Endocrine parameters and heavy metals in storks affect the HPA axis function of wild birds. The association we found occurs at Pb levels below those reported to cause sublethal effects in avian species, and further supports the idea that HPA axis–related end points may be use- ful indicators of metal exposure and potential toxicity in wild animals. Experimental studies in this regard are strongly encouraged. Our results indicate that a simple handling and restraint protocol may be a better approach to assess the HPA axis function than a single measurement taken after capture. Plasma corticosterone is a static measure by itself and does not provide information concerning the dynamic nature of the system (Hinson and Raven 2006; Norris 2000). This work also supports the idea that greater attention to the problems arising from interactions between different types of stressors must certainly be considered, especially when working in the wild, given that such a scenario (and not the exposure to isolated sources of stress) constitutes the environmental reality. The persistence of Pb, combined with the sensitivity of the HPA axis to contaminant insult, suggests that exposure to this metal even at sublethal levels may cause delayed functional deficits later in development. Endangered species and/or populations exposed to addi- tional stress factors (e.g., nutrition deﬁciency, parasitism) would be of special concern. Further ﬁeld studies involving birds from these and other metal-polluted sites are required to determine the extent to which the cortico- sterone response systems of wild birds are affected by exposure to Pb as well as the long- term effects on survival and overall ﬁtness. Thyroid status was not related to metal pollution, although we found differences between colony locations that deserve further investigation. REFERENCES ATSDR (Agency for Toxic Substances and Disease Registry). 1999. Toxicological Proﬁle for Lead. Available: http://www. atsdr.cdc.gov/toxproﬁles/tp13.html [accessed 16 June 2006]. ATSDR (Agency for Toxic Substances and Disease Registry). 2005. 2005 CERCLA Priority List of Hazardous Substances. Available: http://www.atsdr.cdc.gov/cercla/05list.html [accessed 16 June 2006]. Baos R, Jovani R, Forero MG, Tella JL, Gómez G, Jiménez B, et al. 2006. Relationships between T-cell-mediated immune response and Pb, Zn, Cu, Cd, and As concentrations in blood of nestling white storks (Ciconia ciconia) and black kites (Milvus migrans) from Doñana (southwestern Spain) after the Aznalcóllar toxic spill. Environ Toxicol Chem 25(4):1153–1159. Barrot M, Marinelli M, Abrous DN, Rouge-Pont F, Le Moal M, Piazza PV. 2000. The dopaminergic hyper-responsiveness of the shell of the nucleus accumbens is hormone-dependent. Eur J Neurosci 12:973–979. Benito V, Devesa V, Muñoz O, Suñer MA, Montoro R, Baos R, et al. 1999. Trace elements in blood collected from birds feeding in the area around Doñana National Park affected by the toxic spill from the Aznalcóllar mine. Sci Total Environ 242:309–323. Beyer WN, Dalgarn J, Dudding S, French JB, Mateo R, Miesner J, et al. 2004. Zinc and lead poisoning in wild birds in the Tri-State mining district (Oklahoma, Kansas, and Missouri). Arch Environ Contam Toxicol 48:108–117. Blas J, Baos R, Bortolotti GR, Marchant T, Hiraldo F. 2005. A multi-tier approach to identifying environmental stress in altricial nestling birds. Funct Ecol 19:315–322. Blas J, Baos R, Bortolotti GR, Marchant TA, Hiraldo F. 2006. Age- related variation in the adrenocortical response to stress in nestling white storks (Ciconia ciconia) supports the develop- mental hypothesis. Gen Comp Endocrinol 148:172–180; doi:10.1016/j.ygcen.2006.02.011 [Online 19 April 2006]. Burger J, Gochfeld M. 1997. Age differences in metals in the blood of herring (Larus argentatus) and Franklin’s (Larus pipixcan) gulls. Arch Environ Contam Toxicol 33:436–440. Chozas P. 1983. Estudio general sobre la dinámica de población de la cigüeña Ciconia ciconia en España [PhD Thesis]. Madrid:Universidad Complutense de Madrid. Cory-Slechta DA, McCoy L, Richﬁeld EK. 1997. Time course and regional basis of Pb-induced changes in MK-801 binding: reversal by chronic treatment with the dopamine agonist apomorphine but not the D1 agonist SKF-28958. J Neurochem 68:2012–2023. Cory-Slechta DA, Virgolini MB, Thiruchelvam M, Weston DD, Bauter MR. 2004. Maternal stress modulates the effects of developmental lead exposure. Environ Health Perspect 112:717–730. Cramp S, Simmons KEL. 1980. The Birds of the Western Palearctic. Oxford, UK:Oxford University Press. Damstra T, Barlow S, Bergman A, Kavlock R, Van der Kraak G, eds. 2002. Global Assessment of the State-of-the-Science of Endocrine Disruptors. Geneva:World Health Organization, International Programme on Chemical Safety. Available: http://www.who.int/ipcs/publications/new_issues/ endocrine_disruptors/en/ [accessed 17 August 2006]. Der R, Yousef M, Fahim Z, Fahim M. 1977. Effects of lead and cadmium on adrenal and thyroid function in rats. Res Comm Chem Pathol Pharmacol 17:237–253. Di Giulio RT, Scanlon PF. 1985. Effects of cadmium ingestion and food restriction on energy metabolism and tissue metal concentrations in mallard ducks (Anas platyrhynchos). Environ Res 37:433–444. Eeva T, Hasselquist D, Langefors A, Tummeleht L, Nikinmaa M, Ilmonen P. 2005. Pollution related effects on immune func- tion and stress in a free-living population of pied ﬂycatcher Ficedula hypoleuca. J Avian Biol 36:405–412. Eeva T, Lehikoinen E, Nikinmaa M. 2003. Pollution-induced nutritional stress in birds: an experimental study of direct and indirect effects. Ecol Appl 13:1242–1249. Ellegren H. 1996. First gene on the avian W chromosome (CHD) provides a tag for universal sexing of nonratite birds. Proc R Soc Lon B 263:1635–1641. Fox GA. 2001. Wildlife as sentinels of human health effects in the Great Lakes–St. Lawrence Basin. Environ Health Perspect 109(suppl 6):853–861. Franson J. 1996. Interpretation of tissue lead residues in birds other than waterfowl. In: Environmental Contaminants in Wildlife: Interpreting Tissue Concentrations (Beyer NW, Heinz GH, Redmon-Norwood AW, eds). Boca Raton, FL:Lewis Publishers, 265–279. Green A. 2001. Mass/length residuals: measures of body condition or generators of spurious results? Ecology 82:1473–1483. Grimalt JO, Ferrer M, Macpherson E. 1999. The mine tailing accident in Aznalcóllar. Sci Total Environ 242:3–11. Hinson JP, Raven PW. 2006. Effects of endocrine-disrupting chemicals on adrenal function. Best Pract Res Clin Endocrinol Metab 20(1):111–120. Kendall RJ, Dickerson RL, Giesy JP, Suk W, eds. 1998. Principles and Processes for Evaluating Endocrine Disruptors in Wildlife. Pensacola, FL:SETAC Press. Kerr DS, Campbell LW, Applegate MD, Brodish A, Landfield PW. 1991. Chronic stress-induced acceleration of electro- physiologic and morphometric biomarkers of hippocampal aging. J Neurosci 11:1316–1324. Kitaysky AS, Kitaiskaia EV, Piatt JF, Wingfield JC. 2003. Beneﬁts and costs of increased levels of corticosterone in seabird chicks. Horm Behav 43:140–149. Leatherland JF. 2000. Contaminant-altered thyroid function in wildlife. In: Environmental Endocrine Disrupters (Guillette L, Crain DA, eds). New-York:Taylor and Francis, 155–181. Littell RC, Stroup WW, Milliken GA, Wolfinger RD. 1996. Generalized linear mixed models. In: SAS System for Mixed Models. Cary, NC:SAS Institute Inc., 423–459. Lorenzen A, Moon TW, Kennedy SW, Fox GA. 1999. Relationships between environmental organochlorine contaminant residues, plasma corticosterone concentrations, and inter- mediary metabolic enzyme activities in Great Lakes herring gull embryos. Environ Health Perspect 107:179–186. Mayne GJ, Bishop CA, Martin PA, Boermans HJ, Hunter B. 2005. Thyroid function in nestling tree swallows and east- ern bluebirds exposed to non-persistent pesticides and p, p’-DDE in apple orchards of southern Ontario, Canada. Ecotoxicology 14:381–396. Mayne GJ, Martin PA, Bishop CA, Boermans HJ. 2004. Stress and immune responses of nestling tree swallows (Tachycineta bicolor) and eastern bluebirds (Sialis sialis) exposed to non- persistent pesticides and p,p´-dichlorodiphenyldichloro- ethylene in apple orchards of southern Ontario, Canada. Environ Toxicol Chem 23:2930–2940. McArthur MLB, Fox GA, Peakall DB, Philogène BJR. 1983. Ecological signiﬁcance of behavioral and hormonal abnor- malities in breeding ring doves fed an organochlorine chemical mixture. Arch Environ Contam Toxicol 12:343–353. McNabb FMA. 2000. Thyroids. In: Sturkie’s Avian Physiology (Whittow GC, ed). New York:Academic Press, 461–471. Mullner A, Linsenmair KE, Wikelski M. 2004. Exposure to eco- tourism reduces survival and affects stress response in hoatzin chicks (Opisthocomus hoazin). Biol Conserv 118:549–558. Norris DO. 2000. Endocrine disruptors of the stress axis in natural populations: how can we tell? Am Zool 40:393–401. Pastor N, Baos R, López-Lázaro M, Jovani R, Tella JL, Hajji N, et al. 2004. A 4-year follow-up analysis of genotoxic damage in birds of the Doñana area (SW Spain) in the wake of the 1998 mining waste spill. Mutagenesis 19:61–65. Refowitz RM. 1984. Thyroid function and lead: no clear relation- ship. J Occup Med 26:579–583. Robins JM, Cullen MR, Connors BB, Kayne RD. 1983. Depressed thyroid indexes associated with occupational exposure to inorganic lead. Arch Int Med 143:220–224. Sapolsky RM, Romero LM, Munck AU. 2000. How do gluco- corticoids inﬂuence stress responses? Integrating permis- sive, suppressive, stimulatory, and preparative actions. Endocr Rev 21:55–89. SAS Institute, Inc. 2001. SAS/STAT Guide for Personal Computers, Version 8. Cary, NC:SAS Institute Inc. Scanes CG, McNabb FMA. 2003. Avian models for research in toxicology and endocrine disruption. Avian Poul Biol Rev 14:21–52. Schantz SL, Widholm JJ. 2001. Cognitive effects of endocrine- disrupting chemicals in animals. Environ Health Perspect 109:1197–1206. Scheuhammer AM. 1987. The chronic toxicity of aluminium, cadmium, mercury, and lead in birds: a review. Environ Poll 46:263–295. Silverin B. 1998. Territorial behaviour and hormones of pied ﬂy- catchers in optimal and suboptimal habitats. Anim Behav 56:811–818. Smits JE, Fernie KJ, Bortolotti GR, Marchant TA. 2002. Thyroid hormone suppression and cell-mediated immunomodulation in American kestrels (Falco sparverius) exposed to PCBs. Arch Environ Contam Toxicol 43:338–344. Smits JEG, Bortolotti GR, Baos R, Blas J, Hiraldo F, Xie Q. 2005. Skeletal pathology in white storks (Ciconia ciconia) asso- ciated with heavy metal contamination in southwestern Spain. Toxicol Pathol 33(4):441–448. Snoeijs T, Dauwe T, Pinxten R, Darras VM, Arckens L, Eens M. 2005. The combined effect of lead exposure and high or low dietary calcium on health and immunocompetence in the zebra ﬁnch (Taeniopygia guttata). Environ Poll 134:123–132. Van Eeden PH, Schoonbee HJ. 1996. Metal concentrations in liver, kidney, bone and blood of three species of birds from a metal-polluted wetland. Water SA 22:351–358. Vos JG, Dybing E, Greim HA, Ladefoged O, Lambré C, Tarazona JV, et al. 2000. Health effects of endocrine-disrupt- ing chemicals on wildlife, with special reference to the European situation. Crit Rev Toxicol 30:71–133. Vyskocil A, Fiala Z, Ettlerova E, Tenjnorova I. 1990. Inﬂuence of chronic lead exposure on hormone levels in developing rats. J Appl Toxicol 10:301–302. Wayland M, Gilchrist HG, Marchant T, Keating J, Smits JE. 2002. Immune function, stress response, and body condition in Arctic-breeding common eiders in relation to cadmium, mer- cury, and selenium concentrations. Environ Res A 90:47–60. Wayland M, Smits JEG, Gilchrist HG, Marchant T, Keating J. 2003. Biomarker responses in nesting, common eiders in the Canadian Arctic in relation to tissue cadmium, mercury and selenium concentrations. Ecotoxicology 12:225–237. Wingfield JC, Kitaysky AS. 2002. Endocrine responses to unpredictable environmental events: stress or anti-stress hormones? Integr Comp Biol 42:600–609. Environmental Health Perspectives • VOLUME 114 | NUMBER 10 | October 2006 1501