The transfer of lipoprotein-bound cholesterol into adrenal cells was examined. Adrenal glands from unstimulated or corticotropin stimulated hypophysectomized rats were incubated with high density lipoprotein (HDL) or low density lipoprotein LDL containing radiolabeled cholesterol. The rate of transfer of labeled cholesterol from HDL into the glands was two to three times greater than from LDL. Corticotropin stimulation increased the transfer of cholesterol from HDL but not LDL. The effects of corticotropin were not dependent on subsequent cholesterol utilization for steroidogenesis. The process of cholesterol transfer from HDL was linear with time over 2 hr at 37 degrees and greatly reduced at 4 degrees. In addition, the transfer process became saturated above an HDL cholesterol concentration of 900 mug/ml. About 25% of the labeled adrenal cholesterol arising from HDL was recovered within the mitochondria. The labeled cholesterol within isolated mitochondria could undergo mitochondrial conversion to pregnenolone. Finally, the delipidated HDL apolipoproteins, apoA-I and apoA-II, when added to incubations containing less than saturating concentrations of HDL, stimulated transfer of labeled cholesterol from HDL to adrenal cells. These studies suggest that rat adrenal tissue possesses an HDL specific hormonally-responsive mechanism for accumulating extracellular cholesterol and that apoA-I and apoA-II have a significant function in the uptake process.