Characterization of methylaminoisobutyric acid transport by system A in rat mammary gland

During lactation, the mammary gland has a large demand for amino acids for the synthesis of milk proteins and fatty acids, Arteriovenous differences i n amino acids across the mammary gland show an elevated uptake of small neu tral amino acids that are mainly transported via system A, The purpose of t his study was to characterize the transport of methylaminoisobutyric acid ( MeAIB), an amino acid analog used to model transport by system A in lactati ng rat mammary gland explants. MeAIB accumulation in mammary gland cells in creased steadily, and after 3 hours of incubation, the intracellular concen tration of the analog was 8-fold higher than the concentration in the mediu m. MeAIB transport into mammary gland explants showed a K-m of 3.3 +/- 0.4 mmol/L and a maximal velocity (Vmax) of 555 +/- 23 pmol/mu L intracellular fluid (ICF) . min, indicating a system with high capacity but low affinity for its substrate. MeAIB transport into mammary tissue depended highly on N a+, and the uptake was inhibited by addition of natural and analog small ne utral amino acids. Cationic, anionic, and large neutral amino acids did not reduce MeAIB transport into mammary gland explants, Preincubation of mamma ry gland explants in an amino acid-free medium stimulated MeAIB transport, suggesting an adaptive regulation. The addition of an equimolar mixture of alanine, glycine, and serine to the preincubation medium inhibited stimulat ion of MeAIB transport. Furthermore, stimulation of MeAIB uptake by amino a cid starvation was also prevented by the addition of actinomycin D, cyclohe ximide, tunicamycin, and colchicine, Dibutyryl cyclic adenosine monophospha te (cAMP) increased MeAIB uptake, whereas phorbol 12-myristate 13-acetate ( PMA) did not stimulate MeAIB transport. During the first postweaning days, kinetic analyses showed a decrease of 27% in the Vmax. Injection of rat lac tating mammary gland mRNA into Xenopus laevis oocytes induced expression of the MeAIB transport system; however, the induction was only 83% above back ground MeAIB uptake. The results of this study provide a partial explanatio n for the formation of the metabolic pool of small neutral amino acids in t he lactating mammary gland. Copyright (C) 2000 by W.B. Saunders Company.