The entire second thoracic mammary glands of 4-week-old BALB/c female mice primed with oestradiol plus progesterone were cultivate in organ culture medium containing the "growth-promoting" hormone combinations: insulin, prolactin, growth hormone, oestradiol, progesterone and aldosterone or insulin, prolactin and aldosterone. Full lobulo-alveolar development was induced after 5-6 days of incubation and could be maintained for 15-16 days in organ culture in medium containing either hormone combination. After the initial 5-6 days in the "growth-promoting" medium, subsequent cultivation of the glands in a medium with the "lactogenic hormones", insulin, prolactin plus cortisol, led to accumulation of "milk-like" secretory material in the ductal and alveolar lumina. Incubation of the lobulo-alveolar gland in medium with insulin alone for 7-9 days resulted in complete regression of the alveoli leaving only a ductal parenchyma. Incubation in insulin, prolactin, growth hormone or insulin plus the steriod hormones for 7-9 days led to considerable alveolar degeneration without a complete regression. The results indicate that both pituitary and steroid hormones are essential for development and maintenance of mammary alveoli; insulin can only sustain the basal ductal structure.