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1. Ouabain-sensitive 86Rb+ uptake in slices of lactating rabbit mammary gland significantly increased after 20 min or 1 hr of incubation with ovine prolactin (NIH-P-S12; 1 microgram/ml.). 2. Total K+ content of the tissue significantly increased at 20 min of incubation with prolactin. 3. Neither vasopressin nor oxytocin, at concentrations of 2,20 or 40 mui.u./ml., had a significant effect on ouabain-sensitive 86Rb+ uptake or total K+ of the tissue after 30 min or 1 hr of incubation. 4. Tissue slices incubated in cycloheximide at 10 micrograms/ml. for up to 260 min showed a reduction in ouabain-sensitive 86Rb+ uptake and total K+ content, with half-lives of 115 and 63 min, respectively. 5. No consistent in vitro effect of prolactin on (Na+ + K+)-activated ATPase activity in homogenates, crude microsomal fractions or NaI-activated membrane fractions from lactating rabbit mammary gland was found. 6. After 3 hr of incubation of tissue slices in the presence or absence of prolactin (5 micrograms/ml.), no significant differences in the number of [G-3H]ouabain molecules bound per cell (5.2 X 10(4) and 6.2 X 10(4), respectively) or in the dissociation constant (KD) for ouabain binding (5.4 X 10(-7) M and 5.9 X 10(-7) M, respectively) were observed. 7. Incubation of the tissue with cycloheximide (10 micrograms/ml.) for 1-6 hr caused an exponential decrease in [G-3H]ouabain binding with a half-life of 3 hr. 8. It is concluded that prolactin stimulates the activity of the (Na+ + K+)-activated ATPase in slices of lactating rabbit mammary gland within one hour but over this period does not affect the number of ouabain-binding sites, which are apparently turned over with a half-life of 1-3 hr.

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