 4. Changes in cortisol concentrations following social interaction as a function of relationship status in control participants.

Discussion

Supporting information

Acknowledgments

Data Availability

Funding Statement

References

Decision Letter 0

Alexander N Sokolov

Roles

Author response to Decision Letter 0

Decision Letter 1

Alexander N Sokolov

Roles

Author response to Decision Letter 1

Decision Letter 2

Alexander N Sokolov

Roles

Acceptance letter

Alexander N Sokolov

Roles

Associated Data

Supplementary Materials

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

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