Evaluating Mixtures of Urinary Phthalate Metabolites and Serum Per-/Polyfluoroalkyl Substances in Relation to Adolescent Hair Cortisol: The HOME Study.

Results of toxicological studies indicate that phthalates and per-/polyfluoroalkyl substances (PFAS), 2 classes of endocrine-disrupting chemicals, may alter the functioning of the hypothalamic-pituitary-adrenocortical (HPA) axis. We evaluated the associations of urinary phthalate metabolites and serum PFAS during gestation and childhood with adolescent hair cortisol concentrations (pg/mg hair) at age 12 years, an integrative marker of HPA axis activity (n = 205 mother-child pairs; Cincinnati, Ohio; enrolled 2003-2006). We used quantile-based g-computation to estimate associations between mixtures of urinary phthalate metabolites or serum PFAS and hair cortisol. We also examined whether associations of individual phthalate metabolites or PFAS with cortisol varied by the timing of exposure. We found that a 1-quartile increase in all childhood phthalate metabolites was associated with 35% higher adolescent hair cortisol (phthalate mixture ψ = 0.13; 95% confidence interval: 0.03, 0.22); these associations were driven by monoethyl phthalate, monoisobutyl phthalate, and monobenzyl phthalate. We did not find evidence that phthalate metabolites during gestation or serum PFAS mixtures were related to adolescent hair cortisol concentrations. We found suggestive evidence that higher childhood concentrations of individual PFAS were related to higher and lower adolescent hair cortisol concentrations. Our results suggest that phthalate exposure during childhood may contribute to higher levels of chronic HPA axis activity.

Are hair cortisol levels dependent on hair growth rate? A pilot study in rhesus macaques.

Research incorporating the analysis of glucocorticoids, specifically cortisol, in hair samples has exploded over the past 10-15 years, yet factors contributing to the accumulation of cortisol in hair are not yet fully characterized. In particular, it is not clear whether cortisol accumulation in hair is dependent on hair growth rate, a possibility raised by prior rodent studies reporting glucocorticoid-mediated inhibition of hair growth. Using rhesus macaque monkeys (Macaca mulatta), an extensively studied nonhuman primate species, the present pilot study evaluated the hypothesis that hair cortisol accumulation is inversely related to hair growth rate (i.e., slower hair growth leading to elevated cortisol levels). Hair samples were collected from 19 adult female macaques and 17 infants (9 males) 3 months apart using a shave-reshave procedure from the same site below the posterior vertex of the scalp. The second hair samples were measured to the nearest millimeter (mm) for growth rate over the previous 3 months and assayed for hair cortisol concentrations (HCCs) using enzyme immunoassay. Because of the possibility of age-related differences in hair growth rate, correlational analyses were performed separately for adults and infants to determine whether HCC values were associated with growth rate in each age group. These analyses revealed that neither group displayed a significant correlation of HCCs with hair growth. The results additionally showed that overall, adults had a faster hair growth rate than infants and, as expected from previous studies, had lower HCCs than infants. Our results suggest that higher HCCs within the non-stress range do not result from cortisol-mediated inhibition of hair growth. Moreover, similarities between humans and macaque monkeys in both HPA axis regulation and hair growth rates argue that these findings are relevant for human hair cortisol studies. Extrapolation to other species in which the features of hair growth and the relevant regulatory mechanisms are less well understood should be done with caution.

Transient refugees' social support, mental health, and physiological markers: Evidence from Serbian asylum centers.

OBJECTIVES: Refugees seeking safety across international borders are often exposed to a wide breadth of psychosocially stressful experiences that may fracture existing sources of social support and impair the generation of new social relationships, with implications for their long-term health and resilience. Using data from recently settled refugees in two asylum centers in Serbia, we examined the associations between social support, mental health, and physiological markers. METHODS: In this mixed-method study of refugees (age 18-50 years, n = 76), we collected key socio-demographic information and conducted semi-structured interviews about refugees' journey and stay in Serbia, trauma/loss, and their sources of social support. We also collected self-reported measures of mental well-being as well as physiological markers relevant to repeated exposure to chronic psychosocial stress (fingernail cortisol and dried blood spots for analysis of Epstein-Barr virus [EBV] antibody titers). RESULTS: We found that refugees with longer journeys reported lower social support than those with shorter journeys. Refugees with lower social support reported poorer mental well-being, greater PTSD-related symptoms, and higher recent perceived stress than those with higher social support. We also observed that refugees with lower social support and higher recent stress, respectively, tended to exhibit higher fingernail cortisol levels. However, we did not observe comparable patterns linking EBV antibodies with psychosocial functioning. CONCLUSION: Our cross-sectional findings are consistent with the notion that social support is likely to be a critical component in effective interventions aimed at mitigating the adverse health effects of relocation-related illnesses and poor social functioning as they await resettlement.

Social Fear in US Infants: The Roles of Hair and Salivary Cortisol.

Elevated social fear in infancy poses risk for later social maladjustment and psychopathology. Hair cortisol concentration (HCC), an index of cumulative cortisol exposure, and diurnal salivary cortisol slope, a biomarker of acute stress regulation, have been associated with social fear behaviors in childhood; however, no research has addressed their relations in infancy. Elucidating potential biomarkers of infant social fear behaviors, as well as environmental factors associated with these biomarkers, may grant insights into the ontogeny of fear behaviors that increase risk for internalizing and externalizing psychopathologies later in life. The current study used multiple linear regression to examine if infant HCC, infant diurnal cortisol slope, and income-to-needs ratios (ITN) were differentially associated with observed social fear responses to a Stranger Approach task at 12 months. Using a sample of 90 infants (Mage = 12.26m, SD = 0.81m, 50% female), results indicated that increased infant HCC was associated with increased distress vocalizations during the Stranger Approach task, while steeper diurnal cortisol slope was associated with fewer distress vocalizations. Ordinary least squares path analyses did not reveal group differences between economically strained and non-strained infants in how cortisol measures and social fear responses related. Findings underscore very early psychobiological correlates of fearfulness that may increase risk for fear-related disorders and adverse mental health symptomology across childhood.

Infant diurnal cortisol predicts sleep.

The sleep-wake system is immature at birth and develops in parallel with the hypothalamic-pituitary-adrenal axis, a biological stress system of which the end product is cortisol. Perturbations in one system during infancy can maladaptively influence the maturation of the other system, leading to lasting sleep and cortisol system dysregulation and heightening the risk of enduring health problems. To better understand the early interplay between these systems, we examined whether actigraphy-derived measures of night-time sleep duration and onset were associated with cumulative exposure to cortisol, indexed by hair cortisol concentration, in 12-month-old children. Overall, early sleep onset predicted lower hair cortisol above and beyond sleep duration, family income and chaos experienced at home. Furthermore, both sleep and cortisol levels vary day to day, and temporal dependencies between daily sleep and cortisol regulation are not well understood. Thus, we assessed how the sleep characteristics on a particular evening related to salivary cortisol levels the following day and how daytime and evening cortisol related to the sleep characteristics on the same night. Lower total exposure to cortisol on a particular day was related to longer night-time sleep duration the same night, but not sleep onset. Lower salivary cortisol levels on a given evening related to earlier sleep onset the same night, but not to night-time sleep duration. Sleep duration and onset on a given night were unrelated to total cortisol exposure the following day. Findings suggest that in early development, the day-to-day relation between sleep and cortisol is not bidirectional, but more driven by diurnal cortisol.

Hair cortisol in captive corral-housed baboons.

Hair cortisol concentrations (HCCs) are measures of long-term hypothalamic-pituitary-adrenocortical (HPA) activity and can be used as indicators of chronic stress. However, intrinsic factors such as an animal's age and sex can also have an impact on resulting HCCs. Although baboons are commonly studied in captivity, little is known about baseline HCC in this population. Here we measured HCC in two same-sex groups of captive olive (Papio hamadryas anubis) baboons and olive/yellow baboon (Papio hamadryas cynocephalus) crosses housed in large outdoor corrals, and we assessed the impact of age and sex on HCC as major variables of interest. Hair was gently shaved from the back of the neck when the animals were sedated for routine physicals. Subjects were divided into three age categories: juvenile (2-4 years), adult (9-12 years), and senior (13-19 years). The "senior" category contained only males. Results confirm an effect of sex and age on HCCs. Females had higher levels of hair cortisol than males, and juveniles had higher levels than adults. There was also a significant sex × age interaction. There were no sex differences in HCCs in juveniles, but there was a greater decline in HCCs in adult males than in adult females. Within males, there was a significant difference in levels of hair cortisol across the three age categories. Juveniles had higher levels than did adults and seniors, but adults and seniors were not significantly different from one another. These results provide baseline measures of hair cortisol in captive baboons and demonstrate effects of sex and age on HCCs.

Pregnancy and Infant Development (PRIDE)-a preliminary observational study of maternal adversity and infant development.

BACKGROUND: Children from socioeconomically disadvantaged families have a markedly elevated risk for impaired cognitive and social-emotional development. Children in poverty experience have a high risk for developmental delays. Poverty engenders disproportionate exposure to psychological adversity which may contribute to impaired offspring development; however the effect may be mitigated by social support and other aspects of resilience. Our objective was to determine the association between maternal stress, adversity and social support and early infant neurobehavior and child behavior at two and three years. METHODS: We conducted a longitudinal mother-infant cohort study nested within a regional home visiting program in Cincinnati, Ohio. Four home study visits were completed to collect measures of maternal stress, adversity and social support and infant and child behavior. A measure of infant neurobehavior ('high-arousal' infant) was derived from the NICU Network Neurobehavioral Scale (NNNS) at 1 month and externalizing and internalizing symptoms were measured by the Child Behavior Checklist (CBCL) at 24 and 36 months. Linear and logistic regression identified associations between maternal risk/protective factors and infant and child behavioral measures. We used stratification and multiplicative interaction terms to examine potential interactions. RESULTS: We enrolled n = 55 pregnant mothers and follow 53 mother-offspring dyads at 1 month, 40 dyads at 24 months and 27 dyads at 36 months. Maternal adversity and protective factors were not associated with neurobehavior at one month. However, maternal depression and measures of distress in pregnancy were significantly associated with internalizing and externalizing symptoms at 24 and 36 months. CONCLUSIONS: This pilot study established the feasibility of conducting longitudinal research within a community intervention program. In addition, although there were no statistically significant associations between maternal psychosocial factors in pregnancy and infant neurobehavior, there were several associations at 24 months, primarily internalizing symptoms, which persisted through 36 months. Future work will replicate findings within a larger study as well as explore mediators and modifiers of these associations.

Assessment of prenatal stress-related cortisol exposure: focus on cortisol accumulation in hair and nails.

Prenatal stress adversely affects offspring development. Although cortisol is hypothesized to be a key mediator of stress-induced developmental deficits, determining the amount of fetal cortisol exposure produced by maternal stress has proved challenging. Current approaches, such as measuring cortisol concentrations in maternal plasma, saliva, or urine, amniotic fluid, fetal plasma, or cord blood, all have significant limitations for assessing cumulative fetal cortisol exposure over time. A recently emerging approach is to measure cortisol concentrations in maternal hair and/or newborn hair or nail samples. Maternal hair cortisol potentially shows long-term production across each trimester of pregnancy, whereas neonatal hair or nail cortisol is thought to reflect mainly third trimester hormone accumulation. This review first describes fetal adrenocortical development, placental cortisol metabolism, and the various sources of fetal cortisol exposure across pregnancy. We then summarize the results obtained from "classical" methods of assessing prenatal cortisol exposure prior to the advent of hair and nail cortisol measurement. Lastly, we discuss the initial development and validation of the hair cortisol methodology, its subsequent application to studies of chronic stress, and recent findings regarding maternal and neonatal hair or nail cortisol concentrations in relation to prenatal stress and other variables of interest.

Socioeconomic factors, stress, hair cortisol, and white matter microstructure in children.

Socioeconomic disadvantage has been linked to increased stress exposure in children and adults. Exposure to stress in childhood has been associated with deleterious effects on cognitive development and well-being throughout the lifespan. Further, exposure to stress has been associated with differences in brain development in children, both in cortical and subcortical gray matter. However, less is known about the associations among socioeconomic disadvantage, stress, and children's white matter development. In this study, we investigated whether socioeconomic disparities would be associated with differences in white matter microstructure in the cingulum bundle, as has been previously reported. We additionally investigated whether any such differences could be explained by differences in stress exposure and/or physiological stress levels. White matter tracts were measured via diffusion tensor imaging in 58 children aged 5-9 years. Results indicated that greater exposure to stressful life events was associated with higher child hair cortisol concentrations. Further, physiological stress, as indexed by hair cortisol concentrations, were associated with higher fractional anisotropy in the cingulum bundle. These results have implications for better understanding how perceived and physiological stress may alter neural development during childhood.

Infants of mothers with higher physiological stress show alterations in brain function.

Chronic stress has been increasingly linked with aberrations in children's behavioral, cognitive, and social development, yet the effect of chronic physiological stress on neural development during the first year of life is largely unknown. The present study aims to link a physiological index of chronic stress (maternal hair cortisol concentration) to maturational differences in infant functional brain development during the first year of life. Participants were 94 mother-infant dyads. To index chronic physiological stress, maternal hair samples were assayed for the previous three months' cortisol output. To examine the development of brain function during the first year of life, six-to-twelve-month-old infants (N = 94) completed a resting electroencephalography (EEG) recording. Infants of mothers with evidence of higher physiological stress showed increased relative low-frequency (theta) power and reduced relative high-frequency (alpha, high-gamma) power, compared to infants of mothers with evidence of low physiological stress. This pattern of findings is consistent with other studies suggesting that early life stress may lead to alterations in patterns of infant brain development. These findings are important given that maturational lags in brain development can be long-lasting and are associated with deficits in cognitive and emotional development. The present research also suggests that reducing maternal physiological stress may be a useful target for future interventions aiming to foster neurodevelopment during the first year of life.

Cortisol and socioeconomic status in early childhood: A multidimensional assessment.

The hypothalamic-pituitary-adrenal (HPA) axis is sensitive to early life stress, with enduring consequences for biological stress vulnerability and health (Gunnar & Talge, 2008). Low socioeconomic status (SES) is associated with dysregulation of the stress hormone cortisol in early childhood. However, a mechanistic understanding of this association is lacking. Multidimensional assessment of both SES and cortisol is needed to characterize the intricate relations between SES and cortisol function in early childhood. We assessed parent-reported family income, parent education, occupational prestige, neighborhood risk, food insecurity, and household chaos for 12-month-old infants (N = 90) and 3.5-year-old children (N = 91). Hair cortisol concentration (HCC) was obtained from parent and child, indexing chronic biological stress, and diurnal salivary cortisol was measured in the children. Controlling for parent HCC, parent education uniquely predicted infant and child HCC and, in addition, neighborhood risk uniquely predicted infant HCC. Household chaos predicted bedtime salivary cortisol concentration (SCC) for both infants and children, and infant daily cortisol output. Food insecurity was associated with flattened cortisol slope in 3.5-year-old children. Parental sensitivity did not mediate relations between SES and cortisol. Results highlight the utility of SES measures that index unpredictable and unsafe contexts, such as neighborhood risk, food insecurity, and household chaos.

Developmental outcomes of early adverse care on amygdala functional connectivity in nonhuman primates.

Despite the strong link between childhood maltreatment and psychopathology, the underlying neurodevelopmental mechanisms are poorly understood and difficult to disentangle from heritable and prenatal factors. This study used a translational macaque model of infant maltreatment in which the adverse experience occurs in the first months of life, during intense maturation of amygdala circuits important for stress and emotional regulation. Thus, we examined the developmental impact of maltreatment on amygdala functional connectivity (FC) longitudinally, from infancy through the juvenile period. Using resting state functional magnetic resonance imaging (MRI) we performed amygdala-prefrontal cortex (PFC) region-of-interest and exploratory whole-brain amygdala FC analyses. The latter showed (a) developmental increases in amygdala FC with many regions, likely supporting increased processing of socioemotional-relevant stimuli with age; and (b) maltreatment effects on amygdala coupling with arousal and stress brain regions (locus coeruleus, laterodorsal tegmental area) that emerged with age. Maltreated juveniles showed weaker FC than controls, which was negatively associated with infant hair cortisol concentrations. Findings from the region-of-interest analysis also showed weaker amygdala FC with PFC regions in maltreated animals than controls since infancy, whereas bilateral amygdala FC was stronger in maltreated animals. These effects on amygdala FC development may underlie the poor behavioral outcomes associated with this adverse experience.

Maternal expressive suppression moderates the relations between maternal and child hair cortisol.

Self-reports and physiological indicators of stress such as cortisol levels are correlated in maternal and child samples. This relationship is likely to be influenced by maternal emotion regulation. Herein, we investigate the moderating role of maternal regulation strategies on the association between maternal and child hair cortisol levels. Mother-child dyads (N = 63, child mean age = 49.74 months) participated in the study. Hair samples were collected from mother and child, and cortisol was assayed. Mothers reported on their own emotion regulation strategies, namely expressive suppression and cognitive reappraisal. As expected, mother and child hair cortisol levels were significantly correlated. Interestingly, the relation between maternal and child hair cortisol was moderated by maternal suppression of emotion. Mother and child hair cortisol levels were related at low levels of maternal suppression, but not at higher levels of suppression. Maternal cognitive reappraisal of emotion was not associated with cortisol levels.

Maternal hair cortisol levels as a novel predictor of neonatal abstinence syndrome severity: A pilot feasibility study.

Neonatal abstinence syndrome (NAS) after in-utero opioid exposure remains a poorly understood condition with multiple factors contributing to severity. Exposure to maternal stress may be one contributing factor. Hair cortisol measurement represents a novel technique for assessing prenatal stress. In this pilot study, the association between maternal hair cortisol levels and NAS severity was examined in 70 postpartum women with opioid use disorder within 72 hr of delivery. Infants were monitored for NAS and treated according to institutional protocol. Forty-four (63%) of the infants were pharmacologically treated for NAS, with a mean length of hospital stay (LOS) for all infants of 14.2 (SD 9.0) days. The mean cortisol level in the mothers was 131.8 pg/mg (SD 124.7). In bivariate analysis, higher maternal hair cortisol levels were associated with shorter infant LOS (R = -.26, p = .03) and fewer infant opioid treatment days (R = -.28, p = .02). Results were no longer statistically significant in regression models after adjusting for maternal opioid and smoking. In conclusion, we demonstrated the feasibility of hair cortisol assaying within the first few days after delivery in mothers with opioid use disorder as a novel marker for NAS. The findings suggest that maternal stress may impact the severity of infant opioid withdrawal.

Alopecia in rhesus macaques (Macaca mulatta): Association with pregnancy and chronic stress.

BACKGROUND: Alopecia in captive non-human primates is often presented as a welfare issue. However, it is a complex condition with a number of possible causes. The purpose of this study was to assess the impact of pregnancy and hair cortisol concentrations on alopecia in rhesus macaques. METHODS: Subjects were 113 socially housed adult female rhesus macaques (27 pregnant, 35 nursing infants, 51 controls). During routine physicals, photographs were taken for alopecia assessment and hair samples were collected for cortisol assay. RESULTS: Alopecia was more prevalent in pregnant than in control females, but there was no association between alopecia and hair cortisol. However, there was a significant effect of pregnancy on hair cortisol. Nursing females had higher hair cortisol levels than pregnant females, which had higher levels than control females. CONCLUSIONS: Although alopecia does not appear to be associated with hair cortisol, both alopecia and hair cortisol were associated with pregnancy.

Aggression and social support predict long-term cortisol levels in captive tufted capuchin monkeys (Cebus [Sapajus] apella).

Many nonhuman primates live in complex social groups in which they regularly encounter both social stressors such as aggression and social support such as that provided by long-term affiliative relationships. Repeated exposure to social stressors may result in chronically elevated cortisol levels that can have deleterious physical effects such as impaired immune function, cardiovascular disease, and reduced brain function. In contrast, affiliative social relationships may act as a buffer, dampening the release of cortisol in response to acute and chronic stressors. Understanding how social stressors and social support predict cortisol levels is therefore essential to understanding how social situations relate to health and welfare. We studied this relationship in 16 socially housed captive brown capuchin monkeys (Cebus [Sapajus] apella) by comparing long-term hair cortisol levels with behavioral measures of social stress (dominance rank, rank certainty, and amount of aggression received) and social support (amount of affiliation and centrality in the affiliative social network of the group). Dominance rank, rank certainty, amount of affiliation, and age were not significant predictors of long-term cortisol levels in this population. Instead, long-term cortisol levels were positively related to the amount of aggression received and negatively related to centrality in the affiliative social network of the group. This pattern may be attributed to the species' socially tolerant dominance system and to the availability of social support across the dominance hierarchy.

Emotion regulation moderates the association between parent and child hair cortisol concentrations.

Successful emotion regulation facilitates children's coping with everyday stress. It develops rapidly in the early preschool period. However, no work has been done to investigate the potential buffering role of emotion regulation from cumulative physiological effects of stress. In this study, we examined hair cortisol concentration (HCC), an early marker of chronic physiological stress, socioeconomic status (SES), parental sensitivity, and emotion regulation and reactivity in a sample of 3.5-year-old children (N = 86). Emotion regulation and emotional reactivity were independent of child HCC. However, emotion regulation moderated the relationship between parent and child HCC. For children with better emotion regulation, there was no association between parent and child HCC, suggesting that emotion regulation skills buffered the transgenerational effects of chronic physiological stress. Emotional reactivity moderated the relationship between SES and child HCC, and attenuated the association between parental sensitivity and child HCC. Taken together, these findings demonstrate that children who were less emotionally reactive were less susceptible to their environments. Results provide support that child emotion regulation and emotional reactivity can reduce or strengthen the relationship between established risk factors and levels of chronic physiological stress in early childhood.

Salivary cortisol reactivity in preschoolers is associated with hair cortisol and behavioral problems.

The interplay between children's cortisol reactivity to challenge and cumulative cortisol exposure is not well understood. Examining the role of cortisol reactivity in early childhood may elucidate biological mechanisms that contribute to children's chronic physiological stress and behavioral dysregulation. In a sample of 65 preschool-aged children, we examined the relation between children's salivary cortisol reactivity to challenging tasks and their hair cortisol concentration (HCC). While both are biomarkers of the hypothalamic-pituitary-adrenal (HPA) axis, salivary cortisol reactivity reflects an acute cortisol response to a stressor and HCC reflects cumulative cortisol exposure. In addition, we examined the relations of these stress biomarkers with internalizing and externalizing problems. Salivary cortisol reactivity was associated with higher HCC and with increased externalizing behaviors. Child HCC also was positively correlated with parent HCC. Results highlight the contributions of salivary cortisol reactivity to children's cumulative cortisol exposure, which may add to their biological risk for health problems later. The observed association between externalizing problems and salivary cortisol reactivity indicates concordances between dysregulated behavioral reactions and dysregulated cortisol responses to challenges. The finding that salivary cortisol reactivity to challenge in early childhood plays a role in children's cumulative cortisol exposure and behavioral development suggests pathways through which cortisol reactivity may influence long-term physical and mental health.

Investigating relations among stress, sleep and nail cortisol and DHEA.

In the current study, we present data investigating the relationships among stress, sleep disturbance, self-control, and levels of cortisol (CORT) and dehydroepiandrosterone (DHEA) in fingernail clippings. Currently, hair CORT is the only routinely used noninvasive, validated, biomarker of chronic exposure to stress-related hormones. Nail clippings represent an important potential alternative sample matrix for assessing chronic hormone exposure, as it offers a different timeline of hormone incorporation than scalp hair, and may be obtainable from populations in which hair either is lacking or is unavailable for cultural reasons. Moreover, there is established precedent for using fingernail clippings to attain biomarker data. However, the value of nail hormone assessment for psychological research is currently unknown due to a paucity of information on the relations between nail hormone concentrations and environmental or psychological variables. In the present study, we collected data from a low income, minority population (N = 47; 97% African American) to demonstrate feasibility and acceptability of nail collection and analysis of the adrenal steroids CORT and DHEA. Participants reported on perceived stress, sleep and self-control abilities. Correlational analyses suggest that exposure to stressful events, disturbances in sleep and waking were associated with higher levels of nail DHEA, while self-control was associated with higher levels of nail CORT. We discuss the potential importance of this methodology for investigating biological, behavioral, and subjective indices of stress and well-being.

Cortisol in Neonatal Mother's Milk Predicts Later Infant Social and Cognitive Functioning in Rhesus Monkeys.

Milk provides not only the building blocks for somatic development but also the hormonal signals that contribute to the biopsychological organization of the infant. Among mammals, glucocorticoids (GCs) in mother's milk have been associated with infant temperament. This study extended prior work to investigate rhesus monkey (Macaca mulatta) mother-infant dyads (N = 34) from birth through 8 months postpartum. Regression analysis revealed that cortisol concentrations in milk during the neonatal period predicted impulsivity on a cognitive task, but not global social behaviors, months later. During this time period, sex-differentiated social behavior emerged. For female infants, milk cortisol concentrations predicted total frequency of play. Collectively, these findings support and extend the "lactational programming" hypothesis on the impact of maternal-origin hormones ingested via milk.