Early nighttime testosterone peaks are correlated with GnRH-induced testosterone in a diurnal songbird.

Experimental manipulation has established testosterone as a potent, pleiotropic regulator coordinating morphology, physiology and behavior. However, the relationship of field-sampled, unmanipulated testosterone concentrations with traits of interest is often equivocal. Circulating testosterone varies over the course of the day, and recent reports indicate that testosterone is higher during the night in diurnal songbirds. Yet, most field studies sample testosterone during the morning. Sampling at times when levels and individual variation are low may be one reason relationships between testosterone and other traits are not always observed. Testosterone is regulated by the hypothalamic-pituitary-gonadal axis, with gonadotropin-releasing hormone (GnRH) initiating the endocrine cascade. Research has examined GnRH-induced testosterone levels with traits of interest, yet the relevance of these induced levels and their relationship with endogenously produced levels are not fully clear. Using photostimulated male great tits (Parus major) we tested the hypotheses that circulating testosterone levels peak during the night and that GnRH-induced testosterone concentrations are positively related to nightly testosterone peaks. Blood was sampled during first, middle or last third of night. One week later, baseline and GnRH-induced testosterone levels were sampled during mid-morning. Morning baseline testosterone levels were low compared with night-sampled levels that peaked during the first third of the night. Further, GnRH-induced testosterone was strongly positively correlated with levels observed during the first third of the night. These data suggest that morning testosterone samples likely do not reflect an individual's endogenous peak. Instead, GnRH-induced testosterone levels do approximate an individual's nightly peak and may be an alternative for birds that cannot easily be sampled at night in the field. These findings are likely to have implications for research aimed at relating traits of interest with natural variation in sex steroid hormone levels.

Sex steroids modulate circadian behavioral rhythms in captive animals, but does this matter in the wild?

Nearly all organisms alter physiological and behavioral activities across the twenty-four-hour day. Endogenous timekeeping mechanisms, which are responsive to environmental and internal cues, allow organisms to anticipate predictable environmental changes and time their daily activities. Among-individual variation in the chronotype, or phenotypic output of these timekeeping mechanisms (i.e. timing of daily behaviors), is often observed in organisms studied under naturalistic environmental conditions. The neuroendocrine system, including sex steroids, has been implicated in the regulation and modulation of endogenous clocks and their behavioral outputs. Numerous studies have found clear evidence that sex steroids modulate circadian and daily timing of activities in captive animals under controlled conditions. However, little is known about how sex steroids influence daily behavioral rhythms in wild organisms or what, if any, implication this may have for survival and reproductive fitness. Here we review the evidence that sex steroids modulate daily timing in vertebrates under controlled conditions. We then discuss how this relationship may be relevant for the reproductive success and fitness of wild organisms and discuss the limited evidence that sex steroids modulate circadian rhythms in wild organisms.

Adult Provisioning Influences Nestling Corticosterone Levels in Florida Scrub Jays (Aphelocoma coerulescens).

We studied Florida scrub jay (Aphelocoma coerulescens) nestlings to examine the relationship between parental feeding rates and levels of corticosterone (CORT), a metabolic and stress-related steroid hormone hypothesized to play a role in mediating begging behavior. It has been documented that nutritional deficiency results in increased glucocorticoid levels in nestling birds. Further, previous studies have found that CORT levels of Florida scrub jay nestlings are negatively correlated with parental nest attendance and provisioning rates; however, the behavioral observations were made several days before the collection of samples to assess CORT levels. Few studies have investigated whether experience immediately before sampling impacts nestling glucocorticoid levels, especially in a free-living species. By monitoring parental activity at the nest before sample collection, we found that nestling CORT levels varied as a function of parental provisioning rate and the time since their last feed. However, counter to our predictions, higher provisioning rates and more recent feedings were associated with higher CORT levels in nestlings rather than lower CORT levels. These results suggest that some aspect of parental provisioning results in increased CORT levels in nestling Florida scrub jays.

Influence of corticosterone treatment on nestling begging in Florida scrub-jays (Aphelocoma coerulescens).

Altricial young are dependent on adults for protection and food, and they display nutritional need by begging to elicit feeding from parents. Begging at high levels can be energetically expensive and attract predators; thus, an individual must balance its nutritional needs with these potential costs. Further, because a parent is limited in the amount of food it can provide, begging can contribute to both parent-offspring conflict and sibling-sibling competition. Many extrinsic and intrinsic factors may contribute to begging behavior. One intrinsic factor of interest is corticosterone (CORT), a metabolic hormone hypothesized to play a role in regulating a nestling's begging behavior. We investigated the hypothesis that increased exposure to CORT influences nestling begging behavior in an altricial species, the Florida scrub-jay (Aphelocoma coerulescens). We treated one nestling per treatment nest with a twice-daily dose of exogenous hormone via a CORT-injected waxworm, whereas a second individual received a vehicle-injected waxworm. We monitored individual nestling and adult behavior at all nests with the use of high-definition video cameras on several days during treatment. We found no difference in begging rate between CORT fed and vehicle fed nestlings within a treatment nest. Further, to determine whether CORT treatment had indirect effects on the entire brood, we monitored additional nests, in which nestlings were not manipulated. When treatment and controls were compared, overall begging rates of nestlings in treatment nests were greater than those in control nests. This result suggests that CORT treatment of an individual altered its behavior, as well as that of its siblings.

Stress-responsiveness influences baseline glucocorticoid levels: Revisiting the under 3min sampling rule.

Plasma glucocorticoid (CORT) levels collected within 3min of capture are commonly believed to reflect pre-stressor, baseline CORT levels. Differences in these "baseline" values are often interpreted as reflecting differences in health, or the amount of social and environmental stress recently experienced by an individual. When interpreting "baseline" values it is generally assumed that any effect of capture-and-handling during the initial sampling period is small enough and consistent enough among individuals to not obscure pre-capture differences in CORT levels. However, plasma CORT increases in less than 3min post-capture in many free-living, endothermic species in which timing has been assessed. In addition, the rate of CORT secretion and the maximum level attained (i.e., the degree of stress-responsiveness) during a severe stressor often differs among individuals of the same species. In Florida scrub-jays (Aphelocoma coerulescens), an individual's stress-responsiveness during a 30min post-capture stressor is correlated with CORT levels in samples collected within 1.5min of capture, suggesting there is an intrinsic connection between stress-responsiveness and pre-capture CORT levels. Although differences in stress-responsiveness accounted for just 11% of the variance in these samples, on average, higher stress-responsive jays (top third of individuals) had baseline values twice that of lower stress-responsive jays (bottom third). Further, plasma CORT levels begin to increase around 2min post-capture in this species, but the rate of increase between 2 and 3min differs markedly with CORT increasing more rapidly in jays with higher stress-responsiveness. Together, these data indicate that baseline CORT values can be influenced by an individual's stress response phenotype and the differences due to stress-responsiveness can be exaggerated during sample collection. In some cases, the effects of differences in stress-responsiveness and the increase in CORT during sample collection could obscure, or supersede, differences in pre-capture plasma CORT levels that are caused by extrinsic factors.