Sodium-coupled amino acid and sugar transport byNecturus small intestine

Summary Necturus small intestine actively absorbs sugars and amino acids by Na-coupled mechanisms that result in increases in the transepithelial electrical potential difference ( ^ms ) and the short-circuit current (I _sc) which can be attributed entirely to an increase in the rate of active Na absorption. Studies employing conventional microelectrodes indicate that the addition of alanine or galactose to the mucosal solution is followed by a biphasic response. Initially, there is a rapid depolarization of the electrical potential difference across the apical membrane ( ^ms ) which reverses polarity (i.e. cell interior becomes positive with respect to the mucosal solution) and a marked decrease in the ratio of the effective resistance of the mucosal membrane to that of the serosal membrane (R ^m /R ^s ); these events do not appear to be dependent on the availability of metabolic energy. These initial, rapid events are followed by a slow increase in (R ^m /R ^s ) toward control values which is paralleled by a repolarization of ^ms and increases in ^ms andI _sc; this slow series of events is dependent upon the availability of metabolic energy.The results of these studies indicate that: (i) the Na-coupled mechanisms that mediate the entry of sugars and amino acids across the apical membrane are rheogenic (conductive) and result in a decrease inR ^m and a depolarization of ^ms ; and (ii) the subsequent increase in (R ^m /R ^s ) and repolarization of ^ms are the results of a decrease inR ^s which is associated with an increase in the activity of the Na pump at the basolateral membrane.The physiologic implications of these findings are discussed and an equivalent electrical circuit model for rheogenic Na-coupled solute transport processes is analyzed.     

Modification of amino acid and sugar transport in uncoupler-adaptedEuglena gracilis

Uncoupler-adaptedEuglena gracilis have a greatly impaired capacity to take up and incorporate some exogenously supplied amino acids and sugars. The degree of inhibition varies widely from >90% in the case of valine or glucose to none in the case of histidine. The inhibition is due to a decreased activity of the transport mechanism itself and is not due to either a lesion in the control mechanism for endogenous amino acid or sugar synthesis nor to a direct inhibition of the transport mechanism by uncouplers.No preferential labeling of mitochondrial membranes by [¹⁴C]amino acids occurs during the process of adaptation, a time when no cell division occurs. Apparently, during the long time required for adaptation, there occurs no major modification of mitochondrial proteins which could explain the subsequent resistance to uncouplers.Contribution from the Department of Biochemistry, School of Agriculture and Life Sciences and School of Physical and Mathematical Sciences. Paper No. 5179 of the Journal Series of the North Carolina Agricultural Experiment Station, Raleigh, North Carolina 27607.     

Small intestinal sugar and amino acid transport in semistarvation.

The effect of semistarvation on small intestinal transport of D-glucose, L-valine, and NaCl was studied in an in vitro system of isolated rat brush border membrane vesicles. Whereas semistarvation enhanced the transport rate for L-valine by 19-29%, there was no change in D-glucose transport. When energy in the form of a NaSCN gradient was supplied to the membrane vesicles prepared from semistarved animals, L-valine was concentrated to a greater extent than those from well-fed animals. Strain differences were observed in the manner semistarvation affected NaCl transport across the brush border membrane. Semistarvation increased the NaCl transport rate by a factor of 3.5 in one rat strain and not at all in another. These results provide a partial explanation for the cellular basis of elevated neutral amino acid absorption by the small intestine in semistarvation.     

Effect of phenethylbiguanide on sugar and amino acid transport in rabbit ileum

Phenethylbiguanide has been shown to be an inhibitor of sugar and amino acid uptake in both $\text{in vivo}$ and $\text{in vitro}$ conditions. This action could be due to a competition for sodium sites on the sugar and amino acid carrier molecules. The effects of phenethylbiguanide on $\text{in vitro}$ intestinal preparations indicate that this compound has a time-dependent effect, it is most effective when placed on the mucosal surface but is also effective on the serosal surface. Furthermore, competition studies indicate that it is a competitive inhibitor of sugar uptake and a non-competitive inhibitor of amino acid uptake. These results are consistent with the differences in the mechanism of coupled transport between sugars and amino acids, but, do not substantiate the idea that phenethylbiguanide competes for the sodium site on the ternary carrier.     

On the mechanism of sugar and amino acid interaction in intestinal transport.

The influence of amino acids on D-glucose transport was studied in isolated vesicles of brush border membrane from rat small intestine. It is demonstrated that: (a) Uptake of D-glucose by the membranes is inhibited by simultaneous flow of L- and D-alanine into the vesicles. (b) Addition of L-alanine to membranes pre-equilibrated with D-glucose causes efflux of this sugar. (c) The influence of amino acids on D-glucose is dependent on the presence of Na+. (d) The ionophorous agents monactin and valinomycin are able to prevent the transport interaction of D-glucose and amino acids. Monactin is effective in the presence of Na+ without further addition of other cations, while valinomycin is effective only with added K+, in accordance with the known specificity of these antibiotics. (e) The inhibitory effect increases with L-alanine concentration up to about 50 mM after which it levels off. The experiments provide evident that the Na+-dependent sugar and amino acid fluxes across the brush border membrane are coupled electrically.     

Neutral amino acid transport in an androgen-dependent epithelium

• 1.1. Transport of neutral amino acids by the isolated seminal vesicle epithelium of normal and gonadectomized guinea pigs has been investigated by measurement of the uptake of 2-amino[1-¹⁴C]-isobutyric acid and 2-methylamino[1-¹⁴C]isobutyric acid.
• 2.2. The V_max for Na-dependent and -independent transport of both amino acids was reduced by gonadectomy but the general transport characteristics appeared to be unchanged by this treatment.
• 3.3. The most likely explanation of the decreased transport is the loss of transporter molecules associated with the tissue regression that follows rapidly on gonadectomy.

     

Calcium movements accompanying the transport of sugar or amino acid by rabbit enterocytes

Isolated rabbit enterocytes can be loaded with radioactive calcium most of which is presumably in intracellular stores. In the presence of sugar or amino acid there is a transient loss of calcium, followed by replenishment. It is suggested that this movement might be related in the signalling leading to increased potassium permeability observed in enterocytes transporting sugar and amino acids.     

pH-dependent heterogeneity of acidic amino acid transport in rabbit jejunal brush border membrane vesicles.

Initial rates of Na(+)-dependent L-glutamic and D-aspartic acid uptake were determined at various substrate concentrations using a fast sampling, rapid filtration apparatus, and the resulting data were analyzed by nonlinear computer fitting to various transport models. At pH 6.0, L-glutamic acid transport was best accounted for by the presence of both high (Km = 61 microM) and low (Km = 7.0 mM) affinity pathways, whereas D-aspartic acid transport was restricted to a single high affinity route (Km = 80 microM). Excess D-aspartic acid and L-phenylalanine served to isolate L-glutamic acid flux through the remaining low and high affinity systems, respectively. Inhibition studies of other amino acids and analogs allowed us to identify the high affinity pathway as the X-AG system and the low affinity one as the intestinal NBB system. The pH dependences of the high and low affinity pathways of L-glutamic acid transport also allowed us to establish some relationship between the NBB and the more classical ASC system. Finally, these studies also revealed a heterotropic activation of the intestinal X-AG transport system by all neutral amino acids but glycine through an apparent activation of Vmax.     

Urea and amino acid transport by an isolated human placenta]

A method of bilateral perfusion of the isolated human placenta was used to study the urea transport from the fetal placental stream into the maternal one, and of amino acid transport in the opposite direction. Experiments demonstrated that the method provided a sufficiently full perfusion of the intervillous space and offered possibilities for studying the placental transport. With equal amino nitrogen concentration in both circulations, its content in the fetal stream increased during the experiment. This elevation was more expressed when amino acid was added to the maternal circulation. The idea of amino acid "secretion" by the trophoblast cell elements into the fetal circulation was confirmed by the above experiments.     

Monoiodo-d-tyrosine,an artificial amino acid radiopharmaceutical for selective measurement of membrane amino acid transport in the pancreas

Using ¹¹C-labeled natural amino acids, the functional diagnosis of tissue metabolism has been actively studied. Our interest has been focused on developing a clinically available ¹²³I-labeled artificial amino acid with a single metabolic function. For this study, [¹²³I]3-iodo-d-tyrosine ([¹²³I]d-MIT) was selected. In vitro and in vivo studies using ¹²⁵I-labeled d-MIT indicated that it showed a high pancreatic accumulation, selective affinity for membrane active transport systems, and was stable against enzymatic deiodination. A canine scintigraphic study using ¹²³I-labeled d-MIT and kinetic analysis showed that it behaved as an “artificial amino acid” radiopharmaceutical with selective membrane amino acid transport affinity in the pancreas.