This proposal is a request for an ADAMHA RSDA Level II. I am currently an RSDA Level I awardee. This continuation proposal focuses on the importance of the circadian rhythm in cortisol secretion in maintaining proper HPA axis regulation. This proposal involves both basic science animal work and human studies focusing on HPA axis dysregulation in depression. Previous studies by other investigators in rats have demonstrated that during the nadir of the cortisol rhythm when CRH is not driving the system, steroids are not necessary to prevent corticotroph secretion. During this nadir the levels of corticosterone falls to extremely low levels; this fall may be necessary to prevent glucocorticoid receptor desensitization. Studies in this application propose the administration of corticosterone during the trough of circadian rhythm in rats and subsequent evaluation of negative feedback in vivo and in vitro. The length of time that corticosterone will remain elevated will be varied by the use of either short acting (corticosterone in saline) or long acting (corticosterone in sesame oil and corticosterone pellets) preparations. Alterations in feedback secondary to changes in the level of corticosterone will be correlated with measures of glucocorticoid receptors in brain regions and pituitary. In humans the studies focus on evaluating the level of endogenous CRH drive across the circadian rhythm and evaluating the interaction of CRH drive and steroid feedback at different points in the circadian rhythm in depressed subjects and normal controls. One series of studies involves the use of metyrapone, a 21 beta hydroxylase inhibitor to remove steroid feedback at different points in the circadian rhythm; another series of studies involves the infusion of cortisol to turn off active secretion at different points in the circadian rhythm. The final series compares circadian variation in CRH response both with and without steroid feedback. In addition to carrying out these studies I would like to build the basis for future studies and collaborations. I would like to visit the laboratory of Wylie Vale to interact with Catherine Rivier, who is an expert on in vivo CRH studies and the physiology of stress. Should the future data suggest that changes in steroid feedback can occur without changes in glucocorticoid receptors, I plan to collaborate with local experts in the area of second messenger systems to explore possible mechanisms for glucocorticoids actions on release. Finally, I would like to use my ongoing collaboration with the laboratories of Drs. Watson and Akil in the area of molecular biology to pursue training in molecular biology techniques, since these techniques have the advantage of integrating information over a longer time frame than measurement of peptide stores. With the development of intron specific probes, it is now possible to detect increases in gene transcription which can supplement the longer term information on mRNA levels. I will focus on these two measures of mRNA during this collaboration. I also plan to continue collaboration and discussions with my mentors from the two previous outside training periods, Drs. Mary Dallman and Bruce McEwen.
