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 in amino acids across the mammary gland show an elevated uptake of small neutral amino acids that are mainly transported via system A. The purpose of this study was to characterize the transport of methylaminoisobutyric acid (MeAIB), an amino acid analog used to model transport by system A in lactating rat mammary gland explants. MeAIB accumulation in mammary gland cells increased steadily, and after 3 hours of incubation, the intracellular concentration of the analog was 8-fold higher than the concentration in the medium. MeAIB transport into mammary gland explants showed a Km of 3.3 +/- 0.4 mmol/L and a maximal velocity (Vmax) of 555 +/- 23 pmol/microL intracellular fluid (ICF) x min, indicating a system with high capacity but low affinity for its substrate. MeAIB transport into mammary tissue depended highly on Na+, and the uptake was inhibited by addition of natural and analog small neutral amino acids. Cationic, anionic, and large neutral amino acids did not reduce MeAIB transport into mammary gland explants. Preincubation of mammary 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 stimulation of MeAIB transport. Furthermore, stimulation of MeAIB uptake by amino acid starvation was also prevented by the addition of actinomycin D, cycloheximide, tunicamycin, and colchicine. Dibutyryl cyclic adenosine monophosphate (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 lactating mammary gland mRNA into Xenopus laevis oocytes induced expression of the MeAIB transport system; however, the induction was only 83% above background MeAIB uptake. The results of this study provide a partial explanation for the formation of the metabolic pool of small neutral amino acids in the lactating mammary gland.
Departamento de Fisiología de la Nutrición, Instituto Nacional de la Nutrición Salvador Zubirán, México, DF.
A R
AR
Tovar
E
E
Avila
S
S
DeSantiago
N
N
Torres
D026905:Amino Acid Transport Systems; D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000502:physiology*; D005260:Female; D007774:Lactation; D008099:Liver / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012964:Sodium / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014982:Xenopus laevis; D015091:beta-Alanine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics
D026905:Amino Acid Transport Systems; D002352:Carrier Proteins; D015091:beta-Alanine; C012924:2,2-dimethyl-beta-alanine; D012964:Sodium; D013755:Tetradecanoylphorbol Acetate