Brigitte M. Kudielka

Salivary cortisol as a biomarker in stress research.

Salivary cortisol is frequently used as a biomarker of psychological stress. However, psychobiological mechanisms, which trigger the hypothalamus-pituitary-adrenal axis (HPAA) can only indirectly be assessed by salivary cortisol measures. The different instances that control HPAA reactivity (hippocampus, hypothalamus, pituitary, adrenals) and their respective modulators, receptors, or binding proteins, may all affect salivary cortisol measures. Thus, a linear relationship with measures of plasma ACTH and cortisol in blood or urine does not necessarily exist. This is particularly true under response conditions. The present paper addresses several psychological and biological variables, which may account for such dissociations, and aims to help researchers to rate the validity and psychobiological significance of salivary cortisol as an HPAA biomarker of stress in their experiments.

HPA axis responses to laboratory psychosocial stress in healthy elderly adults, younger adults, and children: impact of age and gender

Data from five independent studies were reanalyzed in order to investigate the impact of age and gender on HPA axis responses to an acute psychosocial laboratory stress task. The total sample consisted of 102 healthy subjects with 30 older adults (mean age: 67.3 y), 41 young adults (mean age: 23.5 y), and 31 children (mean age: 12.1 y). All participants were exposed to the Trier Social Stress Test (TSST). The stress protocol caused highly significant ACTH and total plasma cortisol responses in older and younger male and female adults (all p<0.0001) as well as salivary free cortisol responses in all six age and gender groups (all p<0.0001). Three-way ANOVAs for repeated measurement were applied to investigate the impact of age and gender on ACTH and cortisol responses. Results showed that the ACTH response to stress was higher in younger adults compared to older adults (main effect: p=0.009, interaction: p=0.06). Post hoc analyses revealed that there was no age effect in the subgroup of women (p=n.s.), while younger men had higher ACTH responses compared to older men (p=0.01). For total plasma cortisol, ANOVA results showed that the pattern of reactivity did not differ between age and gender groups (all interactional effects p=n.s.), although older females had hightened overall cortisol levels compared to the other groups, as proofed in post hoc analyses (all p<0.05). For free salivary cortisol, a significant main effect of gender (p=0.05) and an almost significant three-way-interaction (p=0.09) emerged. Post hoc analyses showed an elevated overall free salivary cortisol response in elderly men compared to elderly women (p=0.006), while no gender differences emerged in neither young adults nor children (both p=n.s.). In sum, the stressor induced significant HPA axis responses in all age and gender groups. The observed ACTH response patterns in young and elderly adults may suggest that a heightened hypothalamic drive in young men decreases with age, resulting in similar ACTH responses in elderly men and women. Alternative interpretations are also discussed. The data also supports the idea of a greater adrenal cortex sensitivity to ACTH signals in young females. Free salivary cortisol responses were elevated in elderly men compared to elderly women, an effect which cannot be explained by gender differences in perceived stress responses to the TSST. It can be speculated if corticosteroid binding globulin (CBG) and/or sex steroids are important modulators of these effects.

Why do we respond so differently? Reviewing determinants of human salivary cortisol responses to challenge

Stress and stress-related health impairments are major problems in human life and elucidating the biological pathways linking stress and disease is of substantial importance. However, the identification of mechanisms underlying a dysregulation of major components of the stress response system is, particularly in humans, a very challenging task. Salivary cortisol responses to diverse acute challenge paradigms show large intra- and interindividual variability. In order to uncover mechanisms mediating stress-related disorders and to potentially develop new therapeutic strategies, an extensive phenotyping of HPA axis stress responses is essential. Such a research agenda depends on substantial knowledge of moderating and intervening variables that affect cortisol responses to different stressors and stimuli. The aim of this report is, therefore, to provide a comprehensive summary of important determinants of, in particular, human salivary cortisol responses to different kinds of laboratory stimuli including acute psychosocial stress as well as pharmacological provocation procedures. This overview demonstrates the role of age and gender, endogenous and exogenous sex steroid levels, pregnancy, lactation and breast-feeding, smoking, coffee and alcohol consumption as well as dietary energy supply in salivary cortisol responses to acute stress. Furthermore, it briefly summarizes current knowledge of the role of genetic factors and methodological issues in terms of habituation to repeated psychosocial stress exposures and time of testing as well as psychological factors, that have been shown to be associated with salivary cortisol responses like early life experiences, social factors, psychological interventions, personality as well as acute subjective-psychological stress responses and finally states of chronic stress and psychopathology.

Day-to-day dynamics of experience--cortisol associations in a population-based sample of older adults.

In 156 older adults, day-to-day variations in cortisol diurnal rhythms were predicted from both prior-day and same-day experiences, to examine the temporal ordering of experience–cortisol associations in naturalistic environments. Diary reports of daily psychosocial, emotional, and physical states were completed at bedtime on each of three consecutive days. Salivary cortisol levels were measured at wakeup, 30 min after awakening, and at bedtime each day. Multilevel growth curve modeling was used to estimate diurnal cortisol profiles for each person each day. The parameters defining those profiles (wakeup level, diurnal slope, and cortisol awakening response) were predicted simultaneously from day-before and same-day experiences. Prior-day feelings of loneliness, sadness, threat, and lack of control were associated with a higher cortisol awakening response the next day, but morning awakening responses did not predict experiences of these states later the same day. Same-day, but not prior-day, feelings of tension and anger were associated with flatter diurnal cortisol rhythms, primarily because of their association with higher same-day evening cortisol levels. Although wakeup cortisol levels were not predicted by prior-day levels of fatigue and physical symptoms, low wakeup cortisol predicted higher levels of fatigue and physical symptoms later that day. Results are consistent with a dynamic and transactional function of cortisol as both a transducer of psychosocial and emotional experience into physiological activation and an influence on feelings of energy and physical well-being.

Awakening cortisol responses are influenced by health status and awakening time but not by menstrual cycle phase

Recent evidence suggested that the free cortisol response to awakening is influenced by awakening time in healthy younger adults (Edwards et al., 2001). In order to investigate this association further, 179 community-dwelling subjects of a large age range (4-75 yrs) participated in the present study. The sample consisted of 99 women, 67 men and 13 children. Subjects were instructed to obtain saliva samples directly after awakening as well as 15, 30, 45, and 60 minutes thereafter. A first analysis revealed that salivary cortisol profiles after awakening in healthy subjects differed from profiles in subjects who reported health problems or a chronic disease (p = 0.02) with healthy subjects showing a larger cortisol response. Therefore, only healthy subjects were included in the following analyses. Subjects woke up between 0455 and 1203 h. Time of awakening strongly influenced the course of morning cortisol levels. Cortisol profiles differed significantly between two wake-up groups (p<0.001). Similarly, group differences for cortisol increase (p = 0.03) and area under the curve (p = 0.05) were also significant, with more pronounced responses in early awakeners compared to late awakeners. The findings are discussed with respect to the circadian cortisol rhythm and the effects of light exposure. Age was correlated with the cortisol levels immediately after awakening (r = 0.2, p = 0.04), the area under the cortisol curve ( r = -0.20, p = 0.05), and with time of awakening (r = -0.21, p = 0.04), respectively. No differences were found between males and females, or between profiles obtained during the follicular or luteal menstrual cycle phase. Also, no differences were observed between habitual smokers vs. non-smokers. These data suggest that the morning cortisol response is influenced by the awakening time but not by menstrual cycle phase. Moreover, health status and age appear to have an impact on this marker of adrenocortical activity. Wake-up time, health status and age should therefore be controlled for in future studies measuring cortisol responses to awakening.

Acute HPA axis responses, heart rate, and mood changes to psychosocial stress (TSST) in humans at different times of day

There is evidence showing that HPA axis responses to pharmacological provocation depend on time of day with larger cortisol responses in the afternoon and evening compared to the morning hours. However, it is still unknown whether HPA axis responses to psychological stress are affected by time of day and whether they can be assessed with equal reliability in the morning and afternoon, respectively. The present reanalysis is based on five independent studies conducted in the same laboratory by and. All subjects were confronted with the Trier Social Stress Test (TSST) either in the morning or in the afternoon. The total sample consisted of 180 adults with 115 younger (49 females, 66 males) and 65 older adults (32 females, 33 males). All ANCOVA results controlled for possible age and gender effects. Stress-related free salivary cortisol, total plasma cortisol and ACTH net increases did not differ according to time of day (all p = n.s.). However, as expected pre-stress free salivary and total plasma cortisol levels differed significantly between the morning and afternoon group (both p < 0.005), leading to a significantly higher free cortisol area under the curve (AUC) in the morning (p = 0.02). Taken together, these observations suggest that the adrenal glands may be more sensitive to ACTH in the morning. Additionally, higher basal salivary cortisol levels were related to a lower stress-related net increase in salivary cortisol (p = 0.02), total plasma cortisol (p < 0.0001), and marginally ACTH (p = 0.09). Stress-related heart rate increases did not differ between groups (p = n.s.). The finding that the TSST-induced mood change was differentially affected by time of day requires further exploration. We conclude that comparable HPA axis and heart rate stress responses to psychosocial stress can be measured in the morning and afternoon.

Trier Social Stress Test

More than 10 years ago, the Trier Social Stress Test (TSST) was introduced as a standardized protocol for the induction of moderate psychosocial stress in laboratory settings. This article provides an up-to-date description of the TSST protocol and a brief review of a decade of research with the TSST.

Human models in acute and chronic stress: Assessing determinants of individual hypothalamus–pituitary–adrenal axis activity and reactivity

Stress is one of the most significant health problems in modern societies and the 21st century. This explains a pressing need for investigations into the biological pathways linking stress and health. Besides the locus coeruleus–noradrenaline/autonomic (sympathetic) nervous system (), the hypothalamus–pituitary–adrenal (HPA) axis is the major physiological stress response system in the body. Since alterations in HPA axis regulation under basal conditions and in response to acute stress appear to be a close correlate or even a determining factor of the onset of different diseases or disease progression (; ; ; ; ; ; ), the characterization of an individuals HPA axis activity as well as reactivity pattern to psychosocial stress appears to be of major interest. It is obvious that such a research agenda substantially depends on the availability of appropriate measures. However, since the HPA axis is a highly adaptive system which is characterized by marked inter- and intraindividual variability (; ), the development of such markers of HPA axis regulation in humans was—and still is—a rather challenging task. In this brief review, we focus on findings on two HPA axis measures, namely the cortisol-awakening response (CAR) to assess HPA axis basal activity and the Trier social stress test (TSST) to investigate HPA axis stress reactivity.

Assessment of the cortisol awakening response: Expert consensus guidelines

The cortisol awakening response (CAR), the marked increase in cortisol secretion over the first 30-45 min after morning awakening, has been related to a wide range of psychosocial, physical and mental health parameters, making it a key variable for psychoneuroendocrinological research. The CAR is typically assessed from self-collection of saliva samples within the domestic setting. While this confers ecological validity, it lacks direct researcher oversight which can be problematic as the validity of CAR measurement critically relies on participants closely following a timed sampling schedule, beginning with the moment of awakening. Researchers assessing the CAR thus need to take important steps to maximize and monitor saliva sampling accuracy as well as consider a range of other relevant methodological factors. To promote best practice of future research in this field, the International Society of Psychoneuroendocrinology initiated an expert panel charged with (i) summarizing relevant evidence and collective experience on methodological factors affecting CAR assessment and (ii) formulating clear consensus guidelines for future research. The present report summarizes the results of this undertaking. Consensus guidelines are presented on central aspects of CAR assessment, including objective control of sampling accuracy/adherence, participant instructions, covariate accounting, sampling protocols, quantification strategies as well as reporting and interpreting of CAR data. Meeting these methodological standards in future research will create more powerful research designs, thus yielding more reliable and reproducible results and helping to further advance understanding in this evolving field of research.

Two weeks of transdermal estradiol treatment in postmenopausal elderly women and its effect on memory and mood: verbal memory changes are associated with the treatment induced estradiol levels

The present randomized double blind study investigated the effects of a 2 week transdermal estradiol treatment on memory performance in 38 healthy elderly women. Cognitive performance was tested at baseline and after 2 weeks of estradiol or placebo treatment using verbal, semantic, and spatial memory tests as well as a mental rotation task and the Stroop. Initial results showed no differences after treatment between placebo or estradiol treated subjects. However, within treatment group analysis revealed that estradiol treated subjects who reached higher estradiol levels (larger than 29 pg/ml) performed significantly better after treatment in the delayed recall of the paired associate test (verbal memory) than subjects who reached lower estradiol levels (P < 0.05). A nonsignificant trend was observed for the immediate recall condition (P < 0.10). These findings were strengthened by correlations between treatment-induced estradiol levels and changes in verbal memory performance. In addition, there was an association between estradiol levels and mood changes. However mood changes were not significantly associated with changes in verbal memory performance (P > 0.20). The present study supports the idea that estradiol replacement has specific effects on verbal memory in healthy postmenopausal women, with delayed recall being more affected. It suggests that these effects can occur relatively rapidly, and that there may be a dose response relationship of estradiol to memory enhancement. Furthermore, the fact that these results were obtained in women who had been menopausal for an average of 17 years before entering the study indicates that the brain maintains a sensitivity for estrogens even after years of low estradiol plasma concentrations.