B.G. Munck

Amino acid transport by the small intestine of the rat. The existence and specificity of the transport mechanism of imino acids and its relation to the transport of glycine.

Abstract From experiments on sacs of the everted small intestine of the rat it was shown: 1. 1. That the small intestine of the rat possesses a separate transport mechanism for imino acids. 2. 2. That this transport mechanism is used by glycine, betaine, leucine, and alanine in addition to proline, hydroxyproline and sarcosine. 3. 3. That the demonstration of inhibition of the transport of glycine by imino acids is complicated by the affinity of glycine for two transport mechanisms; that of the neutral amino acids in addition to that of the imino acids. 4. 4. That this complication can be eliminated by competitive exclusion of glycine from the carrier of the neutral amino acids. It is suggested that the A-mediation of amino acid transport by the Ehrlich ascites tumor cell may be a manifestation of an imino acid transport mechanism in this cell.

Transport of imino acids and non-α-amino acids across the brush-border membrane of the rabbit ileum

SummaryThe transport of β-alanine and MeAIB and their effects as inhibitors of the transport of alanine, leucine and lysine across the brush-border membrane of the intact epithelium from the rabbits distal ileum has been examined. Two separate transport systems have been characterized: 1) A sodium-dependent, β-alanine-accepting system, which is a high-affinity transport system for α-amino-monocarboxylic acids (neutral a.a.) and for cationic a.a., accepts non-α-amino acids as well as non-α-imino acids, is moderately stereospecific, and for which the affinity of a neutral a.a. is greatly reduced by N-methylation. 2) A sodium-dependent transport system for imino acids, which is inaccessible to cationic amino acids and non-α-amino acids but accepts cyclic, non-α-imino acids, is moderately stereospecific, and for which neutral a.a. have much lower affinities than their N-methylated derivatives. On the basis of the observations of this and the preceding paper five transport systems for amino acids are ascribed to the rabbit ileum. Some discrepancies between the present results and those obtained with brush-border membrane microvesicles from the rabbit small intestine are discussed.

Transport of neutral and cationic amino acids across the brush-border membrane of the rabbit ileum

SummaryThe transport of sugars and amino acids across the brush-border membrane of the distal rabbit ileum has been studied. The kinetics of the transport of glucose demonstrated that the data obtained with the present technique are less distorted by unstirred layers than those obtained with the same technique adapted to the use of magnetic stirring. The role of depolarization of the electrical potential difference across the brush-border membrane in mutual inhibition between different classes of amino acids was estimated by measurements of the effects of high concentrations of alanine and lysine on the transport of galactose. It was found that this role would be insignificant in the present study. By measurements of the transport of alanine, leucine and lysine and the inhibitory interactions between these amino acids the function of three transport systems has been delineated. The transport of lysine is resolved in a high- and a low-affinity contribution. At 140mm sodium these transport systems may also function as respectively high- and low-affinity contributors to the transport of neutral amino acids. At 0mm sodium the high-affinity system remains a high-affinity system for cationic and neutral amino acids with reduced capacity especially for the neutral amino acids. At 0mm sodium the low-affinity systems affinity for lysine is reduced and it is inaccessible to neutral amino acids. In addition to the two systems for lysine transport the existence of a lysine-resistant, sodium-dependent, high-affinity system for the transport of neutral amino acids has been confirmed. It seems unlikely that the distal ileum is equipped with a low-affinity, sodium-independent system for the transport of neutral amino acids.

Amino acid transport by the small intestine of the rat. An evaluation of the possibility of vitamin B6 as an intestinal amino acid carrier.

Abstract The transintestinal amino acid transport by the averted sac from the small intestine of the rat was tested for vitamin B 6 dependence by means of the vitamin B 6 antagonist 4-deoxypyridoxine. The transports of amino acids representing the three known amino acid-transport systems were inhibited almost equally, whereas the transport of glucose was unaffected by the vitamin antagonist. This inhibition of the amino acid transport was found only when the intestinal sacs were preincubated with the vitamin antagonist. On the assumption that the three amino acid-transport systems are operated by three separate carriers it is concluded, that vitamin B 6 cannot be an amino acid carrier of the small intestinal epithelium; rather the vitamin may be necessary to some metabolic process common to the three transport systems. This process could be one by which the affinity of the carriers is diminished inside the cells, whereby an accumulative transport of the amino acids is permitted. This effect of vitamin B 6 could be expressed as an inhibition of a mediated amino acid flux from the serosal to the mucosal fluid.

Amino acid transport by the small intestine of the rat on the counterflow phenomenon as a cause of the accelerating effect of leucine on the transintestinal transport of diamino acids

Abstract The accelerating effect of leucine on the transintestinal transport of lysine has been studied. The relationship of the transport of lysine to that of leucine was such as would be expected if the accelerating effect of leucine was a result of the combined function of the transport mechanisms of the neutral and the diamino acids. Of the two transport mechanisms that of the neutral amino acids created and maintained, on the serosal side of the transport mechanisms, a concentration of leucine sufficient to exert a counterflow effect on the transport of lysine by the transport mechanism of the diamino acids. The transintestinal transport of arginine and ornithine was accelerated by leucine to the same extent as that of lysine.

Effects of pH changes on systems ASC and B in rabbit ileum.

Influx ofd-aspartate (d-Asp),l-glutamate (l-Glu), and serine (Ser) across the brush-border membrane of the intact mucosa from rabbit ileum has been examined. l-Glu influx is chloride independent and completely sodium dependent.d-Asp andl-Glu share a transport system with a maximum transport rate of 1 μmol ⋅ cm-2 ⋅ h-1and an apparent affinity constant ( K ½) of ∼0.3 mM. The function of this transport system is pH insensitive between pH 5.65 and 8.2, and bipolar amino acids do not affect the way in which the transport system handlesd-Asp andl-Glu. The characteristics of this transport system match those of system[Formula: see text].l-Glu and Ser share a transporter for which the inhibitor constant ( K i) ofl-Glu against Ser decreases from 54 to 10 mM when pH is reduced from 7.2 to 5.65, while the maximum rate of transport remains unaffected at ∼10 μmol ⋅ cm-2 ⋅ h-1. The K i values (5 mM) of Ser against l-Glu influx and the l-Glu-sensitive contribution to Ser influx (0.8 μmol ⋅ cm-2 ⋅ h-1at 1 mM Ser) are the same at both pH values. Thel-Glu-sensitive transport of Ser together with the contribution of system bo,+ account for ∼50% of Ser influx at pH 7.2. The remaining 50% can be ascribed to system B. Transport of Ser by system B is reduced by >95% at pH 5.65. At pH 7.2 K i of Ser against transport of leucine (Leu) by system B is 18 mM and K i of Leu against transport of Ser is 1.7 mM. The low-affinity transport ofl-Glu and thel-Glu-sensitive transport of Ser are performed by an equivalent of system ASC. Supplementary experiments using the jejunum confirm the validity of these results for a major portion of the rabbit small intestine.Influx of D-aspartate (D-Asp), L-glutamate (L-Glu), and serine (Ser) across the brush-border membrane of the intact mucosa from rabbit ileum has been examined. L-Glu influx is chloride independent and completely sodium dependent. D-Asp and L-Glu share a transport system with a maximum transport rate of 1 micromol. cm-2. h-1 and an apparent affinity constant (K1/2) of approximately 0.3 mM. The function of this transport system is pH insensitive between pH 5.65 and 8.2, and bipolar amino acids do not affect the way in which the transport system handles D-Asp and L-Glu. The characteristics of this transport system match those of system X-AG. L-Glu and Ser share a transporter for which the inhibitor constant (Ki) of L-Glu against Ser decreases from 54 to 10 mM when pH is reduced from 7.2 to 5.65, while the maximum rate of transport remains unaffected at approximately 10 micromol. cm-2. h-1. The Ki values (5 mM) of Ser against L-Glu influx and the L-Glu-sensitive contribution to Ser influx (0.8 micromol. cm-2. h-1 at 1 mM Ser) are the same at both pH values. The L-Glu-sensitive transport of Ser together with the contribution of system bo,+ account for approximately 50% of Ser influx at pH 7.2. The remaining 50% can be ascribed to system B. Transport of Ser by system B is reduced by >95% at pH 5.65. At pH 7. 2 Ki of Ser against transport of leucine (Leu) by system B is 18 mM and Ki of Leu against transport of Ser is 1.7 mM. The low-affinity transport of L-Glu and the L-Glu-sensitive transport of Ser are performed by an equivalent of system ASC. Supplementary experiments using the jejunum confirm the validity of these results for a major portion of the rabbit small intestine.

Lysine transport across the small intestine. Stimulating and inhibitory effects of neutral amino acids

SummaryThe ordinary aliphatic, neutral amino acids and phenylalanine have been examined for cis-inhibition of influx of alanine (Jmcala) and lysine (Jmclys) and trans-stimulation ofJmclys across the brush border membrane of rat small intestines: and their effects on the unidirectional mucosa-to-serosa flux (Jmslys) across the short circuited intestine have been studied. The effects of alanine, α-amino-n-butyric acid, leucine, and methionine on the steady-state epithelial uptake of lysine [Lys]c have also been measured. In addition the trans-effects of alanine and leucine have been examined for sodium-dependence, and alanine was tested as trans-stimulator of influx of galactose across the brush border membrane (Jmcgal).All the neutral amino acids were found to be competitive cis-inhibitors ofJmclys, and all, except isoleucine, were trans-stimulators ofJmclys. The magnitude of the trans-effect was unrelated to the efficiency of the amino acid as cis-inhibitor. As illustrated by alanine, the trans-effects are probably completely sodium-dependent. Alanine was also effective as trans-stimulator ofJmcgal. With respect to effects on [Lys]c andJmslys the neutral amino acids fall into two groups: One which reduces [Lys]c and stimulatesJmslys, and one which increases [Lys]c and relatively inhibitsJmslys. These effects are not correlated with the affinities of the neutral amino acids for the two carriers involved.It is proposed that the trans-effects onJmclys are induced by an electrogenic, sodium-coupled efflux of the neutral amino acid across the brush border membrane, that the stimulation ofJmslys is brought about by a selective stimulation (of unknown nature) of efflux of lysine across the basolateral membrane (Jcslys), assisted by competitive inhibition of lysine efflux across the brush border membrane (Jcmlys), and that the amino acids which do not stimulateJcmlys increase [Lys]c by competitively inhibitingJcslys andJcmlys.The inhibitory effect of the neutral amino acids onJmclys support the view that the carrier of basic amino acids serves as a second carrier of these amino acids.

Variation in amino acid transport along the rabbit small intestine. Mutual jejunal carriers of leucine and lysine

The jejuno-ileal variation of amino and imino acid transport across the brush-border membrane of intact rabbit small intestine was studied. For the amino acids tested--beta-alanine, leucine, lysine, MeAIB, proline--and for D-glucose, the rates of transport at constant concentrations increase from very low values in the proximal jejunum to maximum values in the most distal 30 cm of the ileum. The apparent affinity constant for jejunal taurine transport is identical to that of the distal ileum, while the jejunal transport capacity is less than half. In the jejunum, as in the distal ileum, leucine and lysine share both sodium-dependent and sodium-independent carriers. Approx. 50% of the quantitative difference in transport capacity is accounted for by the absence of the beta-alanine carrier in the jejunum. These data indicate that the gradients of transport along the small intestine reflect gradients of transport capacities rather than affinities. In comparison with hamster, man and rat, the rabbit seems unique with respect to the location of transport maximum and the steepness of the gradient along the intestine.

CHLORIDE-DEPENDENCE OF AMINO ACID TRANSPORT IN RABBIT ILEUM

Chloride-dependence of influx across the brush-border membrane of distal rabbit ileum was examined for beta-alanine, 2-methylaminoisobutyric acid (MeAIB), leucine, lysine, proline and D-glucose. Influx of leucine at 2 mM and of D-glucose at 0.5 mM was chloride-independent indicating that substitution of isethionate for chloride has no unspecific effect on sodium gradient driven transport processes. In contrast influx of beta-alanine and MeAIB was totally dependent on the presence of chloride ions. In the absence of chloride, proline transport was reduced to 20% of its control level. This remaining transport can be accounted for by the function of the carrier of alpha-amino-monocarboxylic acids. Transport of leucine at 0.1 mM was reduced by absence of chloride. This is in accordance with the observation of leucine transport by the beta-alanine carrier. The kinetics of chloride and sodium activation of transport of MeAIB were examined at 1 mM MeAIB. Chloride activation was characterized by a Hill coefficient of 1 and a K1/2 of 23.5 mM, and sodium activation by a Hill coefficient of 2 and a K1/2 of 51 mM. Thus cotransport of chloride with an imino acid would be compatible with the known rheogenic nature of this transport. This study adds the imino acid carrier and the beta-alanine carrier to the group of chloride-dependent, epithelial amino acid transport systems.

Transport of l;lycine and lysine on the chloride-dependent β-alanine (B0,+) carrier in rabbit small intestine

Transport of glycine, lysine and β-alanine in rabbit, guinea pig and rat small intestine has been examined by measurements of the unidirectional influx across the brush border membrane of the intact epithelium. In rabbit distal ileum the chloride-dependent fraction of glycine transport, and all sodium- and chloride-dependent lysine transport is carried on the β-alanine carrier. Lysine eliminates all saturable, sodium-independent transport of glycine. The saturable, sodium-dependent, and lysine resistant influx of glycine is characterized by a K12Gly of 60 mM. Glycine transport in the mid intestine of the guinea pig is chloride-independent and in the rat only a minute fraction may be chloride-dependent. These species do not possess an equivalent of the rabbit β-alanine carrier. In conclusion, glycine transport in rabbit distal ileurn is by the sodium-dependent carrier of neutral amino acids, by the sodium-independent lysine carrier, and by the sodium- and chloride-dependent /β-alanine carrier which closely resembles the B0,+ carrier described in mouse blastocysts. All sodium dependent lysine transport in rabbit distal ileum is by the chloride- and sodium-dependent /gb-alanine carrier. It is proposed that the (β-alanine carrier in rabbit distal ileum be renamed the B0,+ carrier.