Abstract Cellular penetration of α-aminoisobutyric acid and of 2-deoxyglucose was measured in sex accesory tissues of the castrated male rat 3 to 18 h after a single intravenous injection of testosterone. The hormone had no significant effect on water content or extracellular (inulin) volumes over 18 h in ventral prostate, seminal vesicle and levator ani muscle. α-Aminoisobutyric acid distribution in prostate and seminal vesicle was increased significantly 6 (but not 3) h after testosterone injection and the tissues were concentrating α-aminoisobutyric acid against a concentration gradient by the 18th h; levator ani was not affected until the 12th h. The hormone did not stimulate uptake of 2-deoxyglucose in vivo except in levator ani at h 18. Pretreatment with testosterone for 12 h caused significant increases in α-aminoisobutyric acid uptake in all tissues over a 60-min incubation period; 2-deoxyglucose vitro also was taken up in larger amounts, and against a gradient, in all hormone-treated tissues except levator ani. In each tissue in vitro, the increased α-aminoisobutyric acid distribution due to testosterone was reduced to or below controls by low temperature (20°; Q 10 of 1.9 to 2.0), anoxia or the absence of Na, and by the presence of dinitrophenol, ouabain or alanine. The accumulation of 2-deoxyglucose in vitro after hormone pretreatment was depressed below untreated controls in the presence of glucose or phlorizin. From these experiments it may be concluded that within 6 or 12 h of injection, testosterone increased cellular penetration of a model amino acid and a hexose sugar in male sex accessory organs. The process requires energy, is sodium dependent and apparently utilizes carrier systems in common with natural substrates. The temporal and causal relationships between these events and hormone-stimulated metabolism are discussed.