Physiological studies by Setchell and others have described the existence of a blood-testis barrier (BTB) surrounding the seminiferous tubules of the mammalian testis. These studies were initiated to better define the role of the BTB with regard to the penetration of exogenous chemicals to male germ cells. The rete testis was cannulated in rats and fluid was collected. Test chemicals or drugs were usually administered by continuous i.v. infusion. Permeability of nonelectrolytes of various molecular sizes, acidic compounds with varying partition coefficients and pKalpha values, such as salicylic acid, barbiturates, and sulfonamides, across the BTB were studied. Permeability of nonelectrolytes was demonstrated to be dependent upon their molecular size, suggesting bulk flow through water-filled pores. On the other hand, permeability of acidic drugs with varying pKalpha values depended upon their partition coefficients. Transport of these chemicals from blood to seminferous tubules closely resembled their transport from blood to cerebrospinal fluid. It appears that the BTB is a complex multicellular system composed of membranes surrounding the semiferous tubules and the several layers of spermatogenic cells organized within the tubules, which restrict the permeability to the male germ cells of many foreign compounds. This must be borne in mind when extra-polating data from in vitro mutagenic test systems to man.