The quantitative light microscope radioautographic approach was utilized to locate specific binding sites for blood-borne 125I-labeled ACTH1-24 in the rat brain. Specific binding occurred exclusively in the external zone of the median eminence, adjacent to the primary portal plexus, and to a lesser extent in the ventral arcuate nucleus. This localization may indicate that the hypophysiotropic median eminence is a new target site for circulating adrenocorticotropin. Alternatively, ACTH binding sites in the median eminence-arcuate region may provide the hypothalamus with a hormone-selective capacity to concentrate pituitary ACTH for possible retrograde transport and redistribution within the central nervous system.