Lactating heifers were treated for 4 weeks with recombinant bovine growth hormone (bGH, n = 9) or were untreated (n = 9). In addition, two mammary glands of each heifer were milked four times daily rather than the normal twice daily for the same 4 weeks, and for the following 2 weeks. Over the 4 weeks, milk yield was increased 12-8% by bGH, 14-0% by frequent milking and 28'5% by the combined treatment. The effect of bGH as administered here was slower in onset than that of frequency, but eventually produced a higher peak yield. ANOVA revealed significant effects of each stimulus independently and an additive, but not synergic effect of the combined treatment. The effect of the combined treatment tended to persist beyond the end of treatment; most of this response was related to the milking frequency component rather than the bGH. Mammary differentiation was assessed in biopsies of mammary tissue obtained prior to and at the end of treatment. Mammary enzyme activities (expressed on a per cell basis) indicated minimal differentiative response to either treatment, but synthesis rates for lactose and casein determined in vitro were increased by bGH treatment. Histological examination revealed a stimulatory effect of milking frequency on epithelial cell size. The results indicate that these two galactopoietic stimuli operate through independent mechanisms, and neither stimulus alone is sufficient to maximize milk yield in dairy heifers.The nature of the galactopoietic stimulus provided by bovine growth hormone (bGH) has been investigated intensively in recent years. The major effect is a homeorhetic repartitioning of nutrients towards the mammary gland (Bauman et al. 1985), although there is also evidence of cardiovascular adaptation (Davis et al. 1988) and possibly an indirect effect of bGH on mammary tissue itself, mediated through IGF-1 (Prosser et al. 1990). Nevertheless, the balance of published information shows that bGH's systemic action outweighs any localized effect it has on the gland. Milk yield is also responsive to milking frequency; more frequent milking increases milk yield and less frequent milking reduces it, and in this case the physiological | Present address: