Placental amino acid uptake. II. Tissue preincubation, fluid distribution, and mechanisms of regulation.

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Extract: Human placental villous tissue was incubated with the nonmetabolizable amino acid analog, α-aminoisobutyric acid (AIB). A previous investigation demonstrated that AIB uptake into intracellular fluid is markedly increased by preincubation of tissue in a balanced salt, glucose-containing medium. This increase in transport capacity with preincubation has now been found to be blocked completely by cycloheximide, dinitrophenol, or cyanide, which indicates that the process requires protein synthesis and aerobic metabolism. Inasmuch as increases in AIB uptake with preincubation were reported in other tissues to be regulated by amino acids in the medium, effects of amino acids were studied in the placenta. Preincubation of placental tissue with alanine did result in a transport activity which was lower than that of control preincubated tissue. However, two types of experiments indicated that this difference results from inhibition of transport activity by intracellular alanine accumulated during preincubation rather than from blockage of formation of new transport capacity during preincubation. (1) Much of the difference between control and alanine-preincubated tissue was eliminated by short additional preincubations in cycloheximide-containing, alanine-free medium; a treatment which should lower intracellular alanine concentration but not permit protein synthesis. (2) Preincubation with AIB permitted measurement of both intracellular concentration and transport activity. Addition of AIB to control preincubated tissue caused concentrating ability to fall as intracellular concentration increased. When AIB and control preincubated tissue were tested with the same intracellular AIB concentration, their transport activities were the same. The rate of increase in concentrating ability with preincubation was inversely related to the amount of tissue per volume of preincubation medium and was enhanced by changing the medium at intervals. Preincubation of new placental tissue in medium which had been used previously for preincubation resulted in a smaller increase in concentrating ability than did preincubation of the same tissue in fresh medium. These findings were not dependent on glucose concentration or pH. During preincubation, some as yet unidentified tissue substance is apparently released into the medium and inhibits the increase in transport activity. Speculation: Amino acids required by the fetus must be supplied by the placenta. In vitro incubation of placental tissue has identified and partially characterized two potential mechanisms for regulation of placental amino acid uptake. The first mechanism increases transport activity when tissue is preincubated in vitro before measurement of uptake. The second mechanism apparently decreases existing uptake activity when amino acids are present intracellularly in high concentrations. In vivo these mechanisms could be the basis for regulation of amino acid transfer from mother to fetus.