Adrenocorticotropic hormone (ACTH) stimulates the secretion of glucocorticoids by acting upon melanocortin type 2 receptors (MC2R) located in the adrenal zona fasciculata. However, translocation of MC2R from the endoplasmic reticulum to its active state in the cell membrane requires its interaction with the melanocortin receptor associated protein (MRAP). To determine whether adrenocortical activation during stress involves regulation of MRAP, we examined the effects of restraint stress and ACTH injection on HPA axis activity, MC2R mRNA, and the levels of MRAP primary transcript (hnRNA) and mRNA. Blood levels of ACTH and corticosterone showed peaks at 15 min and 1 hour restraint, respectively, and both return to basal levels by 2 hours. Real time PCR analysis of MC2R mRNA levels showed a progressive decrease up to 2h, and returned near basal levels by 4h after restraint. Conversely, there were rapid increases in MRAP hnRNA and mRNA levels, with peaks at 15 min and 1h, respectively. Similar changes in MC2R mRNA and MRAP hnRNA and mRNA levels were observed after a single injection of ACTH (4ng) in methylprednisolone-suppressed rats, suggesting that the effects of stress are mediated by ACTH. Studies in collagenase dispersed rats adrenal fasciculata cells revealed a dose dependent effect of ACTH on MRAP hnRNA levels with an ED50 of about 0.1nM. Using 10nM ACTH, the maximal stimulation was at 30 min and levels remained elevated up to 2h. The PKA inhibitor, H89, prevented the stimulatory effect of all ACTH concentrations on MRAP hnRNA while the MAP kinase inhibitor, UO126 inhibited only the effect of low concentrations of ACTH. The data show rapid changes in MRAP transcription during stress or ACTH exposure. This effect is mediate by ACTH in a cyclic AMP and PKA-dependent manner. The rapid and transient increases in MRAP transcription during adrenal fasciculata activation suggest that regulation of MRAP levels plays an important role controlling ACTH receptor activity in the adrenals. Sources of Research Support: Intramural Research program, NICHD. Nothing to Disclose: YL, AGC, VH, FS, SL, GA