Uncovering a 'sensitive window' of multisensory and motor neuroplasticity in the cerebrum and cerebellum of male and female starlings.

Traditionally, research unraveling seasonal neuroplasticity in songbirds has focused on the male song control system and testosterone. We longitudinally monitored the song behavior and neuroplasticity in male and female starlings during multiple photoperiods using Diffusion Tensor and Fixel-Based techniques. These exploratory data-driven whole-brain methods resulted in a population-based tractogram confirming microstructural sexual dimorphisms in the song control system. Furthermore, male brains showed hemispheric asymmetries in the pallium, whereas females had higher interhemispheric connectivity, which could not be attributed to brain size differences. Only females with large brains sing but differ from males in their song behavior by showing involvement of the hippocampus. Both sexes experienced multisensory neuroplasticity in the song control, auditory and visual system, and cerebellum, mainly during the photosensitive period. This period with low gonadal hormone levels might represent a 'sensitive window' during which different sensory and motor systems in the cerebrum and cerebellum can be seasonally re-shaped in both sexes.

In vivo online monitoring of testosterone-induced neuroplasticity in a female songbird.

Adult neuroplasticity in the song control system of seasonal songbirds is largely driven by photoperiod-induced increases in testosterone. Prior studies of the relationships between testosterone, song performance and neuroplasticity used invasive techniques, which prevent analyzing the dynamic changes over time and often focus on pre-defined regions-of-interest instead of examining the entire brain. Here, we combined (i) in vivo diffusion tensor imaging (DTI) to assess structural neuroplasticity with (ii) repeated monitoring of song and (iii) measures of plasma testosterone concentrations in thirteen female photosensitive starlings (Sturnus vulgaris) who received a testosterone implant for 3 weeks. We observed fast (days) and slower (weeks) effects of testosterone on song behavior and structural neuroplasticity and determined how these effects correlate on a within-subject level, which suggested separate contributions of the song motor and anterior forebrain pathways in the development of song performance. Specifically, the increase in testosterone correlated with a rapid increase of song rate and RA volume, and with changes in Area X microstructure. After implant removal, these variables rapidly reverted to baseline levels. In contrast, the more gradual improvement of song quality was positively correlated with the fractional anisotropy values (DTI metric sensitive to white matter changes) of the HVC-RA tract and of the lamina mesopallialis, which contains fibers connecting the song control nuclei. Thus, we confirmed many of the previously reported testosterone-induced effects, like the increase in song control nuclei volume, but identified for the first time a more global picture of the spatio-temporal changes in brain plasticity.

Plasma Biochemical and Lipid Panel Reference Intervals in Common Mynahs (Acridotheres tristis).

Mynahs are popular pet and aviary birds. Captive mynahs are prone to metabolic disorders, such as obesity, hepatic lipidosis, and atherosclerosis, and determination of a plasma biochemical reference interval is essential for diagnosis, treatment, and monitoring of these disorders. To establish reference intervals for the lipoprotein panel and plasma activity of liver enzymes in common mynahs (Acridotheres tristis), glucose, cholesterol, triglycerides, high and low density lipoprotein concentrations, and activities of aspartate aminotransferase, alkaline phosphatase, gamma-glutamyl transferase, and lactate dehydrogenase were measured in 56 adult healthy individuals. The data were grouped according to sex and body weight. For all variables, no significant differences were found between male and female mynahs, and body weight had no effect on lipoprotein fractions. Our results provide a reliable reference interval for some blood biochemical values in healthy common mynahs that could be useful for diagnosing and monitoring disease problems.

Variation in developmental trajectories of physiological and somatic traits in a common songbird approaching fledging.

In avian species, little is known about the development of physiological traits in the days preceding fledging, a critical life history transition marked by a high mortality rate. Developmental trajectory during this period may be flexible based on ecological context or hardwired, with potential costs for variation in growth in the form of oxidative stress. Patterns in development are likely to relate to variation in life history, for which seabirds and aerial insectivores have been well studied, while our focal species is a grassland ground forager, the European starling (Sturnus vulgaris). We show that changes in haematocrit, body mass and wing length are independent of year and brood quality, while changes in haemoglobin concentration are higher in low-quality broods. Moreover, we also identify higher oxidative stress in low-quality year and second broods, a potential cost for maintaining a hardwired developmental trajectory in a lower quality environment. Finally, we experimentally test the effects of food supplementation on development and maturity of chicks at fledging to show that although food increases body mass early in development, it does not change the trajectory or final maturity of chicks at fledging. Collectively this study demonstrates that some developmental changes prior to fledging may be hardwired, but may have long-term oxidative costs in low-quality environments.

Detecting Methemoglobinemia in Animals with a Drop of Blood.

A major concern during pesticide development and use is the impact on non-target species, such as raptors or domestic cats and dogs. Sodium nitrite and para-aminopropiophenone (PAPP) are two toxicants currently being studied for the control of invasive species, such as starlings and feral swine. When given to an animal these compounds oxidize hemoglobin, which renders it unable to carry oxygen resulting in methemoglobinemia. This study developed a method to estimate methemoglobin levels in mammals and birds by examining the efficacy of sodium nitrite to induce the conversion of hemoglobin to methemoglobin. Varying concentrations of sodium nitrite were added to aliquots of coyote, vole, feral swine, starling, and duck blood, collected from captive animals. The blood samples were analyzed spectrophotometrically to determine percent methemoglobin and digitally to determine red color values (RCV) associated with different methemoglobin levels. The avian and mammalian blood reached 100% methemoglobin levels at 200 mM and 15 mM sodium nitrite, respectively. All animals had similar RCV for a given percent methemoglobin. In conclusion, this study developed a procedure to quickly determine methemoglobin levels in mammals and birds. Furthermore, percent methemoglobin can be estimated with one standard curve from any animal species and an image of a blood spot. The technique will be useful during field studies, in agricultural areas, or in a veterinarian's office for the rapid diagnosis of methemoglobinemia in non-target animals that have eaten toxicants/baits or baited animals.

Contributions of testosterone and territory ownership to sexually-motivated behaviors and mRNA expression in the medial preoptic area of male European starlings.

Animals integrate social information with their internal endocrine state to control the timing of behavior, but how these signals are integrated in the brain is not understood. The medial preoptic area (mPOA) may play an integrative role in the control of courtship behavior, as it receives projections from multiple sensory systems, and is central to the hormonal control of courtship behavior across vertebrates. Additionally, data from many species implicate opioid and dopaminergic systems in the mPOA in the control of male courtship behavior. We used European starlings to test the hypothesis that testosterone (T) and social status (in the form of territory possession) interact to control the timing of courtship behavior by modulating steroid hormone-, opioid- and dopaminergic-related gene expression in the mPOA. We found that only males given both T and a nesting territory produced high rates of courtship behavior in response to a female. T treatment altered patterns of gene expression in the mPOA by increasing androgen receptor, aromatase, mu-opioid receptor and preproenkephalin mRNA and decreasing tyrosine hydroxylase mRNA expression. Territory possession did not alter mRNA expression in the mPOA, despite the finding that only birds with both T and a nesting territory produced courtship behavior. We propose that T prepares the mPOA to respond to the presence of a female with high rates of courtship song by altering gene expression, but that activity in the mPOA is under a continuous (i.e. tonic) inhibition until a male starling obtains a nesting territory.

Two seconds is all it takes: European starlings (Sturnus vulgaris) increase levels of circulating glucocorticoids after witnessing a brief raptor attack.

Researchers typically study "acute" activation of the hypothalamic-pituitary-adrenal (HPA) axis by measuring levels of circulating glucocorticoids in animals that have been exposed to a predator or a cue from a predator (e.g., odor), or have experienced a standardized capture-and-restraint protocol, all of which are many minutes in duration. However, exposure to predators in the "wild", either as the subject of an attack or as a witness to an attack, is generally much shorter as most depredation attempts upon free-living animals last <5s. Yet, whether a stimulus lasting only seconds can activate the HPA axis is unknown. To determine if a stimulus of a few seconds triggers a glucocorticoid response, we measured levels of corticosterone (CORT; the primary avian glucocorticoid) in wild-caught European starlings (Sturnus vulgaris) after they witnessed a brief (<2-8s) raptor attack upon a conspecific, a human "attack" (i.e., a researcher handling a conspecific), and an undisturbed control. Witnesses of a raptor attack responded with CORT levels comparable to that induced by a standardized capture-and-restraint protocol. Glucocorticoid levels of individuals following the control treatment were similar to baseline levels, and those that witnessed a human "attack" had intermediate levels. Our results demonstrate that witnessing a predator attack of very brief duration triggers a profound adrenocortical stress response. Given the considerable evidence of a role for glucocorticoids in learning and memory, such a response may affect how individuals learn to recognize and appropriately react to predators.

Reproductive state modulates testosterone-induced singing in adult female European starlings (Sturnus vulgaris).

European starlings (Sturnus vulgaris) exhibit seasonal changes in singing and in the volumes of the neural substrate. Increases in song nuclei volume are mediated at least in part by increases in day length, which is also associated with increases in plasma testosterone (T), reproductive activity, and singing behavior in males. The correlations between photoperiod (i.e. daylength), T, reproductive state and singing hamper our ability to disentangle causal relationships. We investigated how photoperiodic-induced variation in reproductive state modulates the effects of T on singing behavior and song nuclei volumes in adult female starlings. Female starlings do not naturally produce measureable levels of circulating T but nevertheless respond to exogenous T, which induces male-like singing. We manipulated photoperiod by placing birds in a photosensitive or photorefractory state and then treated them with T-filled or empty silastic implants. We recorded morning singing behavior for 3 weeks, after which we assessed reproductive condition and measured song nuclei volumes. We found that T-treated photosensitive birds sang significantly more than all other groups including T-treated photorefractory birds. All T-treated birds had larger song nuclei volumes than with blank-treated birds (despite photorefractory T-treated birds not increasing song-rate). There was no effect of photoperiod on the song nuclei volumes of T-treated birds. These data show that the behavioral effects of exogenous T can be modulated by reproductive state in adult female songbirds. Furthermore, these data are consistent with other observations that increases in singing rate in response to T are not necessarily due to the direct effects of T on song nuclei volume.

Evidence for baseline glucocorticoids as mediators of reproductive investment in a wild bird.

Determining the mechanisms that mediate investment decisions between current and future reproductive attempts is still a key goal of life-history studies. Since baseline levels of stress hormones (glucocorticoids - GCs) act as predictive and labile regulators of daily energetic balance in vertebrates they remain excellent candidates for mediating investment decisions both within and across reproductive attempts. Using free-living female European starlings (Sturnus vulgaris) we experimentally reduced investment in current reproduction (number of offspring raised in the first brood) to examine whether baseline corticosterone (CORT) acted as a hormonal mediator preparing individuals for a predictable increase in future investment (number of offspring raised in the second brood). Although treatment and control birds raised the same total amount of offspring across two broods, the experimental birds increased reproductive investment in second broods to compensate for the reduced investment in the first brood. Data on both mean and intra-individual changes in baseline CORT support the idea that an increase in baseline CORT between the incubation stages in treatment birds strongly predicted this increase in investment. Importantly, we measured the increase in baseline CORT during late incubation prior to the increase in energetic demand associated with increased reproductive investment in offspring, indicating that flexible within-individual changes in baseline GCs can act as a labile mechanism preparing individuals for predictable increases in reproductive investment. As such, our experimental results indicate that elevated baseline GCs can prepare individuals for investment in energetically expensive life-history stages, rather than simply being elevated as a consequence of increased effort or demand. This suggests that short-term preparative increases in baseline GCs benefit individuals by successfully allowing them to maximize fitness under varying environmental conditions.

Testosterone increases repertoire size in an open-ended learner: an experimental study using adult male European starlings (Sturnus vulgaris).

Song in songbirds is a learned secondary sexual behavior, first acquired during a sensitive phase of juvenile development, which is affected by hormones such as testosterone (T). While the latter has received much attention, the potential involvement of T in the adult repertoire changes observed in a number of species is much less understood. Yet, this may prove essential to understand the role of song as a sexually selected trait. We therefore performed a T-implantation experiment during the non-breeding season (when T is basal), using adult male European starlings (Sturnus vulgaris), a songbird species in which song repertoire size (and composition) changes seasonally and increases with age. Repertoire size increased rapidly in T-males, but not in control males, indicating a role for T in repertoire size changes. This increase resulted from a lower proportion of dropped song types in T-males than in control males, while the proportion of added song types did not differ between both groups. Interestingly, the observed repertoire turnover (adding and removing song types from the repertoire) in both groups, suggests that elevated plasma T levels were not essential for changes in repertoire composition (contrary to repertoire size). Finally, T-males (but not control males) significantly increased their song rate, while neither group showed a significant change in their song bout length and phrase repetition rate. Taken together, our results suggest a role for T in adult song learning and provide new insights into the information content of repertoire size and song bout length as sexually selected traits.

DHEA and estradiol levels in brain, gonads, adrenal glands, and plasma of developing male and female European starlings.

Traditionally, sexual differentiation of the brain was thought to be driven by gonadal hormones, particularly testosterone (T). However, recent studies in songbirds suggest that other steroids may also be important. For example, dehydroepiandrosterone (DHEA) can be synthesized by the gonads, adrenal glands, and/or brain and locally metabolized into T and 17ß-estradiol (E(2)). Here, we examined DHEA and E(2) levels in the brain, peripheral tissues, and plasma of wild European starlings (Sturnus vulgaris). In Study 1, samples were collected from males and females at P0 (day of hatch), P6, and P8. In Study 2, samples were collected at P4. At P0, DHEA levels in the diencephalon were higher in males than females. DHEA levels were generally high in the gonads and adrenals, and they were higher in testes than ovaries at P8. Further, E(2) levels were non-detectable in most brain samples, suggesting that DHEA was not metabolized to E(2) or that locally produced E(2) was rapidly inactivated. At P4, DHEA levels in telencephalic regions were lower in males than females. Taken together, these data suggest that sex differences in peripheral DHEA secretion and neural DHEA metabolism at specific ages during development might play a role in sexual differentiation of the songbird brain.

Seasonal changes in courtship behavior, plasma androgen levels and in hypothalamic aromatase immunoreactivity in male free-living European starlings (Sturnus vulgaris).

In songbirds from temperate latitudes, singing during spring has an essential role in mate attraction, while during the non-breeding season it is connected to territorial aggression and/or maintaining dominance hierarchies or flock cohesion. Courtship behavior is regulated by plasma testosterone (T) levels. Other androgens, like dehydroepiandrosterone (DHEA) could be responsible for aggression. The aromatization of androgens in the brain is an essential step in mediating their effects on behavior. Our goal was to determine whether the seasonal changes in male courtship behavior (measured by average song bout length and wing-waving/flicking) are related to seasonal changes in androgen activity (measured by plasma T, DHEA levels) and aromatase (ARO) immunoreactivity in the preoptic area/medial preoptic nucleus (POA/POM) of free-living male starlings. DHEA increased during pair formation, decreased at nesting and remained at low levels. The number of ARO cells - in line with the T levels - increased during the courtship and nesting periods, but outside the breeding season it was low. Song bout length showed a similar pattern, namely the peak was reached during the courtship period, and after that males stopped singing when chicks started to hatch. Short and fast wing-flicking and wing-waving behavior was observed only during the breading season. Summarizing, we have found that song bout length of male starlings changes parallel with plasma T levels and ARO immunoreactivity in the POA/POM. Furthermore, DHEA levels were low during the sexually inactive period which suggests that other mechanisms could be involved in the aggressive non-courtship behavior/vocalization in these birds.

Seasonal changes in concentrations of plasma LH and prolactin associated with the advance in the development of photorefractoriness and molt by high temperature in the starling.

In a study on starlings (Sturnus vulgaris) kept on a simulated annual cycle in photoperiod, temperature had no effect on the timing or rate of testicular maturation but high temperature resulted in an advance in the timing of testicular regression and molt (Dawson, 2005). This study asks whether the earlier gonadal regression in response to higher temperature represents a central neuroendocrine response to temperature, and secondly, whether prolactin plays a role in the earlier regression. Castrated starlings were kept on a simulated annual cycle of photoperiod at either 8 or 18 degrees C. Circulating LH and prolactin concentrations were measured and the progress of the post-nuptial molt was recorded as an external indicator of the development of photorefractoriness. Additionally plasma prolactin was measured in samples taken from intact male and female starlings in the 2005 study. In castrated birds, LH concentrations decreased three weeks earlier at 18 degrees C. These birds also showed the same three week advance in molt as males and females in the earlier study. This demonstrates that the advance in regression caused by higher temperatures probably results from a central neuroendocrine mechanism, i.e., an advance in photorefractoriness, rather than an effect at the level of the gonads. Temperature had a highly significant effect on the changes in prolactin - peak prolactin occurred three weeks earlier at 18 degrees C. However, there was no clear consistent significant difference in prolactin between the two temperatures in advance of the onset of photorefractoriness, so the advance in photorefractoriness may not be mediated by prolactin. The higher temperature resulted in a significantly earlier decrease in prolactin and this may be causally related to the advance in molt.

Chronic psychological stress alters body weight and blood chemistry in European starlings (Sturnus vulgaris).

One hallmark of chronic stress is a decrease in body weight that rebounds once chronic stress is alleviated. We applied chronic psychological stress by exposing European starlings (Sturnus vulgaris) to a previously validated chronic stress protocol (CSP) consisting of 4 different randomly applied stressors per day. Experimental design consisted of a 21 day CSP (CSP1), a 60day recovery (R1), a second 14 day CSP (CSP2), and a second 30 day recovery (R2). Body weight decreased by approximately 5% during CSP1, but overshot to 5-10% above initial body weight during R1. To investigate underlying mechanisms, we periodically measured 12 biochemical analytes, including aspartate aminotransferase (AST), creatine kinase (CK), bile acids, total protein, albumin, globulin, glucose, uric acid, calcium (Ca(++)), phosphorus (PHOS), potassium (K(+)), and sodium (Na(+)). AST and CK increased at the beginning of CSP1, suggesting muscle breakdown. Additionally, decreases in albumin and total protein paired with stable uric acid, but no associated change in glucose, suggested protein breakdown as a secondary energy source. Changes in blood parameters that occurred during CSP1 did not reverse during R1. During CSP2 and R2, weight loss and gain occurred in different proportions. CSP2 produced an approximate 15% decrease in body weight, but R2 resulted in only re-gaining 5% of this weight, although this was equivalent to the pre-CSP1 weight. In summary, protein metabolism appeared to mediate weight loss during chronic stress, but over-gaining weight was not a good indicator of recovery.

Structural changes between seasons in the songbird auditory forebrain.

The song control system (SCS) of seasonal songbirds shows remarkable seasonal plasticity. Male starlings (Sturnus vulgaris) sing throughout the year, but in the breeding season, when concentrations of testosterone are elevated, the song is highly sexually motivated. The main goal of this study was to investigate structural seasonal changes in regions involved in auditory processing and in socio-sexual behavior. Using in vivo Diffusion Tensor Imaging (DTI), we measured in breeding and nonbreeding seasons volume and tissue characteristics of several brain regions of nine adult male starlings. We demonstrate that the songbird brain exhibits an extreme seasonal plasticity not merely limited to the SCS. Volumetric analysis showed seasonal telencephalon volume changes and more importantly also a volumetric change in the caudal region of the nidopallium (NCM), a region analogous to the mammalian secondary auditory cortex. Analysis of the DTI data allowed detection of seasonal changes in cellular attributes in NCM and regions involved in social behavior. This study extends our view on a seasonally dynamic avian brain which not only hones its song control system but also auditory and social systems to be prepared for the breeding season.

Reproductive conflict and the costs of social status in cooperatively breeding vertebrates.

Conflict over reproduction is an inherent part of group living. In many social vertebrates, conflict may be reflected as allostatic load, or the costs of social status and dominance rank, which may be quantified by measuring glucocorticoid stress hormones. Here, we develop the first quantitative model of allostatic load based on the tug-of-war model of reproductive skew to generate insights into the mechanisms underlying reproductive conflict in cooperative breeders and to determine whether glucocorticoids can be used to assess conflict levels in group-living vertebrates. It predicts that subordinates have higher allostatic loads than dominants under most conditions, but when body condition is lower in dominants than in subordinates, dominants experience higher allostatic load. Group structure is also important, as dominants generally have higher allostatic loads than subordinates when there is a large number of subordinates in the group, but this cost can be reduced by increasing the number of dominants, as in plural breeding societies. Using glucocorticoid data from cooperatively breeding superb starlings Lamprotornis superbus, we found empirical support for both predictions. Our model is useful for understanding how the costs of social status influence reproductive sharing, and it suggests that glucocorticoids can be used to examine reproductive conflict and cooperation in social species.

Increased energy expenditure but decreased stress responsiveness during molt.

Baseline and stress-induced corticosterone (CORT), heart rate (fH), and energy expenditure were measured in eight captive European starlings Sturnus vulgaris during and following a prebasic molt. The fH and oxygen consumption (V O2 ) were measured simultaneously across a range of heart rates, and energy expenditure (kJ/d) was then calculated from data. Energy expenditure and fH were strongly and positively correlated in each individual. Baseline fH and energy expenditure were significantly higher during molt. Molting starlings expended 32% more energy over 24 h than nonmolting birds, with the most significant increase (60%) occurring at night, indicating a substantial energetic cost to molt. Furthermore, the cardiac and metabolic responses to stress during molt were different than during nonmolt. Birds were subjected to four different 30-min acute stressors. The fH and CORT responses to these stressors were generally lower during molt. Although restraint caused a 64% increase in daily energy expenditure during nonmolt, no other stressor caused a significant increase in energy expenditure. Overall, our data suggest that molt is not only energetically expensive but that it also alters multiple stress response pathways. Furthermore, most acute stressors do not appear to require a significant increase in energy expenditure.

The effect of chronic psychological stress on corticosterone, plasma metabolites, and immune responsiveness in European starlings.

Although increases in glucocorticoid concentrations during acute stress are believed to help animals survive stressful events, chronic changes in glucocorticoid concentrations can alter metabolism and lead to disease. We studied the effect of chronic psychological stress on corticosterone (CORT), corticosterone binding globulin (CBG), glucose, and triglyceride concentrations as well as immune responsiveness to a T-cell mitogen challenge in European starlings, Sturnus vulgaris. To induce chronic stress we used a chronic stress protocol consisting of five stressors (loud radio, cage tapping, cage rolling, human voice, and bag restraint) administered in random order for 30 min for 4 times/day over 18 days. Total CORT decreased throughout the chronic stress period, which parallels a previous study with starlings. CBG capacity did not significantly change with chronic stress, thus free CORT followed the same pattern of attenuation as total CORT during chronic stress. Despite the change in regulation of CORT release, daytime glucose and triglyceride concentrations did not change with chronic stress. In addition, immune responsiveness in chronically stressed and unstressed birds was similar. Our results, together with a previous study using a similar CSP in European starlings, suggest that starlings physiologically dampen the HPA axis during chronic psychological stress to avoid pathology associated with chronically augmented CORT concentrations such as hyperglycemia and impaired immune function.

Chronic stress in free-living European starlings reduces corticosterone concentrations and reproductive success.

Chronic increases in stress hormones such as glucocorticoids are maladaptive, yet studies demonstrating a causal relationship among chronic stress, increases in glucocorticoid concentrations, and subsequent fitness costs in free-living animals are lacking. We experimentally induced chronic psychological stress in female European starlings (Sturnus vulgaris) by subjecting half of the females at our study site to a chronic stress protocol consisting of 4, 30 min stressors (loud radio, predator calls, a novel object, or predator decoys including a snake, rat, and owl) administered in random order daily for 8 days after clutch completion. Experimental females were captured at the end of the chronic stress protocol (9 days after the onset of the chronic stress protocol), and unstressed control females were captured at the same stage of the nesting cycle. Chronically stressed females had lower baseline corticosterone (CORT, the avian glucocorticoid) concentrations and lower reproductive success than unstressed females. Furthermore, surviving nestlings in experimentally stressed broods showed sensitization of the CORT response to acute stress, which is a physiological change that could persist to adulthood. Attenuation of baseline CORT concentrations in adult females is contrary to the general assumption that elevated CORT concentrations indicate stress, suggesting that more research is necessary before CORT concentrations can be used to accurately assess chronic stress in field studies.

Accumulation, tissue-specific distribution and debromination of decabromodiphenyl ether (BDE 209) in European starlings (Sturnus vulgaris).

In this study we investigated the accumulation, tissue-specific distribution and possible debromination of BDE 209 in a terrestrial songbird species, the European starling, using silastic implants as a method of exposure. BDE 209 accumulated in the blood of the exposed starlings to a mean peak concentration of 16+/-4.1 ng/ml on day 10. After this peak, there was a decline to 3.3+/-0.4 ng/ml blood at the end of the exposure period of 76 days, which suggests elimination of BDE 209. In the exposed group, the muscle concentrations (461 ng/g lipid weight [lw], 430 ng/g lw) were about twofold those in liver (269 ng/g lw, 237 ng/g lw). In addition to BDE 209, other PBDE congeners, particularly octa- and nonaBDEs, were also present in the muscle and liver, suggesting bioformation from BDE 209. To our knowledge, these results are the first indications for the debromination of BDE 209 in birds.