15 Environmental signals regulate a variety of key processes in animal physiology. In many genera, environmental signals such as changes in day length or temperature signal the onset or termination of reproduction (Steger and Bartke, 1996), thus ensuring that reproduction occurs at a time that maximizes the survival of the offspring (Bronson, 1985). These environmental signals typically stimulate the release of neuro- endocrine signaling molecules that initiate endocrine cascades culminating in physiological responses to the environmental cue. A well characterized neuro-endocrine response to environmental signaling is the detection of photoperiod changes by the pineal gland and its regulation of hypothalamic- pituitary cascades via the action of melatonin in vertebrates (Steger and Bartke, 1996). Similar photoperiod-initiated neuro- endocrine cascades have been identified in insects, though the organs, hormones and regulated events differ significantly from those characterized in vertebrates (Horie et al., 2000; Yamashita, 1996). The terpenoid hormone methyl farnesoate is emerging in crustaceans as a major hormone responsible for transducing environmental signals. Methyl farnesoate is synthesized by the mandibular organ of Decapod crustaceans (Laufer et al., 1987) and its secretion is negatively regulated by members of the crustacean hyperglycemic hormone (CHH) family of neuropeptides that are synthesized by the X-organ/sinus gland complex (Liu and Laufer, 1996). Methyl farnesoate is structurally similar to juvenoid hormones of insects and retinoid hormones of vertebrates, and its activity is probably mediated through interaction with a nuclear receptor. Methyl farnesoate has been associated with a variety of physiological processes in crustaceans related to reproduction, including testicular maturation (Kalavathy et al., 1999), The Journal of Experimental Biology 208, 15-23 Published by The Company of Biologists 2005 doi:10.1242/jeb.01343 Environmental signals can activate neuro-endocrine cascades that regulate various physiological processes. In the present study, we demonstrate that two responses to environmental stress signaling in the crustacean Daphnia magna – hemoglobin accumulation and male offspring production – are co-elevated by the crustacean terpenoid hormone methyl farnesoate and several synthetic analogs. Potency of the hormones with respect to the induction of both hemoglobin and male offspring was highly correlated, suggesting that both processes are regulated by the same terpenoid signaling pathway. Six clones of the D. pulex/pulicaria species complex that were previously characterized as unable to produce male offspring and five clones that were capable of producing males were evaluated for both hemoglobin induction and male offspring production in response to methyl farnesoate. Four of the five male-producing clones produced both hemoglobin and male offspring in response to the hormone. Five of the six non-male-producing clones produced neither hemoglobin nor males in response to the hormone. These results provide additional evidence that both physiological processes are regulated by the same signaling pathway. Furthermore, the results indicate that the non-male-producing clones are largely defective in some methyl farnesoate signaling component, downstream from methyl farnesoate synthesis but upstream from the genes regulated by the hormone. A likely candidate for the site of the defect is the methyl farnesoate receptor. As a consequence of this defect, non-male-producing clones have lost their responsiveness to environmental signals that are transduced by this endocrine pathway. This defect in signaling would be likely to enhance population growth in stable environments due to the elimination of males from the population, assuming that other processes critical to population growth are not also compromised by this defect. Key words: Cladocera, juvenoid, endocrine disruption, evolution, nuclear receptor, Daphnia magna. Summary Introduction Stress signaling: coregulation of hemoglobin and male sex determination through a terpenoid signaling pathway in a crustacean Cynthia V. Rider 1 , Thomas A. Gorr 2 , Allen W. Olmstead 1 , Beth A. Wasilak 1 and Gerald A. LeBlanc 1, * 1 Department of Environmental and Molecular Toxicology, North Carolina State University, Raleigh, NC 27695-7633, USA and 2 Division of Hematology, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA 02115, USA *Author for correspondence (e-mail: ga_leblanc@ncsu.edu) Accepted 13 October 2004 THEJOURNALOFEXPERIMENTALBIOLOGY