The overall goals of this application are to improve our understanding of both the function and regulation of corticotropin-releasing hormone. The hypothalamic-pituitary-adrenal (HPA) axis is an important system mediating many of the body's responses to stress, and CRH is a central component in regulation of the HPA axis. The role and relative importance of CRH in comparison with other regulators of ACTH release, e.g. vasopressin or catecholamines are, however, currently unclear. In addition to its role in regulation of ACTH release, CRH may serve other important functions. CRH and its receptor are widely distributed in the brain, where it may act as a neuromodulator, perhaps mediating some forms of stress-related behavior. CRH is also present in the immune system, where it has been shown to both suppress and augment inflammatory reactions. The study of the regulation of CRH synthesis and release in these physiologic roles has been hampered by the unavailability of suitable cellular models. To address these issues, three lines of investigation are proposed. First, a CRH-deficient mouse model will be created inactivating the endogenous murine CRH gene via homologous recombination in embryonic stem cells. With this model, the role of CRH in development of pituitary corticotrophs and the response of the HPA axis to stress and circadian stimuli will be evaluated. Second, immortalized CRH-producing cell lines to directly investigate mediators of CRH release, and to provide a suitable model system with which to study CRM regulation will be established. Third, the function and regulation of CRH synthesized within the immune system will be investigated. The sponsor, Dr. Joseph Majzoub, is a recognized leader in the molecular biology of CRH. Dr. David Wight, collaborating on development of transgenic mouse lines, has successfully generated numerous transgenic animals in the past. Dr. Raif Geha, who will serve as a consultant on the immune function of CRH, is a prominent immunologist with considerable expertise on mechanisms of immune activation. The proposed studies allow definition of a crucial aspect of the body's response to stress, while allowing me training in techniques of embryo manipulation and gene transfer technology. These techniques will be central to my long term goal of studying the development of the neuroendocrine system.