Aberrant adrenocortical disease (ACD) is a frequently found variant of Cushings syndrome in dogs. ACD is a disease in which cortisol usually remains at normal concentrations (which is misleading), but excessive adrenal intermediate steroids are secreted causing clinical signs of canine Cushings syndrome. The Clinical Endocrinology Service at the University of Tennessee is an internationally recognized center of excellence in ACD diagnostics currently testing over 1500 suspected ACD cases per year. Effective treatment options for ACD are limited. Melatonin currently is used to treat ACE in an effort to decrease secretion of intermediate adrenal steroid; however, its mechanism of action is unknown. Some dogs have a favorable clinical response to melatonin and other dogs do not respond to melatonin. In this proposed research, melatonin will be utilized in a cell culture system. Adrenal cells and canine adrenal tissue explants will be treated with melatonin and cell culture media will be analyzed for concentrations of progesterone, 17-hydroxyprogesterone, androstenedione, testosterone, estradiol, and cortisol (hormones analyzed in ACE patients). Adrenocorticotropic hormone-sensitive cell signaling pathways will be studied for melatonin targeting effects. Results will aid in elucidating melatonins possible mechanism of action on adrenal steroidogenesis.