Volume-activated amino acid efflux from term human placental tissue: stimulation of efflux via a pathway sensitive to anion transport inhibitors.

The effect of a hyposmotic challenge and hence cell-swelling upon the efflux of a variety of solutes from isolated human placental tissue has been examined. A hyposmotic shock increased the fractional release of taurine, the most abundant free amino acid in placental tissue, via a pathway sensitive to niflumic acid, DIDS (4,4'-Diisothiocyanatostilbene-2',2'-disulphonic acid,) NPPB (5-Nitro-2(3-phenylpropylamino)benzoic acid) and DIOA (R(+)[2-n-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden -5-y) oxy] acetic acid). In contrast, tamoxifen was without effect. The cell-swelling induced efflux of taurine was attenuated (40 per cent) by replacing external Cl- with NO3-. The efflux of glutamic acid was also markedly increased by a hyposmotic challenge. Niflumic acid inhibited both basal and volume-activated glutamic acid efflux. A hyposmotic shock also increased alpha-aminoisobutyric acid efflux but not that of 3-O-methylglucose and SO4(2)-. The results suggest that the human placenta can respond to cell-swelling by releasing organic osmolytes such as amino acids via a pathway which is sensitive to anion transport inhibitors. However, it appears that the volume-activated amino acid transport system is independent from the placental anion-exchange pathways. The efflux of these compounds may act with K+ and Cl- efflux to effect a regulatory volume decrease in placental tissue. In addition, volume-activated transport may play a role in transplacental amino acid transfer.

The effect of a hyposmotic shock on amino acid efflux from lactating rat mammary tissue: stimulation of taurine and glycine efflux via a pathway distinct from anion exchange and volume-activated anion channels.

We have examined the effect of a hyposmotic shock, and thus cell swelling, upon the efflux of amino acids, SO4(2-) and I- from lactating mammary tissue. A hyposmotic challenge increased the efflux of taurine and glycine via a 4,4'-diisothiocyanatostilbene-2,2'- disulphonic acid (DIDS)-sensitive pathway. It appears that these amino acids do not exit via an anion-exchange mechanism following cell swelling because sulphate efflux, which uses a DIDS-sensitive exchange mechanism, was unaffected. The hyposmotic-induced efflux of taurine was not dependent upon the Na+ gradient and was not influenced by the nature of the anion in the incubation medium. In addition, taurine efflux was stimulated by incubating mammary tissue in an isosmotic medium that contained urea, suggesting that cell swelling is the stimulating factor rather than a decrease in osmolality per se. The results suggest that mammary tissue uses taurine and glycine as a means of regulating cell volume following swelling. In contrast, the efflux of glutamic acid, alanine and alpha-aminoisobutyric acid was unaffected by a hyposmotic challenge. Similarly, the efflux of I- was unaffected by such a challenge. The results suggest that volume-activated amino acid transport in lactating rat mammary tissue is distinct from volume-regulated anion channels.

Lysine transport in lactating rat mammary tissue: evidence for an interaction between cationic and neutral amino acids.

The transport of lysine by the lactating rat mammary gland has been examined to determine whether there is an interaction between cationic and neutral amino acids. Lysine uptake was time dependent and unaffected by replacing Na+ with choline. In the presence of Na+, lysine influx was inhibited by cationic amino acids (arginine, homoarginine, ornithine and lysine) and by a range of neutral amino acids (methionine, glutamine, leucine, phenylalanine, alanine, asparagine, alpha-aminoisobutyric acid (AIB), 2-aminobicyclo [2,2,1] heptane-2-carboxylic acid (BCH), proline and tryptophan). Leucine and glutamine also inhibited lysine influx in the absence of Na+ but phenylalanine and proline did not. Lysine efflux from mammary tissue was trans-accelerated by various cationic amino acids (lysine, arginine, homoarginine and ornithine). In addition, leucine and glutamine were capable of trans-stimulating lysine efflux in the presence and absence of Na+. It appears that cationic and neutral amino acids stimulated lysine efflux at a single locus.

High affinity (Na(+) + Cl-)-dependent taurine transport by lactating mammary tissue.

The transport of taurine by lactating rat mammary tissue has been examined. Taurine uptake was found to be dependent upon the presence of extracellular Na+ and Cl-, which is consistent with (Na(+) + Cl(-) + taurine) cotransport. The Ka and Vmax of taurine influx were respectively 43 microM and 37.5 mumol/kg cell water per 15 min. It is apparent that the mechanism responsible for taurine uptake is highly selective for beta-amino acids. Taurine efflux consisted of a fast extracellular component and a slow membrane-limited component. The slow component was relatively insensitive to temperature, suggesting that it may represent simple diffusion. Mammary tissue was found to contain a high level of intracellular taurine: 7.29-7.44 mmol/kg cell water. We suggest that taurine is taken up and concentrated by the mammary gland by a (Na(+) + Cl(-) + taurine) cotransport mechanism situated in the blood-facing aspect of the secretory epithelium and that a low outward permeability to taurine allows a high intra-to-extracellular concentration gradient to be maintained. Milk taurine may be derived from taurine diffusing from the cell cytosol across the apical membrane.

Characteristics of alpha-aminoisobutyric acid transport by lactating rat mammary gland.

The transport of alpha-aminoisobutyric acid (AIB) by lactating rat mammary tissue has been examined. AIB uptake by mammary tissue was via both Na(+)-dependent and Na(+)-independent pathways. AIB uptake via the Na(+)-dependent pathway was inhibited by (methylamino)isobutyric acid (MeAIB) whereas AIB uptake via the Na(+)-independent pathway was blocked by 2-aminobicyclo[2,2,1]-heptane-2-carboxylic acid (BCH). A small fraction of AIB influx persisted in the presence of both MeAIB and BCH. The Na(+)-independent moiety of AIB uptake was strongly inhibited by phenylalanine, tryptophan, leucine, isoleucine and methionine. AIB efflux from mammary tissue slices was found to be both Na(+)-dependent and Na(+)-independent. Trans-stimulation of AIB efflux by other amino acids was not observed; in contrast, external phenylalanine, tryptophan and leucine inhibited AIB efflux. The results are largely consistent with the presence of systems A and L in lactating rat mammary tissue. However, the Na(+)-independent fraction of AIB transport may represent transport via a tissue specific form of system L.

Stimulation of taurine efflux from human placental tissue by a hypoosmotic challenge.

The fractional release of taurine from preloaded human placental tissue fragments was stimulated by a hypoosmotic challenge. This stimulation could not be attributed to a change in the Na+ gradient or the ionic strength of the medium. We suggest that taurine fluxes may play a role in placental cell volume regulation.

K+ (Rb+) transport by a mammary secretory cell apical membrane fraction isolated from goats' milk.

The transport of K+ (Rb+) by cell fragments isolated from goats' milk, which are believed to be pieces of mammary epithelial cell cytoplasm encapsulated by apical membrane, has been studied. K+ (Rb+) accumulation was time dependent and was virtually abolished by 0.1% Triton X-100 which indicates that K+ (Rb+) uptake is into a membrane-limited space rather than by binding. Replacing medium Cl- with SCN- increased whereas gluconate replacement of Cl- decreased K+ (Rb+) influx suggesting that uptake is sensitive to changes in membrane potential. Na+, K+, Li+ and Rb+ ions added to the incubation medium reduced the rate of K+ (Rb+) accumulation. K+ (Rb+) uptake was markedly reduced by both barium ions and quinine; the inhibitory effects were not additive suggesting they are acting at a single locus. Increasing the pH of the medium stimulated the Ba2(+)-sensitive component of K+ (Rb+) influx. The results show that the transport properties of the goats' milk membrane fraction used here are similar to the apical aspect of the intact mammary epithelium. Therefore this membrane preparation may be a good source and model to study mammary secretory cell apical membrane ion transport.

An interaction between selenate and a sulfate transporter in the lactating rat mammary gland.

The efficacy of selenate and other divalent anions to stimulate the efflux of radiolabeled sulfate from lactating rat mammary tissue slices has been examined. Both selenate and sulfate markedly increased the fractional release of sulfate via a system that is temperature-sensitive and sensitive to the anion-exchange inhibitor DIDS. The effect of selenate on sulfate efflux was saturable with an apparent affinity constant of approximately 0.27 mM. In addition, molybdate and thiosulfate were also found to increase sulfate efflux from the trans-aspect. It is concluded that sulfate and selenate share a pathway for transport in the lactating rat mammary gland.

Efflux of chloride from lactating rat mammary tissue slices.

1. The efflux of chloride (using 36Cl) from lactating rat mammary tissue slices has been investigated. 2. Chloride efflux was found to be temperature dependent; lowering the temperature of the incubation medium reduced the fractional efflux. 3. The stilbene derivatives DIDS was without effect on the fractional release of Cl when studied at 20 degrees C. However, DIDS was found to attenuate the increase in efflux found upon transferring the tissue from a medium maintained at 4 degrees C to one at 20 degrees C. 4. The loop-diuretic furosemide, also reduced the temperature-sensitive portion of Cl efflux. 5. Chloride efflux was transiently increased when tissue slices were transferred from a medium containing gluconate as the principal anion to one containing Cl. 6. The results appear to confirm that mammary Cl transport is mediated via anion exchange and via (Na + K + Cl) cotransport.